阅读说明：本技术 一种人***瘤病毒18型l1蛋白的突变体 (Mutant of human papilloma virus 18 type L1 protein ) 是由 李少伟 宋硕 何茂洲 史晶洁 顾颖 夏宁邵 于 2019-06-04 设计创作，主要内容包括：本发明涉及一种突变的HPV18L1蛋白(或其变体),其编码序列和制备方法,以及包含其的病毒样颗粒,所述蛋白(或其变体)和病毒样颗粒能够诱发抗至少两个型别的HPV(例如,HPV18和HPV45,或者HPV18、HPV45和HPV59)的中和抗体,从而可用于预防所述至少两个型别的HPV感染以及由所述感染所导致的疾病例如宫颈癌和尖锐湿疣。本发明还涉及上述蛋白和病毒样颗粒用于制备药物组合物或疫苗的用途,所述药物组合物或疫苗可用于预防所述至少两个型别的HPV感染以及由所述感染所导致的疾病例如宫颈癌和尖锐湿疣。(The present invention relates to a mutated HPV18L1 protein (or a variant thereof), a coding sequence and a preparation method thereof, and virus-like particles comprising the same, which are capable of inducing neutralizing antibodies against at least two types of HPV (e.g., HPV18 and HPV45, or HPV18, HPV45 and HPV59), thereby being useful for preventing infection by the at least two types of HPV and diseases caused by the infection, such as cervical cancer and condyloma acuminata. The invention also relates to the use of the above proteins and virus-like particles for the preparation of a pharmaceutical composition or vaccine useful for the prevention of HPV infections of said at least two types and diseases caused by said infections, such as cervical cancer and condyloma acuminata.)

1. A mutant HPV18L1 protein or variant thereof wherein the mutant HPV18L1 protein has the following mutations compared to the wild-type HPV18L1 protein:

(1) The N-terminal is truncated by 40-80 amino acids, such as 40-50, 45-70, 50-70, 55-65, 60-70, 65-75, 60-80 or 70-80 amino acids; and

(2) (a) the amino acid residues at positions 235-243 of the L1 protein of wild-type HPV18 are replaced by the amino acid residues at the corresponding positions of the L1 protein of wild-type HPV of the second type; or

(b) The amino acid residues at the 327-346 position of the wild-type HPV18L1 protein are replaced by the amino acid residues at the corresponding positions of the L1 protein of the wild-type HPV of the second type; or

(c) the amino acid residue at the 114-123 position of the wild-type HPV18L1 protein is replaced by the amino acid residue at the corresponding position of the L1 protein of the wild-type HPV of the second type; or

(d) The amino acid residue at the 176-202 th position of the wild-type HPV18L1 protein is replaced by the amino acid residue at the corresponding position of the L1 protein of the wild-type HPV of the second type;

And, the variant differs from the mutated HPV18L1 protein only in the substitution (preferably conservative substitution), addition or deletion of one or a few (e.g., 1, 2, 3, 4, 5, 6, 7, 8 or 9) amino acids and retains the function of the mutated HPV18L1 protein, i.e., is capable of inducing neutralizing antibodies against at least two additional types of HPV (e.g., HPV18 and HPV45, or HPV18, HPV45 and HPV 59);

Preferably, the mutated HPV18L1 protein has the mutation defined in (2) (a) or (2) (b), and optionally, also has the following mutations:

(3) The amino acid residue at the 112-123 position of the wild-type HPV18L1 protein is replaced by the amino acid residue at the corresponding position of the wild-type HPV L1 protein of the third type;

preferably, the mutated HPV18L1 protein has the mutations defined in (2) (c) or (2) (d), and optionally, also has the following mutations:

(4) The amino acid residue at the 410-421 position of the wild-type HPV18L1 protein is replaced by the amino acid residue at the corresponding position of the wild-type HPV L1 protein of the third type;

Preferably, the mutated HPV18L1 protein has the mutations defined in (2) (c) and (2) (a), and optionally also the mutation defined in (4);

preferably, the mutant HPV18L1 protein is N-terminally truncated by 45, 50, 52, 55, 58, 60, 62, 65, 68, 70, 72, 75 or 78 amino acids compared to the wild-type HPV18L1 protein;

preferably, the mutated HPV18L1 protein is N-terminally truncated by 65 amino acids compared to the wild-type HPV18L1 protein;

Preferably, the wild-type HPV of the second type is HPV 45; preferably, the amino acid residue at the corresponding position in (2) (a) is the amino acid residue at position 201-209 of the wild-type HPV45L1 protein; preferably, the amino acid residue at the corresponding position in (2) (b) is the amino acid residue at position 293-314 of the wild-type HPV45L1 protein; preferably, the amino acid residues at the corresponding positions in (2) (c) are amino acid residues at positions 79-89 of wild-type HPV45L1 protein; preferably, the amino acid residue at the corresponding position in (2) (d) is the amino acid residue at position 142-168 of the wild-type HPV45L1 protein;

Preferably, the wild-type HPV of the third type is HPV 59; preferably, the amino acid residues at the corresponding positions in (3) are amino acid residues at positions 51-62 of wild-type HPV59L1 protein; preferably, the amino acid residue at the corresponding position in (4) is the amino acid residue at position 349-360 of the wild-type HPV59L1 protein;

Preferably, the wild-type HPV18L1 protein has an amino acid sequence as shown in SEQ ID NO 1 or 113;

Preferably, the wild-type HPV45L1 protein has an amino acid sequence shown as SEQ ID NO 2 or 115;

preferably, the wild-type HPV59L1 protein has an amino acid sequence shown as SEQ ID NO. 3;

Preferably, the mutated HPV18L1 protein has an amino acid sequence selected from the group consisting of: 6, 7, 9, 13, 17, 18 or 19 SEQ ID NO.

2. An isolated nucleic acid encoding the mutated HPV18L1 protein of claim 1 or a variant thereof,

Preferably, the isolated nucleic acid has a nucleotide sequence selected from the group consisting of: SEQ ID NOS 25, 26, 28, 32, 36, 37 and 38.

3. A vector comprising the isolated nucleic acid of claim 2.

4. A host cell comprising the isolated nucleic acid of claim 2 and/or the vector of claim 3.

5. an HPV virus-like particle comprising or consisting of the mutated HPV18L1 protein of claim 1 or a variant thereof, or the mutated HPV18L1 protein of claim 1 or a variant thereof.

6. A composition comprising the mutated HPV18L1 protein or variant thereof of claim 1, or the isolated nucleic acid of claim 2, or the vector of claim 3, or the host cell of claim 4, or the HPV virus-like particle of claim 5.

7. A pharmaceutical composition or vaccine comprising the HPV virus-like particle of claim 5, optionally together with a pharmaceutically acceptable carrier and/or excipient,

Preferably, the HPV virus-like particle is present in an effective amount to prevent HPV infection or a disease caused by HPV infection;

Preferably, the HPV infection is one or more types of HPV infection (e.g., HPV18 infection, HPV45 infection, and/or HPV59 infection);

Preferably, the disease caused by HPV infection is selected from cervical cancer and condyloma acuminatum.

8. A method of making the mutant HPV18L1 protein or variant thereof of claim 1, comprising expressing the mutant HPV18L1 protein or variant thereof in a host cell, followed by recovering the mutant HPV18L1 protein or variant thereof from the culture of the host cell;

Preferably, the host cell is escherichia coli;

preferably, the method comprises the steps of: expressing the mutated HPV18L1 protein or variant thereof in Escherichia coli, and purifying the mutated HPV18L1 protein or variant thereof from a lysis supernatant of the Escherichia coli; preferably, said mutated HPV18L1 protein or variant thereof is recovered from the lysis supernatant of said e.coli by chromatography (e.g. cation exchange chromatography, hydroxyapatite chromatography and/or hydrophobic interaction chromatography).

9. A method of making a vaccine comprising admixing the HPV virus-like particle of claim 5 with a pharmaceutically acceptable carrier and/or excipient.

10. A method of preventing HPV infection or a disease caused by HPV infection, comprising administering to a subject a prophylactically effective amount of the HPV virus-like particle of claim 5 or the pharmaceutical composition or vaccine of claim 7,

Preferably, the HPV infection is one or more types of HPV infection (e.g., HPV18 infection, HPV45 infection, and/or HPV59 infection);

Preferably, the disease caused by HPV infection is selected from cervical cancer and condyloma acuminatum.

11. Use of the mutated HPV18L1 protein or variant thereof of claim 1 or the HPV virus-like particle of claim 5 in the manufacture of a pharmaceutical composition or a vaccine for the prevention of HPV infection or a disease resulting from HPV infection,

Preferably, the HPV infection is one or more types of HPV infection (e.g., HPV18 infection, HPV45 infection, and/or HPV59 infection);

Preferably, the disease caused by HPV infection is selected from cervical cancer and condyloma acuminatum.

12. The mutated HPV18L1 protein or variant thereof of claim 1 or the HPV virus-like particle of claim 5 for use in the prevention of HPV infection or a disease caused by HPV infection,

preferably, the HPV infection is one or more types of HPV infection (e.g., HPV18 infection, HPV45 infection, and/or HPV59 infection);

Preferably, the disease caused by HPV infection is selected from cervical cancer and condyloma acuminatum.

Technical Field

The present invention relates to the fields of molecular virology and immunology. In particular, the present invention relates to a mutated HPV18L1 protein (or a variant thereof), a coding sequence and a preparation method thereof, and virus-like particles comprising the same, which are capable of inducing neutralizing antibodies against at least two types of HPV (e.g., HPV18 and HPV45, or HPV18, HPV45 and HPV59), thereby being useful for preventing infection by the at least two types of HPV and diseases caused by the infection, such as cervical cancer and condyloma acuminatum. The invention also relates to the use of the above proteins and virus-like particles for the preparation of a pharmaceutical composition or vaccine useful for the prevention of HPV infections of said at least two types and diseases caused by said infections, such as cervical cancer and condyloma acuminata.

Background

Human Papilloma Virus (HPV) mainly causes warty lesions of the skin and mucous membranes. According to its relation with tumorigenesis, HPV can be classified into high-risk type and low-risk type, wherein HPV infection of high-risk type is confirmed to be a major cause of genital cancer including female cervical cancer; the low risk type mainly causes condyloma acuminatum. The most effective way to prevent and control HPV infection is to administer HPV vaccines, in particular vaccines against high-risk HPV types that can cause cervical cancer.

the major capsid protein L1 of HPV has the property of self-assembling as hollow Virus-Like particles (VLPs). HPV VLPs are 20-hedral stereosymmetric structures composed of pentamers of the 72 major capsid proteins L1 (Doorb, J.and P.H.Gallimore.1987.J Virol,61(9): 2793-9). The structure of HPV VLPs is highly similar to native HPV, retains most of the neutralizing epitopes of the native virus, and induces high titers of neutralizing antibodies (Kirnbauer, R., F. Booy, et al.1992Proc Natl Acad Sci U S A89 (24): 12180-4).

However, existing studies show that HPV VLPs induce mainly neutralizing antibodies against the same type HPV, resulting in protective immunity against the same type HPV, while there is only low cross-protection between some highly homologous types (saral. bissett, Giada Mattiuzzo, et al.2014 vaccine.32: 6548-.

Therefore, there is a need in the art to develop HPV virus-like particles capable of inducing protective neutralizing antibodies against multiple types of HPV to more economically and effectively prevent multiple types of HPV infection and diseases caused thereby, such as cervical cancer and condyloma acuminatum.

Disclosure of Invention

The present invention is based, at least in part, on the following unexpected findings of the inventors: after replacing a specific segment of the Human Papillomavirus (HPV) type 18L1 protein with a corresponding segment of the second type HPV (e.g., HPV45) L1 protein, the resulting mutant HPV18L1 protein is able to induce the body to produce high titer neutralizing antibodies against HPV18 and the second type HPV (e.g., HPV45), with protective effects comparable to mixed HPV18 VLPs and the second type HPV VLPs, and with protective effects against HPV18 comparable to HPV18 VLPs alone, and with protective effects against the second type HPV (e.g., HPV45) comparable to the second type HPV VLPs alone.

Furthermore, on the basis of the above substitution, another specific segment in the HPV18L1 protein may be further substituted with a corresponding segment of the L1 protein of HPV of the third type (e.g., HPV59), whereby the resulting mutant HPV18L1 protein containing the double substitution is capable of inducing the body to produce high-titer neutralizing antibodies against HPV18, HPV of the second type (e.g., HPV45) and HPV of the third type (e.g., HPV59), with a protective effect comparable to that of mixed HPV18 VLPs, HPV VLPs of the second type and HPV VLPs of the third type; and, its protective effect against HPV18 is comparable to HPV18 VLP alone, protective effect against HPV of the second type (e.g., HPV45) is comparable to HPV VLP of the second type alone, and protective effect against HPV of the third type (e.g., HPV59) is comparable to HPV VLP of the third type alone.

Accordingly, in one aspect, the present invention provides a mutant HPV18L1 protein or variant thereof, wherein the mutant HPV18L1 protein has the following mutations compared to the wild-type HPV18L1 protein:

(1) The N-terminal is truncated by 40-80 amino acids, such as 40-50, 45-70, 50-70, 55-65, 60-70, 65-75, 60-80 or 70-80 amino acids; and

(2) (a) the amino acid residues at positions 235-243 of the L1 protein of wild-type HPV18 are replaced by the amino acid residues at the corresponding positions of the L1 protein of wild-type HPV of the second type; or

(b) The amino acid residues at the 327-346 position of the wild-type HPV18L1 protein are replaced by the amino acid residues at the corresponding positions of the L1 protein of the wild-type HPV of the second type; or

(c) The amino acid residue at the 114-123 position of the wild-type HPV18L1 protein is replaced by the amino acid residue at the corresponding position of the L1 protein of the wild-type HPV of the second type; or

(d) The amino acid residue at the 176-202 th position of the wild-type HPV18L1 protein is replaced by the amino acid residue at the corresponding position of the L1 protein of the wild-type HPV of the second type;

And, the variant differs from the mutated HPV18L1 protein only in the substitution (preferably conservative substitution), addition or deletion of one or a few (e.g., 1, 2, 3, 4, 5, 6, 7, 8 or 9) amino acids and retains the function of the mutated HPV18L1 protein, i.e., is capable of inducing neutralizing antibodies against at least two additional types of HPV (e.g., HPV18 and HPV45, or HPV18, HPV45 and HPV 59).

In certain preferred embodiments, the mutated HPV18L1 protein has the mutations defined in (2) (a) or (2) (b), and optionally, further has the following mutations:

(3) The amino acid residues at positions 112 and 123 of the wild-type HPV18L1 protein are replaced by the amino acid residues at the corresponding positions of the wild-type HPV L1 protein of the third type.

In certain preferred embodiments, the mutated HPV18L1 protein has the mutations defined in (2) (c) or (2) (d), and optionally, also has the following mutations:

(4) The amino acid residue at the 410-421 position of the wild-type HPV18L1 protein is replaced by the amino acid residue at the corresponding position of the wild-type HPV L1 protein of the third type.

in certain preferred embodiments, the mutated HPV18L1 protein has the mutations defined in (2) (c) and (2) (a), and optionally, also has the mutation defined in (4).

in certain preferred embodiments, the mutated HPV18L1 protein has the following mutations: the amino acid residues at positions 235-243 of the wild-type HPV18L1 protein are replaced by the amino acid residues at the corresponding positions of the second type wild-type HPV L1 protein, and the amino acid residues at positions 112-123 are replaced by the amino acid residues at the corresponding positions of the third type wild-type HPV L1 protein.

In certain preferred embodiments, the mutated HPV18L1 protein has the following mutations: the amino acid residues at positions 327 and 346 of the wild-type HPV18L1 protein are replaced by the amino acid residues at the corresponding positions of the wild-type HPV L1 protein of the second type, and the amino acid residues at positions 112 and 123 are replaced by the amino acid residues at the corresponding positions of the wild-type HPV L1 protein of the third type.

In certain preferred embodiments, the mutated HPV18L1 protein has the following mutations: the amino acid residues at the 114-123 position of the wild-type HPV18L1 protein are replaced by the amino acid residues at the corresponding positions of the second type wild-type HPV L1 protein, and the amino acid residues at the 410-421 position are replaced by the amino acid residues at the corresponding positions of the third type wild-type HPV L1 protein.

in certain preferred embodiments, the mutated HPV18L1 protein has the following mutations: the amino acid residues at positions 176-202 of the wild-type HPV18L1 protein are replaced by the amino acid residues at the corresponding positions of the second type wild-type HPV L1 protein, and the amino acid residues at positions 410-421 are replaced by the amino acid residues at the corresponding positions of the third type wild-type HPV L1 protein.

In certain preferred embodiments, the mutated HPV18L1 protein has the following mutations: the amino acid residue at the 114-position 123 of the wild-type HPV18L1 protein is replaced by the amino acid residue at the corresponding position of the second type wild-type HPV L1 protein, the amino acid residue at the 235-position 243 is replaced by the amino acid residue at the corresponding position of the second type wild-type HPV L1 protein, and the amino acid residue at the 410-position 421 is replaced by the amino acid residue at the corresponding position of the third type wild-type HPV VL1 protein.

In certain preferred embodiments, the mutant HPV18L1 protein is N-terminally truncated by 45, 50, 52, 55, 58, 60, 62, 65, 68, 70, 72, 75, or 78 amino acids compared to the wild-type HPV18L1 protein.

In certain preferred embodiments, the mutant HPV18L1 protein is truncated at the N-terminus by 65 amino acids compared to the wild-type HPV18L1 protein.

in certain preferred embodiments, the wild-type HPV of the second type is HPV 45. In certain preferred embodiments, the amino acid residues at the corresponding positions described in (2) (a) are the amino acid residues at positions 201-209 of the L1 protein of wild-type HPV 45. In certain preferred embodiments, the amino acid residue at the corresponding position described in (2) (b) is the amino acid residue at position 293-314 of the wild-type HPV45L1 protein. In certain preferred embodiments, the amino acid residues at the corresponding positions described in (2) (c) are amino acid residues 79-89 of wild-type HPV45L1 protein. In certain preferred embodiments, the amino acid residues at the corresponding positions described in (2) (d) are amino acid residues at positions 142 and 168 of the L1 protein of wild-type HPV 45.

In certain preferred embodiments, the wild-type HPV of the third type is HPV 59. In certain preferred embodiments, the amino acid residues at the corresponding positions described in (3) are amino acid residues 51-62 of wild-type HPV59L1 protein. In certain preferred embodiments, the amino acid residues at the corresponding positions described in (4) are amino acid residues at positions 349-360 of the L1 protein of wild-type HPV 59.

in certain preferred embodiments, the wild-type HPV18L1 protein has an amino acid sequence as set forth in SEQ ID No. 1 or 113.

In certain preferred embodiments, the wild-type HPV45L1 protein has an amino acid sequence as set forth in SEQ ID No. 2 or 115.

In certain preferred embodiments, the wild-type HPV59L1 protein has the amino acid sequence shown as SEQ ID No. 3.

in certain preferred embodiments, the sequence of amino acid residues 79-89 of the wild-type HPV45L1 protein is shown in SEQ ID NO 39.

In certain preferred embodiments, the sequence of amino acid residues 142-168 of the wild-type HPV45L1 protein is shown in SEQ ID NO 40.

In certain preferred embodiments, the sequence of amino acid residues 201-209 of the wild-type HPV45L1 protein is shown as SEQ ID NO 41.

In certain preferred embodiments, the sequence of the amino acid residues at positions 293-314 of the wild-type HPV45L1 protein is shown as SEQ ID NO: 42.

in certain preferred embodiments, the sequence of amino acid residues 51-62 of the wild-type HPV59L1 protein is depicted in SEQ ID NO 43.

In certain preferred embodiments, the sequence of amino acid residues 349-360 of the wild-type HPV59L1 protein is shown in SEQ ID NO 44.

In certain preferred embodiments, the mutated HPV18L1 protein has an amino acid sequence selected from the group consisting of seq id no:4, 5, 6, 7, 9, 13, 17, 18 or 19 SEQ ID NO.

In certain more preferred embodiments, the mutated HPV18L1 protein has an amino acid sequence selected from the group consisting of: 6, 7, 9, 13, 17, 18 or 19 SEQ ID NO.

In another aspect, the invention provides an isolated nucleic acid encoding a mutant HPV18L1 protein or variant thereof as described above. In another aspect, the invention provides a vector comprising the isolated nucleic acid. In certain preferred embodiments, the isolated nucleic acid of the invention has a nucleotide sequence selected from the group consisting of: 23, 24, 25, 26, 28, 32, 36, 37 or 38. In certain more preferred embodiments, the isolated nucleic acid of the invention has a nucleotide sequence selected from the group consisting of: 25, 26, 28, 32, 36, 37 or 38 SEQ ID NO.

Vectors useful for inserting a polynucleotide of interest are well known in the art and include, but are not limited to, cloning vectors and expression vectors. In one embodiment, the vector is, for example, a plasmid, cosmid, phage, or the like.

in another aspect, the invention also relates to a host cell comprising the isolated nucleic acid or vector described above. Such host cells include, but are not limited to, prokaryotic cells such as E.coli cells, and eukaryotic cells such as yeast cells, insect cells, plant cells, and animal cells (e.g., mammalian cells, e.g., mouse cells, human cells, etc.). The host cell of the invention may also be a cell line, such as 293T cells.

In another aspect, the invention relates to an HPV virus-like particle, wherein the virus-like particle comprises or consists of a mutated HPV18L1 protein of the invention or a variant thereof.

In certain preferred embodiments, the HPV virus-like particle of the invention comprises a mutated HPV18L1 protein which is N-terminally truncated by 40-80 amino acids, such as 40-50, 45-70, 50-70, 55-65, 60-70, 65-75, 60-80 or 70-80 amino acids, such as 45, 50, 52, 55, 58, 60, 62, 65, 68, 70, 72, 75 or 78 amino acids, compared to the wild-type HPV18L1 protein, and the amino acid residues at positions 235 and 243 of the wild-type HPV18L1 protein are replaced with the amino acid residues at positions 201 and 209 of the wild-type HPV45L1 protein.

In certain preferred embodiments, the HPV virus-like particles of the invention comprise a mutated HPV18L1 protein truncated at the N-terminus by 40-80 amino acids, such as 40-50, 45-70, 50-70, 55-65, 60-70, 65-75, 60-80 or 70-80 amino acids, such as 45, 50, 52, 55, 58, 60, 62, 65, 68, 70, 72, 75 or 78 amino acids, as compared to the wild-type HPV18L1 protein, and the amino acid residue 327 at position glancing 346 of the wild-type HPV18L1 protein is replaced with the amino acid residue at position 293 and glancing 314 of the wild-type HPV45L1 protein.

in certain preferred embodiments, the HPV virus-like particle of the invention comprises a mutated HPV18L1 protein that is N-terminally truncated by 40-80 amino acids, such as 40-50, 45-70, 50-70, 55-65, 60-70, 65-75, 60-80 or 70-80 amino acids, such as 45, 50, 52, 55, 58, 60, 62, 65, 68, 70, 72, 75 or 78 amino acids, compared to the wild-type HPV18L1 protein, and the amino acid residues located at positions 114 and 123 of the wild-type HPV18L1 protein are replaced with the amino acid residues located at positions 79 and 89 of the wild-type HPV45L1 protein.

in certain preferred embodiments, the HPV virus-like particle of the invention comprises a mutated HPV18L1 protein which is N-terminally truncated by 40-80 amino acids, such as 40-50, 45-70, 50-70, 55-65, 60-70, 65-75, 60-80 or 70-80 amino acids, such as 45, 50, 52, 55, 58, 60, 62, 65, 68, 70, 72, 75 or 78 amino acids, compared to the wild-type HPV18L1 protein, and the amino acid residue at position 176-168 of the wild-type HPV18L1 protein is replaced with the amino acid residue at position 142-168 of the wild-type HPV45L1 protein.

In certain preferred embodiments, the HPV virus-like particles of the invention comprise a mutated HPV18L1 protein, compared with wild HPV18L1 protein, the N end is truncated by 40-80 amino acids, e.g.40-50, 45-70, 50-70, 55-65, 60-70, 65-75, 60-80 or 70-80 amino acids, for example 45, 50, 52, 55, 58, 60, 62, 65, 68, 70, 72, 75 or 78 amino acids, and the amino acid residues at the 235-position 243 of the wild-type HPV18L1 protein are replaced by the amino acid residues at the 201-position 209 of the wild-type HPV45L1 protein, and the amino acid residues at the 112-123 position of the wild-type HPV18L1 protein are replaced by the amino acid residues at the 51-62 position of the wild-type HPV59L1 protein.

in certain preferred embodiments, the HPV virus-like particles of the invention comprise a mutated HPV18L1 protein, compared with wild HPV18L1 protein, the N end is truncated by 40-80 amino acids, e.g.40-50, 45-70, 50-70, 55-65, 60-70, 65-75, 60-80 or 70-80 amino acids, for example 45, 50, 52, 55, 58, 60, 62, 65, 68, 70, 72, 75 or 78 amino acids, and the amino acid residues at the 327-346 position of the wild-type HPV18L1 protein are replaced by the amino acid residues at the 293-314 position of the wild-type HPV45L1 protein, and the amino acid residues at the 112-123 position of the wild-type HPV18L1 protein are replaced by the amino acid residues at the 51-62 position of the wild-type HPV59L1 protein.

In certain preferred embodiments, the HPV virus-like particles of the invention comprise a mutated HPV18L1 protein, compared with wild HPV18L1 protein, the N end is truncated by 40-80 amino acids, e.g.40-50, 45-70, 50-70, 55-65, 60-70, 65-75, 60-80 or 70-80 amino acids, for example 45, 50, 52, 55, 58, 60, 62, 65, 68, 70, 72, 75 or 78 amino acids, and the amino acid residues at the 114-123 th positions of the wild-type HPV18L1 protein are replaced by the amino acid residues at the 79-89 th positions of the wild-type HPV45L1 protein, and the amino acid residues at the 410-421 position of the wild-type HPV18L1 protein are replaced by the amino acid residues at the 349-360 position of the wild-type HPV59L1 protein.

In certain preferred embodiments, the HPV virus-like particles of the invention comprise a mutated HPV18L1 protein, compared with wild HPV18L1 protein, the N end is truncated by 40-80 amino acids, e.g.40-50, 45-70, 50-70, 55-65, 60-70, 65-75, 60-80 or 70-80 amino acids, for example 45, 50, 52, 55, 58, 60, 62, 65, 68, 70, 72, 75 or 78 amino acids, and the amino acid residues at the 176-202 th positions of the wild-type HPV18L1 protein are replaced by the amino acid residues at the 142-168 th positions of the wild-type HPV45L1 protein, and the amino acid residues at the 410-421 position of the wild-type HPV18L1 protein are replaced by the amino acid residues at the 349-360 position of the wild-type HPV59L1 protein.

In certain preferred embodiments, the HPV virus-like particle of the invention comprises a mutated HPV18L1 protein which is N-terminally truncated by 40-80 amino acids, such as 40-50, 45-70, 50-70, 55-65, 60-70, 65-75, 60-80 or 70-80 amino acids, such as 45, 50, 52, 55, 58, 60, 62, 65, 68, 70, 72, 75 or 78 amino acids, compared to the wild-type HPV18L1 protein, and the amino acid residue at position 114 and 123 of the wild-type HPV18L1 protein is replaced by the amino acid residue at position 79-89 of the wild-type HPV45L1 protein, and the amino acid residue at position 235 and 243 of the wild-type HPV18L1 protein is replaced by the amino acid residue 201 and 209 of the wild-type HPV 2L 1 protein, and the amino acid residue at position 421 and 84 of the wild-type HPV 4642L 38410 of the wild-type HPV18L1 protein is replaced by the amino acid residue 421 and 685 84 of the wild-type HPV1 protein Amino acid residues 349-360 of the L1 protein.

In a particularly preferred embodiment, the HPV virus-like particle of the invention comprises a mutated HPV18L1 protein having the sequence shown in SEQ ID NOs 4, 5, 6, 7, 9, 13, 17, 18 or 19.

in another aspect, the invention also relates to a composition comprising the above mutated HPV18L1 protein or variant thereof, or the above isolated nucleic acid or vector or host cell or HPV virus-like particle. In certain preferred embodiments, the compositions comprise a mutated HPV18L1 protein of the invention or a variant thereof. In certain preferred embodiments, the composition comprises an HPV virus-like particle of the invention.

In another aspect, the present invention also relates to a pharmaceutical composition or vaccine comprising the HPV virus-like particle of the invention, optionally further comprising a pharmaceutically acceptable carrier and/or excipient. The pharmaceutical composition or vaccine of the present invention can be used for preventing HPV infection or diseases caused by HPV infection such as cervical cancer and condyloma acuminatum.

in certain preferred embodiments, the HPV virus-like particle is present in an effective amount to prevent HPV infection or a disease caused by HPV infection. In certain preferred embodiments, the HPV infection is one or more HPV infection of a type(s) (e.g., HPV18 infection, HPV45 infection, and/or HPV59 infection). In certain preferred embodiments, the disease caused by HPV infection is selected from cervical cancer and condyloma acuminatum.

The pharmaceutical composition or vaccine of the present invention may be administered by methods well known in the art, such as, but not limited to, oral administration or injection. In the present invention, a particularly preferred mode of administration is injection.

in certain preferred embodiments, the pharmaceutical composition or vaccine of the invention is administered in a unit dosage form. For example, and without intending to limit the present invention, the amount of HPV virus-like particles contained per unit dose is 5. mu.g-80. mu.g, preferably 20. mu.g-40. mu.g.

In another aspect, the present invention relates to a method of preparing a mutant HPV18L1 protein or variant thereof as described above, comprising expressing the mutant HPV18L1 protein or variant thereof in a host cell and recovering the mutant HPV18L1 protein or variant thereof from the culture of the host cell.

in certain preferred embodiments, the host cell is E.coli.

In certain preferred embodiments, the method comprises the steps of: expressing the mutant HPV18L1 protein or variant thereof in Escherichia coli, and purifying the mutant HPV18L1 protein or variant thereof from the lysate supernatant of the Escherichia coli. In certain preferred embodiments, the mutated HPV18L1 protein or variant thereof is recovered from the e.coli lysis supernatant by chromatography (e.g., cation exchange chromatography, hydroxyapatite chromatography and/or hydrophobic interaction chromatography).

In another aspect, the invention relates to a method of preparing a vaccine comprising admixing the HPV virus-like particle of the invention with a pharmaceutically acceptable carrier and/or excipient.

In another aspect, the present invention relates to a method of preventing HPV infection or a disease caused by HPV infection, comprising administering to a subject a prophylactically effective amount of an HPV virus-like particle or a pharmaceutical composition or a vaccine according to the invention. In a preferred embodiment, the HPV infection is one or more HPV infection of a type(s) (e.g., HPV18 infection, HPV45 infection, and/or HPV59 infection). In another preferred embodiment, the diseases caused by HPV infection include, but are not limited to, cervical cancer and condyloma acuminatum. In another preferred embodiment, the subject is a mammal, such as a human.

In another aspect, it also relates to the use of a mutated HPV18L1 protein or variant thereof or HPV virus-like particle according to the invention for the preparation of a pharmaceutical composition or a vaccine for the prevention of HPV infection or a disease caused by HPV infection. In a preferred embodiment, the HPV infection is one or more HPV infection of a type(s) (e.g., HPV18 infection, HPV45 infection, and/or HPV59 infection). In another preferred embodiment, the diseases caused by HPV infection include, but are not limited to, cervical cancer and condyloma acuminatum.

In another aspect, the present invention also relates to a mutated HPV18L1 protein or a variant thereof or an HPV virus-like particle according to the present invention, for use in the prevention of an HPV infection or a disease caused by an HPV infection. In a preferred embodiment, the HPV infection is one or more HPV infection of a type(s) (e.g., HPV18 infection, HPV45 infection, and/or HPV59 infection). In another preferred embodiment, the diseases caused by HPV infection include, but are not limited to, cervical cancer and condyloma acuminatum.

Description and explanation of related terms in the present invention

In the present invention, unless otherwise specified, scientific and technical terms used herein have the meanings that are commonly understood by those skilled in the art. Also, cell culture, molecular genetics, nucleic acid chemistry, immunology laboratory procedures, as used herein, are conventional procedures that are widely used in the relevant art. Meanwhile, in order to better understand the present invention, the definitions and explanations of related terms are provided below.

according to the present invention, the term "wild-type HPV of the second type" refers to a wild-type HPV of another type than HPV 18. In the present invention, the wild-type HPV of the second type is preferably wild-type HPV 45. According to the present invention, the term "wild-type HPV of the third type" refers to a wild-type HPV of another type than HPV18 and different from the wild-type HPV of the second type. In the present invention, the wild-type HPV of the third type is preferably wild-type HPV 59.

According to the invention, the expression "corresponding position" refers to an equivalent position in the sequences being compared when the sequences are optimally aligned, i.e. when the sequences are aligned to obtain the highest percentage identity.

According to the present invention, the term "wild-type HPV18L1 protein" refers to the major capsid protein L1 naturally occurring in human papillomavirus type 18 (HPV 18). The sequence of the wild-type HPV18L1 protein is well known in the art and can be found in various public databases (e.g., NCBI database accession numbers ARS43428.1, ARS43407.1, ARS43401.1, AAP20601.1, and ABP 99727.1).

in the present invention, when referring to the amino acid sequence of wild-type HPV18L1 protein, reference is made to SEQ ID NO:1, is described. For example, the expression "amino acid residues 235-243 of wild-type HPV18L1 protein" refers to the amino acid sequence of SEQ ID NO:1 at amino acid residue 235-243 of the polypeptide shown in the specification. However, it is understood by those skilled in the art that wild-type HPV18 may comprise a variety of isolates, and that there may be differences between the amino acid sequences of the L1 proteins of the various isolates. Further, it is understood by those skilled in the art that, despite sequence differences, the L1 proteins of different isolates of HPV18 have very high identity (typically greater than 95%, e.g., greater than 96%, greater than 97%, greater than 98%, or greater than 99%) in amino acid sequence and have essentially the same biological function. Therefore, in the present invention, the term "wild-type HPV18L1 protein" shall include not only the protein shown in SEQ ID NO:1, but also the L1 proteins of various HPV18 isolates (e.g., the HPV18L1 proteins shown in ARS43428.1, ARS43407.1, ARS43401.1, AAP20601.1 and ABP 99727.1). And, when describing a sequence fragment of wild-type HPV18L1 protein, it includes not only SEQ ID NO:1, and also includes the corresponding sequence fragment in the L1 protein of various HPV18 isolates. For example, the expression "amino acid residues 235-243 of the wild-type HPV18L1 protein" includes the amino acid sequences shown in SEQ ID NO:1, amino acid residues 235-243, and the corresponding fragments in the L1 proteins of various HPV18 isolates.

According to the present invention, the term "wild-type HPV45L1 protein" refers to the major capsid protein L1 naturally occurring in human papillomavirus type 45 (HPV 45). The sequence of the wild-type HPV45L1 protein is well known in the art and can be found in various public databases (e.g., NCBI database accession numbers P36741.1, ALV85689.1, ABP99815.1, AGU90600.1, and ALV 85649.1).

in the present invention, when referring to the amino acid sequence of wild-type HPV45L1 protein, reference is made to SEQ ID NO:2, to the sequence shown in figure 2. For example, the expression "amino acid residues 201-209 of wild-type HPV45L1 protein" refers to the amino acid sequence of SEQ ID NO:2 at amino acid residue 201-209 of the polypeptide. However, it is understood by those skilled in the art that wild-type HPV45 may comprise a variety of isolates, and that there may be differences between the amino acid sequences of the L1 proteins of the various isolates. Further, it is understood by those skilled in the art that, despite sequence differences, the L1 proteins of different isolates of HPV45 have very high identity (typically greater than 95%, e.g., greater than 96%, greater than 97%, greater than 98%, or greater than 99%) in amino acid sequence and have essentially the same biological function. Therefore, in the present invention, the term "wild-type HPV45L1 protein" shall include not only the protein shown in SEQ ID NO:2, but also the L1 proteins of various HPV45 isolates (e.g., HPV45L1 proteins shown by P36741.1, ALV85689.1, ABP99815.1, AGU90600.1 and ALV 85649.1). And, when describing a sequence fragment of wild-type HPV45L1 protein, it includes not only SEQ ID NO:2, and also includes the corresponding sequence fragment in the L1 protein of various HPV45 isolates. For example, the expression "amino acid residues 201-209 of the wild-type HPV45L1 protein" includes the amino acid sequences shown in SEQ ID NO:2, amino acid residues 201-209, and the corresponding fragment in the L1 protein of the various HPV45 isolates.

According to the present invention, the term "wild-type HPV59L1 protein" refers to the major capsid protein L1 naturally occurring in human papillomavirus type 59 (HPV 59). The sequence of the wild-type HPV59L1 protein is well known in the art and can be found in various public databases (e.g., NCBI database accession numbers CAA54856.1, AGU90656.1, AEP23088.1, AEP23087.1, and AGU 90672.1).

In the present invention, when referring to the amino acid sequence of wild-type HPV59L1 protein, reference is made to SEQ ID NO:3, is described. For example, the expression "amino acid residues 51-62 of wild-type HPV59L1 protein" means the amino acid sequence of seq id NO:3 at amino acid residues 51-62 of the polypeptide. However, it is understood by those skilled in the art that wild-type HPV59 may comprise a variety of isolates, and that there may be differences between the amino acid sequences of the L1 proteins of the various isolates. Further, it is understood by those skilled in the art that, despite sequence differences, the L1 proteins of different isolates of HPV59 have very high identity (typically greater than 95%, e.g., greater than 96%, greater than 97%, greater than 98%, or greater than 99%) in amino acid sequence and have essentially the same biological function. Therefore, in the present invention, the term "wild-type HPV59L1 protein" shall include not only the protein shown in SEQ ID NO 3, but also the L1 proteins of various HPV59 isolates (e.g., the HPV59L1 proteins shown in CAA54856.1, AGU90656.1, AEP23088.1, AEP23087.1 and AGU 90672.1). And, when describing a sequence fragment of wild-type HPV59L1 protein, it includes not only SEQ ID NO:3, and also includes the corresponding sequence fragment in the L1 protein of various HPV59 isolates. For example, the expression "amino acid residues 51-62 of wild-type HPV59L1 protein" includes SEQ ID NO:3, amino acid residues 51-62, and the corresponding fragments in the L1 protein of various HPV59 isolates.

According to the invention, the expression "corresponding sequence fragment" or "corresponding fragment" refers to the fragments at equivalent positions in the sequences being compared when the sequences are optimally aligned, i.e. when the sequences are aligned to obtain the highest percentage identity.

According to the present invention, the expression "truncated by X amino acids at the N-terminus" means that the amino acid residues 1 to X at the N-terminus of the protein are replaced with a methionine residue encoded by the start codon (for initiating translation of the protein). For example, the HPV18L1 protein truncated by 65 amino acids at the N-terminus refers to a protein obtained by replacing amino acid residues 1 to 65 of the N-terminus of wild-type HPV18L1 protein with a methionine residue encoded by the start codon.

According to the present invention, the term "variant" refers to a protein whose amino acid sequence has a substitution (preferably conservative substitution), addition or deletion of one or several (e.g., 1, 2, 3, 4, 5, 6, 7, 8 or 9) amino acids, or has at least 90%, 95%, 96%, 97%, 98%, or 99% identity, compared to the amino acid sequence of the mutated HPV18L1 protein of the present invention (protein as shown in SEQ ID NO:6, 7, 9, 13, 17, 18 or 19), and which retains the function of the mutated HPV18L1 protein. In the present invention, the term "function of mutated HPV18L1 protein" means: capable of inducing the body to produce neutralizing antibodies against at least two types of HPV (e.g., HPV18 and HPV45, or HPV18, HPV45 and HPV 59). The term "identity" is a measure of similarity of nucleotide or amino acid sequences. Sequences are usually aligned to achieve maximum matching. "identity" itself has a meaning well known in the art and can be calculated using published algorithms (e.g., BLAST).

According to the invention, the term "identity" is used to refer to the match of sequences between two polypeptides or between two nucleic acids. When a position in both of the sequences being compared is occupied by the same base or amino acid monomer subunit (e.g., a position in each of two DNA molecules is occupied by adenine, or a position in each of two polypeptides is occupied by lysine), then the molecules are identical at that position. The "percent identity" between two sequences is a function of the number of matching positions shared by the two sequences divided by the number of positions compared x 100. For example, if 6 of 10 positions of two sequences match, then the two sequences have 60% identity. For example, the DNA sequences CTGACT and CAGGTT share 50% identity (3 of the total 6 positions match). Typically, the comparison is made when the two sequences are aligned to yield maximum identity. Such alignments can be performed by using, for example, Needleman et al (1970) j.mol.biol.48: 443-453. The algorithm of E.Meyers and W.Miller (Compout.Applbiosci., 4:11-17(1988)) which has been incorporated into the ALIGN program (version 2.0) can also be used to determine percent identity between two amino acid sequences using a PAM120 weight residue table (weight residue table), a gap length penalty of 12, and a gap penalty of 4. Furthermore, percent identity between two amino acid sequences can be determined using the Needleman and Wunsch (J MoIBiol.48: 444-.

As used herein, the term "conservative substitution" means an amino acid substitution that does not adversely affect or alter the essential characteristics of the protein/polypeptide comprising the amino acid sequence. For example, conservative substitutions may be introduced by standard techniques known in the art, such as site-directed mutagenesis and PCR-mediated mutagenesis. Conservative amino acid substitutions include those in which an amino acid residue is replaced with an amino acid residue having a similar side chain, e.g., a substitution with a residue that is physically or functionally similar to the corresponding amino acid residue (e.g., of similar size, shape, charge, chemical properties, including the ability to form covalent or hydrogen bonds, etc.). Families of amino acid residues with similar side chains have been defined in the art. These families include amino acids with basic side chains (e.g., lysine, arginine, and histidine), acidic side chains (e.g., aspartic acid, glutamic acid), uncharged polar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine, tryptophan), nonpolar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine), beta-branched side chains (e.g., threonine, valine, isoleucine), and aromatic side chains (e.g., tyrosine, phenylalanine tryptophan, histidine). Thus, a conservative substitution typically refers to the replacement of a corresponding amino acid residue with another amino acid residue from the same side chain family. Methods for identifying conservative substitutions of amino acids are well known in the art (see, e.g., Brummell et al, biochem.32:1180-1187 (1993); Kobayashi et al Protein Eng.12(10):879-884 (1999); and Burks et al, Proc. Natl Acad. set USA 94:412-417(1997), which are incorporated herein by reference).

According to the present invention, the term "E.coli expression system" refers to an expression system consisting of E.coli (strain) derived from a commercially available strain, such as, but not limited to: ER2566, BL21(DE3), B834(DE3), BLR (DE 3).

According to the present invention, the term "vector" refers to a nucleic acid vehicle into which a polynucleotide can be inserted. When a vector is capable of expressing a protein encoded by an inserted polynucleotide, the vector is referred to as an expression vector. The vector may be introduced into a host cell by transformation, transduction, or transfection, and the genetic material elements carried thereby are expressed in the host cell. Vectors are well known to those skilled in the art and include, but are not limited to: a plasmid; bacteriophage; cosmids, and the like.

according to the present invention, the term "pharmaceutically acceptable carrier and/or excipient" refers to carriers and/or excipients that are pharmacologically and/or physiologically compatible with the subject and the active ingredient, which are well known in the art (see, e.g., Remington's pharmaceutical sciences. edited by genomic AR,19th ed. pennsylvania: mack publishing Company,1995) and include, but are not limited to: pH regulator, surfactant, adjuvant, and ionic strength enhancer. For example, pH adjusting agents include, but are not limited to, phosphate buffers; surfactants include, but are not limited to, cationic, anionic or nonionic surfactants, such as Tween-80; adjuvants include, but are not limited to, aluminum adjuvants (e.g., aluminum hydroxide), freund's adjuvant (e.g., complete freund's adjuvant); ionic strength enhancers include, but are not limited to, sodium chloride.

According to the present invention, the term "effective amount" means an amount effective to achieve the intended purpose. For example, a prophylactically effective amount (e.g., HPV infection) is an amount effective to prevent, or delay the onset of a disease (e.g., HPV infection). It is within the ability of those skilled in the art to determine such an effective amount.

According to the present invention, the term "chromatography" includes, but is not limited to: ion exchange chromatography (e.g., cation exchange chromatography), hydrophobic interaction chromatography, adsorption chromatography (e.g., hydroxyapatite chromatography), gel filtration (gel exclusion) chromatography, affinity chromatography.

according to the invention, the term "lysis supernatant" refers to the solution produced by the following steps: host cells (e.g., E.coli) are disrupted in a lysis solution, and insoluble matter is removed from the lysis solution containing the disrupted host cells. Various lysing solutions are known to those skilled in the art and include, but are not limited to, Tris buffer, phosphate buffer, HEPES buffer, MOPS buffer, and the like. In addition, the disruption of host cells can be accomplished by a variety of methods well known to those skilled in the art, including but not limited to homogenizer disruption, sonication, milling, high pressure extrusion, lysozyme treatment, and the like. Methods for removing insoluble materials from the lysate are also well known to those skilled in the art, and include, but are not limited to, filtration and centrifugation.

Advantageous effects of the invention

Studies have shown that although there is some cross-protection between HPV18 and other types of HPV (e.g. HPV45 and HPV59), the ability of such cross-protection is low, typically less than one percent, even less than one thousandth of the level of protection of the VLP of its own type. Therefore, the risk of infection with other types of HPV (e.g. HPV45 and HPV59) remains high for subjects vaccinated with HPV18 vaccine.

The invention provides a mutant HPV18L1 protein and an HPV virus-like particle formed by the same. The HPV virus-like particle of the present invention can provide significant cross-protection capability between HPV18 and other types of HPV (e.g., HPV45 and HPV 59). In particular, the HPV virus-like particle of the present invention is capable of inducing the body to produce high titer neutralizing antibodies against at least two types of HPV (e.g., HPV18 and HPV45, or HPV18, HPV45 and HPV59) at equivalent immune doses, and is comparable in effect to a mixture of multiple types of HPV VLPs (e.g., a mixture of HPV18 VLPs and HPV45 VLPs, or a mixture of HPV18 VLPs, HPV45 VLPs and HPV59 VLPs). Therefore, the HPV virus-like particle of the present invention can be used for simultaneously preventing infection of at least two types of HPV (e.g., HPV18 and HPV45, or HPV18, HPV45 and HPV59) and diseases associated therewith, with significant advantageous technical effects. The method has particularly remarkable advantages in the aspects of expanding the protection range of the HPV vaccine, reducing the production cost of the HPV vaccine and the like.

Embodiments of the present invention will be described in detail below with reference to the drawings and examples, but those skilled in the art will understand that the following drawings and examples are only for illustrating the present invention and do not limit the scope of the present invention. Various objects and advantageous aspects of the present invention will become apparent to those skilled in the art from the accompanying drawings and the following detailed description of the preferred embodiments.

Drawings

FIG. 1 shows the results of SDS polyacrylamide gel electrophoresis of the purified mutein of example 1. Lane M: protein molecular weight markers; lane 1 HPV18N65 (HPV 18L1 protein with 65 amino acids truncated at the N-terminus); lane 2, H18N65-45T 1; lane 3, H18N65-45T 2; lane 4H 18N65-45T 3; lane 5, H18N65-45T 4; lane 6, H18N65-45T 5; lane 7 HPV18N 65; lane 8, H18N65-45T3-59S 1; lane 9, H18N65-45T3-59S 2; lane 10, H18N65-45T3-59S 4; lane 11, H18N65-45T3-59S 5; lane 12, H18N65-45T4-59S 1; lane 13, H18N65-45T4-59S 2; lane 14, H18N65-45T4-59S 3; lane 15 HPV18N 65; lane 16, H18N65-45T4-59S 5; lane 17 HPV18N 65; lane 18, H18N65-45T1-59S 5; lane 19, H18N65-45T2-59S 5; lane 20, H18N65-45T1T3-59S 5. The results show that after chromatographic purification, the purity of the H18N-45T, H18N-45T-59S and H18N-45T 1T-59S protein can reach more than 85%.

FIG. 2 shows the results of Western blot detection of H18N-45T, H18N-45T-59S, H18N-45T 1T-59S prepared in example 1 with the broad-spectrum antibody 4B. Lane M: protein molecular weight markers; lane 1 HPV18N 65; lane 2, H18N65-45T 1; lane 3, H18N65-45T 2; lane 4H 18N65-45T 3; lane 5, H18N65-45T 4; lane 6, H18N65-45T 5; lane 7 HPV18N 65; lane 8, H18N65-45T3-59S 1; lane 9, H18N65-45T3-59S 2; lane 10, H18N65-45T3-59S 4; lane 11, H18N65-45T3-59S 5; lane 12, H18N65-45T4-59S 1; lane 13, H18N65-45T4-59S 2; lane 14, H18N65-45T4-59S 3; lane 15 HPV18N 65; lane 16, H18N65-45T4-59S 5; lane 17 HPV18N 65; lane 18, H18N65-45T1-59S 5; lane 19, H18N65-45T2-59S 5; lane 20, H18N65-45T1T3-59S 5. The results show that the mutant proteins H18N65-45T1, H18N65-45T2, H18N65-45T3, H18N65-45T4, H18N65-45T5, H18N65-45T3-59S 3, H18N 3-45T 3-3659S 3, H18N 3-3659S 3, and H18N 3-3659B 364B antibody can be specifically recognized.

FIGS. 3A-3S show a polypeptide comprising the proteins HPV18N65 (FIG. 3A), HPV45N27 (FIG. 3B), HPV59L1 (FIG. 3C), H18N65-45T1 (FIG. 3D), H18N65-45T2 (FIG. 3E), H18N65-45T3 (FIG. 3F), H18N65-45T4 (FIG. 3G), H18N 4-45T 4 (FIG. 3H), H18N 4-45T 4-59S 4 (FIG. 3I), H18N 4-45T 4-59S 4 (FIG. 3J), H18N 4-45T 4-59S 4 (FIG. 3K), H18N 4-45T 4-59S 4 (FIG. 3L), H18N 4-T4-3659S 4 (FIG. 3M-4S 4), H18N 4-4T 4-3659 (FIG. 3H-4T 4-4S 3659), H18N 4-4S 4 (FIG. 72-4P) and H-4T 4-4 (FIG. 3H-4H-4P-4H-4S 4P) as shown in FIGS. 72-4 (FIG. 72H-4H, Results of molecular sieve chromatographic analysis of samples of H18N65-45T1T3-59S5 (FIG. 3S). The results show that the VLPs comprising proteins H18N65-45T1, H18N65-45T2, H18N 2-45T 2-59S 2, H18N 2-45T 2-59S 2, H18N 2-45T 2-59S 2, VLPs 2-VLPs, VLPs (VLPs) VLPs, and VLPs (VLPs) are all assembled from the same as the HPV protein, and VLPs (VLPs) of the HPV protein, and VLPs (VLPs) of the VLPs, and VLPs (VLPs). This suggests that both of these proteins can assemble into VLPs.

Fig. 4A-4S show transmission electron microscopy observations of various VLP samples (magnification 100,000, Bar 0.1 μm). Wherein: fig. 4A, VLPs assembled from HPV18N 65; fig. 4B, VLPs assembled from HPV45N 27; fig. 4C, VLP assembled by HPV59L 1; FIG. 4D, VLPs assembled from H18N65-45T 1; FIG. 4E, VLPs assembled from H18N65-45T 2; FIG. 4F, VLP assembled from H18N65-45T 3; FIG. 4G, VLPs assembled from H18N65-45T 4; FIG. 4H, VLP assembled from H18N65-45T 5; FIG. 4I, VLP assembled from H18N65-45T3-59S 1; FIG. 4J, VLP assembled from H18N65-45T3-59S 2; FIG. 4K, VLP assembled from H18N65-45T3-59S 4; FIG. 4L, VLP assembled from H18N65-45T3-59S 5; FIG. 4M, VLP assembled from H18N65-45T4-59S 1; FIG. 4N, VLP assembled from H18N65-45T4-59S 2; FIG. 4O, VLP assembled from H18N65-45T4-59S 3; FIG. 4P, VLP assembled from H18N65-45T4-59S 5; FIG. 4Q, VLP assembled from H18N65-45T1-59S 5; FIG. 4R, VLP assembled from H18N65-45T2-59S 5; FIG. 4S, VLP assembled from H18N65-45T1T3-59S 5. The results show that H18N65-45T1, H18N65-45T2, H18N65-45T3, H18N 3-45T 3-59S 3, H3-N3-59S 3, VLP 3-N3-3, VLP 3-VLP 3659 and VLP 3-.

FIGS. 5A-5H show the results of sedimentation rate analysis of HPV18N65 VLPs, HPV45N27 VLPs, HPV59 VLPs, H18N65-45T3 VLPs, H18N65-45T4 VLPs, H18N65-45T3-59S1 VLPs, H18N65-45T4-59S1 VLPs, H18N65-45T1T3-59S5 VLPs. FIG. 5A, HPV18N65 VLP; FIG. 5B, HPV45N27 VLP; FIG. 5C, HPV59 VLP; FIG. 5D, H18N65-45T3 VLP; FIG. 5E, H18N65-45T4 VLP; FIG. 5F, H18N65-45T3-59S1 VLP; FIG. 5G, H18N65-45T4-59S1 VLP; FIG. 5H, H18N65-45T1T3-59S5 VLP. The results showed that the sedimentation coefficients of H18N65-45T3VLP, H18N65-45T4VLP, H18N65-45T1T3-59S5VLP were 143.7S, 173.3S, 167.1S, respectively, which are similar to those of HPV18N65VLP, HPV45N27VLP, and HPV59VLP (HPV18N65VLP, 142.2S; HPV45N VLP 27, 146.5S, and HPV59VLP, 139.3S). This indicates that H18N65-45T3, H18N65-45T4, H18N65-45T1T3-59S5 are able to assemble into virus-like particles of similar size and morphology to wild-type VLPs.

FIGS. 6A-6H show the results of testing the thermal stability of HPV18N65 VLPs, HPV45N27 VLPs, HPV59 VLPs, H18N65-45T3 VLPs, H18N65-45T4 VLPs, H18N65-45T3-59S1 VLPs, H18N65-45T4-59S1 VLPs, H18N65-45T1T3-59S5 VLPs. FIG. 6A, HPV18N65 VLP; FIG. 6B, HPV45N27 VLP; FIG. 6C, HPV59 VLP; FIG. 6D, H18N65-45T3 VLP; FIG. 6E, H18N65-45T4 VLP; FIG. 6F, H18N65-45T3-59S1 VLP; FIG. 6G, H18N65-45T4-59S1 VLP; FIG. 6H, H18N65-45T1T3-59S5 VLP. The results show that VLPs formed from each protein have extremely high thermostability.

FIG. 7A shows the results of the assessment of the immunoprotection in mice of experimental groups H18N65-45T1 VLPs, H18N65-45T2 VLPs, H18N65-45T3 VLPs, H18N65-45T4 VLPs, H18N65-45T5 VLPs and control groups HPV18N65 VLPs, HPV45N27 VLPs, mixed HPV18/45 VLPs. The results show that H18N65-45T1 VLPs, H18N65-45T2, VLPH18N65-45T3 VLPs and H18N65-45T4 VLPs each retain the activity of inducing high titers of neutralizing antibodies against HPV18 in mice, and that their protective effect against HPV18 is comparable to HPV18N65VLP alone, mixed HPV18/HPV45 VLPs, and that their activity of inducing neutralizing antibodies against HPV45 in mice is also higher than that of HPV18N65VLP alone. In particular, H18N65-45T3 VLPs and H18N65-45T4 VLPs each induce high titers of neutralizing antibodies against HPV45 and HPV18 in mice; and its protective effect against HPV18 is comparable to HPV18N65VLP alone, mixed HPV18/HPV45VLP, and significantly higher than HPV45N27VLP alone; and its protective effect against HPV45 is comparable to HPV45N27VLP alone, mixed HPV18/HPV45VLP, and significantly higher than HPV18N65VLP alone. This indicates that, after mutation, the H18N65-45T 1VLP, H18N65-45T2, VLPH18N65-45T 3VLP and H18N65-45T4VLP not only retain immunogenicity against HPV18, but also have improved immunogenicity against HPV45 relative to HPV18N65VLP, and in particular, the H18N65-45T3VLP and the H18N65-45T4VLP have good cross-immunogenicity and cross-protection against HPV18 and HPV45, and are useful as effective vaccines for preventing HPV18 infection and/or HPV45 infection, and as a mixed vaccine replacing HPV18 VLP and HPV45 VLP.

FIG. 7B shows the results of the immunoprotection in mice of experimental groups H18N65-45T3-59S1 VLPs, H18N65-45T3-59S2 VLPs, H18N65-45T3-59S4 VLPs, H18N65-45T3-59S5 VLPs, H18N65-45T4-59S1 VLPs, H18N65-45T4-59S2 VLPs, H18N65-45T4-59S3 VLPs, H18N65-45T4-59S5 and control groups HPV18N65 VLPs, HPV45N27 VLPs, HPV59 VLPs and mixed HPV18/HPV45/HPV59 VLPs. The results show that H18N65-45T3-59S1 VLPs and H18N65-45T4-59S1 VLPs can each induce high titers of neutralizing antibodies against HPV18, HPV45, and HPV59 in mice; and its protective effect against HPV18 is comparable to HPV18N65VLP alone, mixed HPV18/HPV45/HPV59VLP, and significantly higher than HPV45N27VLP alone and HPV59VLP alone; and its protective effect against HPV45 is comparable to HPV45N27VLP alone, mixed HPV18/HPV45/HPV59VLP, and significantly higher than HPV18N65VLP alone and HPV59VLP alone; and the protective effect against HPV59 is equivalent to that of single HPV59VLP, mixed HPV18/HPV45/HPV59VLP and is obviously higher than that of single HPV45N27VLP and single HPV18N65 VLP. This indicates that the H18N65-45T3-59S1 VLPs and H18N65-45T4-59S1 VLPs have good cross immunogenicity and cross-protection to HPV18, HPV45 and HPV59, can be used as an effective vaccine for preventing HPV18 infection, HPV45 infection and/or HPV59 infection, and can be used to replace mixed vaccines containing HPV18 VLPs, HPV45 VLPs and HPV59 VLPs.

FIGS. 8A-8B show the results of the evaluation of neutralizing antibody titers in mouse sera after immunization of mice with H18N65-45T4 VLP. FIG. 8A: 10 μ g dose group (immunization dose 10 μ g, using aluminum adjuvant); FIG. 8B: 1 μ g dose group (immunization dose 1 μ g, using aluminum adjuvant). The results show that the H18N65-45T4VLP can induce mice to generate high-titer neutralizing antibodies against HPV18, the protective effect of the H18N65-45T4VLP is equivalent to that of single HPV18N65VLP and mixed HPV18/45 VLP at the same dose, and the H18N65-45T4VLP is remarkably superior to that of single HPV45N27VLP at the same dose; and the protective effect of the polypeptide can induce mice to generate high-titer neutralizing antibodies against HPV45, is equivalent to that of single HPV45N27VLP and mixed HPV18/45 VLP at the same dose, and is remarkably superior to that of single HPV18N65VLP at the same dose. This indicates that the H18N65-45T4 VLPs have good cross-immunogenicity and cross-protection against HPV18 and HPV 45.

FIGS. 8C-8D show the results of evaluation of neutralizing antibody titers in mouse sera after immunization of mice with H18N65-45T3-59S1 VLPs, H18N65-45T4-59S1 VLPs, H18N65-45T1-59S5 VLPs, H18N65-45T2-59S5 VLPs, H18N65-45T1T3-59S5 VLPs. FIG. 8C: 10 μ g dose group (immunization dose 10 μ g, using aluminum adjuvant); FIG. 8D: 1 μ g dose group (immunization dose 1 μ g, using aluminum adjuvant). The results show that H18N65-45T3-59S1 VLPs, H18N65-45T4-59S1 VLPs, H18N65-45T1-59S5 VLPs, H18N65-45T2-59S5 VLPs and H18N65-45T1T3-59S5 VLPs can induce mice to produce high titer neutralizing antibodies against HPV18, the protective effect thereof is comparable to that of single HPV18N65 VLPs and mixed HPV18/HPV45/HPV59 VLPs at the same dose, and is significantly superior to that of single HPV45N27 VLPs or single HPV59 VLPs at the same dose; and the H18N65-45T3-59S1 VLPs, H18N65-45T4-59S1 VLPs, H18N65-45T1-59S5 VLPs, H18N65-45T2-59S5 VLPs and H18N65-45T1T3-59S5 VLPs can induce mice to produce high-titer neutralizing antibodies against HPV45, the protective effect of the neutralizing antibodies is equivalent to that of single HPV45N27 VLPs and mixed HPV18/HPV45/HPV59 VLPs at the same dosage, and is remarkably superior to that of single HPV18N65 VLPs or single HPV59 VLPs at the same dosage; and the H18N65-45T3-59S 1VLP, H18N65-45T4-59S1VLP, H18N65-45T1-59S 5VLP, H18N65-45T2-59S 5VLP and H18N65-45T1T3-59S5VLP can induce mice to produce high-titer neutralizing antibodies against HPV59, and the protective effect of the neutralizing antibodies is equivalent to that of single HPV59VLP and mixed HPV18/HPV45/HPV59VLP at the same dosage and is remarkably superior to that of single HPV18N65VLP or single HPV45N27VLP at the same dosage. This indicates that H18N65-45T3-59S1 VLPs, H18N65-45T4-59S1 VLPs, H18N65-45T1-59S5 VLPs, H18N65-45T2-59S5 VLPs and H18N65-45T1T3-59S5 VLPs have good cross-immunogenicity and cross-protection against HPV18, HPV45 and HPV 59.

Sequence information

information on the partial sequences to which the present invention relates is provided in table 1 below.

Table 1: description of the sequences

Sequence 1(SEQ ID NO: 1):

MCLYTRVLILHYHLLPLYGPLYHPQPLPLHSILVYMVHIIICGHYIILFLRNVNVFPIFLQMALWRPSDNTVYLPPPSVARVVNTDDYVTRTSIFYHAGSSRLLTVGNPYFRVPAGGGNKQDIPKVSAYQYRVFRVQLPDPNKFGLPDTSIYNPETQRLVWACAGVEIGRGQPLGVGLSGHPFYNKLDDTESSHAATSNVSEDVRDNVSVDYKQTQLCILGCAPAIGEHWAKGTACKSRPLSQGDCPPLELKNTVLEDGDMVDTGYGAMDFSTLQDTKCEVPLDICQSICKYPDYLQMSADPYGDSMFFCLRREQLFARHFWNRAGTMGDTVPQSLYIKGTGMRASPGSCVYSPSPSGSIVTSDSQLFNKPYWLHKAQGHNNGVCWHNQLFVTVVDTTRSTNLTICASTQSPVPGQYDATKFKQYSRHVEEYDLQFIFQLCTITLTADVMSYIHSMNSSILEDWNFGVPPPPTTSLVDTYRFVQSVAIACQKDAAPAENKDPYDKLKFWNVDLKEKFSLDLDQYPLGRKFLVQAGLRRKPTIGPRKRSAPSATTASKPAKRVRVRARK

Sequence 2(SEQ ID NO: 2):

MAHNIIYGHGIIIFLKNVNVFPIFLQMALWRPSDSTVYLPPPSVARVVSTDDYVSRTSIFYHAGSSRLLTVGNPYFRVVPNGAGNKQAVPKVSAYQYRVFRVALPDPNKFGLPDSTIYNPETQRLVWACVGMEIGRGQPLGIGLSGHPFYNKLDDTESAHAATAVITQDVRDNVSVDYKQTQLCILGCVPAIGEHWAKGTLCKPAQLQPGDCPPLELKNTIIEDGDMVDTGYGAMDFSTLQDTKCEVPLDICQSICKYPDYLQMSADPYGDSMFFCLRREQLFARHFWNRAGVMGDTVPTDLYIKGTSANMRETPGSCVYSPSPSGSIITSDSQLFNKPYWLHKAQGHNNGICWHNQLFVTVVDTTRSTNLTLCASTQNPVPSTYDPTKFKQYSRHVEEYDLQFIFQLCTITLTAEVMSYIHSMNSSILENWNFGVPPPPTTSLVDTYRFVQSVAVTCQKDTTPPEKQDPYDKLKFWTVDLKEKFSSDLDQYPLGRKFLVQAGLRRRPTIGPRKRPAASTSTASTASRPAKRVRIRSKK

Sequence 3(SEQ ID NO: 3):

MALWRSSDNKVYLPPPSVAKVVSTDEYVTRTSIFYHAGSSRLLTVGHPYFKVPKGGNGRQDVPKVSAYQYRVFRVKLPDPNKFGLPDNTVYDPNSQRLVWACVGVEIGRGQPLGVGLSGHPLYNKLDDTENSHVASAVDTKDTRDNVSVDYKQTQLCIIGCVPAIGEHWTKGTACKPTTVVQGDCPPLELINTPIEDGDMVDTGYGAMDFKLLQDNKSEVPLDICQSICKYPDYLQMSADAYGDSMFFCLRREQVFARHFWNRSGTMGDQLPESLYIKGTDIRANPGSYLYSPSPSGSVVTSDSQLFNKPYWLHKAQGLNNGICWHNQLFLTVVDTTRSTNLSVCASTTSSIPNVYTPTSFKEYARHVEEFDLQFIFQLCKITLTTEVMSYIHNMNTTILEDWNFGVTPPPTASLVDTYRFVQSAAVTCQKDTAPPVKQDPYDKLKFWPVDLKERFSADLDQFPLGRKFLLQLGARPKPTIGPRKRAAPAPTSTPSPKRVKRRKSSRK

Sequence 4(SEQ ID NO: 4):

MRPSDNTVYLPPPSVARVVNTDDYVTRTSIFYHAGSSRLLTVGNPYFRVVPNGAGNKQAVPKVSAYQYRVFRVQLPDPNKFGLPDTSIYNPETQRLVWACAGVEIGRGQPLGVGLSGHPFYNKLDDTESSHAATSNVSEDVRDNVSVDYKQTQLCILGCAPAIGEHWAKGTACKSRPLSQGDCPPLELKNTVLEDGDMVDTGYGAMDFSTLQDTKCEVPLDICQSICKYPDYLQMSADPYGDSMFFCLRREQLFARHFWNRAGTMGDTVPQSLYIKGTGMRASPGSCVYSPSPSGSIVTSDSQLFNKPYWLHKAQGHNNGVCWHNQLFVTVVDTTRSTNLTICASTQSPVPGQYDATKFKQYSRHVEEYDLQFIFQLCTITLTADVMSYIHSMNSSILEDWNFGVPPPPTTSLVDTYRFVQSVAIACQKDAAPAENKDPYDKLKFWNVDLKEKFSLDLDQYPLGRKFLVQAGLRRKPTIGPRKRSAPSATTASKPAKRVRVRARK

Sequence 5(SEQ ID NO: 5):

MRPSDNTVYLPPPSVARVVNTDDYVTRTSIFYHAGSSRLLTVGNPYFRVPAGGGNKQDIPKVSAYQYRVFRVQLPDPNKFGLPDTSIYNPETQRLVWACAGVEIGRGQPLGIGLSGHPFYNKLDDTESAHAATAVITQDVRDNVSVDYKQTQLCILGCAPAIGEHWAKGTACKSRPLSQGDCPPLELKNTVLEDGDMVDTGYGAMDFSTLQDTKCEVPLDICQSICKYPDYLQMSADPYGDSMFFCLRREQLFARHFWNRAGTMGDTVPQSLYIKGTGMRASPGSCVYSPSPSGSIVTSDSQLFNKPYWLHKAQGHNNGVCWHNQLFVTVVDTTRSTNLTICASTQSPVPGQYDATKFKQYSRHVEEYDLQFIFQLCTITLTADVMSYIHSMNSSILEDWNFGVPPPPTTSLVDTYRFVQSVAIACQKDAAPAENKDPYDKLKFWNVDLKEKFSLDLDQYPLGRKFLVQAGLRRKPTIGPRKRSAPSATTASKPAKRVRVRARK

Sequence 6(SEQ ID NO: 6):

MRPSDNTVYLPPPSVARVVNTDDYVTRTSIFYHAGSSRLLTVGNPYFRVPAGGGNKQDIPKVSAYQYRVFRVQLPDPNKFGLPDTSIYNPETQRLVWACAGVEIGRGQPLGVGLSGHPFYNKLDDTESSHAATSNVSEDVRDNVSVDYKQTQLCILGCAPAIGEHWAKGTLCKPAQLQPGDCPPLELKNTVLEDGDMVDTGYGAMDFSTLQDTKCEVPLDICQSICKYPDYLQMSADPYGDSMFFCLRREQLFARHFWNRAGTMGDTVPQSLYIKGTGMRASPGSCVYSPSPSGSIVTSDSQLFNKPYWLHKAQGHNNGVCWHNQLFVTVVDTTRSTNLTICASTQSPVPGQYDATKFKQYSRHVEEYDLQFIFQLCTITLTADVMSYIHSMNSSILEDWNFGVPPPPTTSLVDTYRFVQSVAIACQKDAAPAENKDPYDKLKFWNVDLKEKFSLDLDQYPLGRKFLVQAGLRRKPTIGPRKRSAPSATTASKPAKRVRVRARK

sequence 7(SEQ ID NO: 7):

MRPSDNTVYLPPPSVARVVNTDDYVTRTSIFYHAGSSRLLTVGNPYFRVPAGGGNKQDIPKVSAYQYRVFRVQLPDPNKFGLPDTSIYNPETQRLVWACAGVEIGRGQPLGVGLSGHPFYNKLDDTESSHAATSNVSEDVRDNVSVDYKQTQLCILGCAPAIGEHWAKGTACKSRPLSQGDCPPLELKNTVLEDGDMVDTGYGAMDFSTLQDTKCEVPLDICQSICKYPDYLQMSADPYGDSMFFCLRREQLFARHFWNRAGVMGDTVPTDLYIKGTSANMRETPGSCVYSPSPSGSIVTSDSQLFNKPYWLHKAQGHNNGVCWHNQLFVTVVDTTRSTNLTICASTQSPVPGQYDATKFKQYSRHVEEYDLQFIFQLCTITLTADVMSYIHSMNSSILEDWNFGVPPPPTTSLVDTYRFVQSVAIACQKDAAPAENKDPYDKLKFWNVDLKEKFSLDLDQYPLGRKFLVQAGLRRKPTIGPRKRSAPSATTASKPAKRVRVRARK

Sequence 8(SEQ ID NO: 8):

MRPSDNTVYLPPPSVARVVNTDDYVTRTSIFYHAGSSRLLTVGNPYFRVPAGGGNKQDIPKVSAYQYRVFRVQLPDPNKFGLPDTSIYNPETQRLVWACAGVEIGRGQPLGVGLSGHPFYNKLDDTESSHAATSNVSEDVRDNVSVDYKQTQLCILGCAPAIGEHWAKGTACKSRPLSQGDCPPLELKNTVLEDGDMVDTGYGAMDFSTLQDTKCEVPLDICQSICKYPDYLQMSADPYGDSMFFCLRREQLFARHFWNRAGTMGDTVPQSLYIKGTGMRASPGSCVYSPSPSGSIVTSDSQLFNKPYWLHKAQGHNNGVCWHNQLFVTVVDTTRSTNLTICASTQNPVPSTYDPTKFKQYSRHVEEYDLQFIFQLCTITLTADVMSYIHSMNSSILEDWNFGVPPPPTTSLVDTYRFVQSVAIACQKDAAPAENKDPYDKLKFWNVDLKEKFSLDLDQYPLGRKFLVQAGLRRKPTIGPRKRSAPSATTASKPAKRVRVRARK

Sequence 9(SEQ ID NO: 9):

MRPSDNTVYLPPPSVARVVNTDDYVTRTSIFYHAGSSRLLTVGNPYFKVPKGGNGRQDVPKVSAYQYRVFRVQLPDPNKFGLPDTSIYNPETQRLVWACAGVEIGRGQPLGVGLSGHPFYNKLDDTESSHAATSNVSEDVRDNVSVDYKQTQLCILGCAPAIGEHWAKGTLCKPAQLQPGDCPPLELKNTVLEDGDMVDTGYGAMDFSTLQDTKCEVPLDICQSICKYPDYLQMSADPYGDSMFFCLRREQLFARHFWNRAGTMGDTVPQSLYIKGTGMRASPGSCVYSPSPSGSIVTSDSQLFNKPYWLHKAQGHNNGVCWHNQLFVTVVDTTRSTNLTICASTQSPVPGQYDATKFKQYSRHVEEYDLQFIFQLCTITLTADVMSYIHSMNSSILEDWNFGVPPPPTTSLVDTYRFVQSVAIACQKDAAPAENKDPYDKLKFWNVDLKEKFSLDLDQYPLGRKFLVQAGLRRKPTIGPRKRSAPSATTASKPAKRVRVRARK

Sequence 10(SEQ ID NO: 10):

MRPSDNTVYLPPPSVARVVNTDDYVTRTSIFYHAGSSRLLTVGNPYFRVPAGGGNKQDIPKVSAYQYRVFRVQLPDPNKFGLPDTSIYNPETQRLVWACAGVEIGRGQPLGVGLSGHPLYNKLDDTENSHVASAVDTKDTRDNVSVDYKQTQLCILGCAPAIGEHWAKGTLCKPAQLQPGDCPPLELKNTVLEDGDMVDTGYGAMDFSTLQDTKCEVPLDICQSICKYPDYLQMSADPYGDSMFFCLRREQLFARHFWNRAGTMGDTVPQSLYIKGTGMRASPGSCVYSPSPSGSIVTSDSQLFNKPYWLHKAQGHNNGVCWHNQLFVTVVDTTRSTNLTICASTQSPVPGQYDATKFKQYSRHVEEYDLQFIFQLCTITLTADVMSYIHSMNSSILEDWNFGVPPPPTTSLVDTYRFVQSVAIACQKDAAPAENKDPYDKLKFWNVDLKEKFSLDLDQYPLGRKFLVQAGLRRKPTIGPRKRSAPSATTASKPAKRVRVRARK

Sequence 11(SEQ ID NO: 11):

MRPSDNTVYLPPPSVARVVNTDDYVTRTSIFYHAGSSRLLTVGNPYFRVPAGGGNKQDIPKVSAYQYRVFRVQLPDPNKFGLPDTSIYNPETQRLVWACAGVEIGRGQPLGVGLSGHPFYNKLDDTESSHAATSNVSEDVRDNVSVDYKQTQLCILGCAPAIGEHWAKGTLCKPAQLQPGDCPPLELKNTVLEDGDMVDTGYGAMDFSTLQDTKCEVPLDICQSICKYPDYLQMSADPYGDSMFFCLRREQLFARHFWNRSGTMGDQLPESLYIKGTDIRANPGSYLYSPSPSGSIVTSDSQLFNKPYWLHKAQGHNNGVCWHNQLFVTVVDTTRSTNLTICASTQSPVPGQYDATKFKQYSRHVEEYDLQFIFQLCTITLTADVMSYIHSMNSSILEDWNFGVPPPPTTSLVDTYRFVQSVAIACQKDAAPAENKDPYDKLKFWNVDLKEKFSLDLDQYPLGRKFLVQAGLRRKPTIGPRKRSAPSATTASKPAKRVRVRARK

sequence 12(SEQ ID NO: 12):

MRPSDNTVYLPPPSVARVVNTDDYVTRTSIFYHAGSSRLLTVGNPYFRVPAGGGNKQDIPKVSAYQYRVFRVQLPDPNKFGLPDTSIYNPETQRLVWACAGVEIGRGQPLGVGLSGHPFYNKLDDTESSHAATSNVSEDVRDNVSVDYKQTQLCILGCAPAIGEHWAKGTLCKPAQLQPGDCPPLELKNTVLEDGDMVDTGYGAMDFSTLQDTKCEVPLDICQSICKYPDYLQMSADPYGDSMFFCLRREQLFARHFWNRAGTMGDTVPQSLYIKGTGMRASPGSCVYSPSPSGSIVTSDSQLFNKPYWLHKAQGHNNGVCWHNQLFVTVVDTTRSTNLTICASTTSSIPNVYTPTSFKQYSRHVEEYDLQFIFQLCTITLTADVMSYIHSMNSSILEDWNFGVPPPPTTSLVDTYRFVQSVAIACQKDAAPAENKDPYDKLKFWNVDLKEKFSLDLDQYPLGRKFLVQAGLRRKPTIGPRKRSAPSATTASKPAKRVRVRARK

Sequence 13(SEQ ID NO: 13):

MRPSDNTVYLPPPSVARVVNTDDYVTRTSIFYHAGSSRLLTVGNPYFKVPKGGNGRQDVPKVSAYQYRVFRVQLPDPNKFGLPDTSIYNPETQRLVWACAGVEIGRGQPLGVGLSGHPFYNKLDDTESSHAATSNVSEDVRDNVSVDYKQTQLCILGCAPAIGEHWAKGTACKSRPLSQGDCPPLELKNTVLEDGDMVDTGYGAMDFSTLQDTKCEVPLDICQSICKYPDYLQMSADPYGDSMFFCLRREQLFARHFWNRAGVMGDTVPTDLYIKGTSANMRETPGSCVYSPSPSGSIVTSDSQLFNKPYWLHKAQGHNNGVCWHNQLFVTVVDTTRSTNLTICASTQSPVPGQYDATKFKQYSRHVEEYDLQFIFQLCTITLTADVMSYIHSMNSSILEDWNFGVPPPPTTSLVDTYRFVQSVAIACQKDAAPAENKDPYDKLKFWNVDLKEKFSLDLDQYPLGRKFLVQAGLRRKPTIGPRKRSAPSATTASKPAKRVRVRARK

Sequence 14(SEQ ID NO: 14):

MRPSDNTVYLPPPSVARVVNTDDYVTRTSIFYHAGSSRLLTVGNPYFRVPAGGGNKQDIPKVSAYQYRVFRVQLPDPNKFGLPDTSIYNPETQRLVWACAGVEIGRGQPLGVGLSGHPLYNKLDDTENSHVASAVDTKDTRDNVSVDYKQTQLCILGCAPAIGEHWAKGTACKSRPLSQGDCPPLELKNTVLEDGDMVDTGYGAMDFSTLQDTKCEVPLDICQSICKYPDYLQMSADPYGDSMFFCLRREQLFARHFWNRAGVMGDTVPTDLYIKGTSANMRETPGSCVYSPSPSGSIVTSDSQLFNKPYWLHKAQGHNNGVCWHNQLFVTVVDTTRSTNLTICASTQSPVPGQYDATKFKQYSRHVEEYDLQFIFQLCTITLTADVMSYIHSMNSSILEDWNFGVPPPPTTSLVDTYRFVQSVAIACQKDAAPAENKDPYDKLKFWNVDLKEKFSLDLDQYPLGRKFLVQAGLRRKPTIGPRKRSAPSATTASKPAKRVRVRARK

Sequence 15(SEQ ID NO: 15):

MRPSDNTVYLPPPSVARVVNTDDYVTRTSIFYHAGSSRLLTVGNPYFRVPAGGGNKQDIPKVSAYQYRVFRVQLPDPNKFGLPDTSIYNPETQRLVWACAGVEIGRGQPLGVGLSGHPFYNKLDDTESSHAATSNVSEDVRDNVSVDYKQTQLCILGCAPAIGEHWTKGTACKPTTVVQGDCPPLELKNTVLEDGDMVDTGYGAMDFSTLQDTKCEVPLDICQSICKYPDYLQMSADPYGDSMFFCLRREQLFARHFWNRAGVMGDTVPTDLYIKGTSANMRETPGSCVYSPSPSGSIVTSDSQLFNKPYWLHKAQGHNNGVCWHNQLFVTVVDTTRSTNLTICASTQSPVPGQYDATKFKQYSRHVEEYDLQFIFQLCTITLTADVMSYIHSMNSSILEDWNFGVPPPPTTSLVDTYRFVQSVAIACQKDAAPAENKDPYDKLKFWNVDLKEKFSLDLDQYPLGRKFLVQAGLRRKPTIGPRKRSAPSATTASKPAKRVRVRARK

sequence 16(SEQ ID NO: 16):

MRPSDNTVYLPPPSVARVVNTDDYVTRTSIFYHAGSSRLLTVGNPYFRVPAGGGNKQDIPKVSAYQYRVFRVQLPDPNKFGLPDTSIYNPETQRLVWACAGVEIGRGQPLGVGLSGHPFYNKLDDTESSHAATSNVSEDVRDNVSVDYKQTQLCILGCAPAIGEHWAKGTACKSRPLSQGDCPPLELKNTVLEDGDMVDTGYGAMDFSTLQDTKCEVPLDICQSICKYPDYLQMSADPYGDSMFFCLRREQLFARHFWNRAGVMGDTVPTDLYIKGTSANMRETPGSCVYSPSPSGSIVTSDSQLFNKPYWLHKAQGHNNGVCWHNQLFVTVVDTTRSTNLTICASTTSSIPNVYTPTSFKQYSRHVEEYDLQFIFQLCTITLTADVMSYIHSMNSSILEDWNFGVPPPPTTSLVDTYRFVQSVAIACQKDAAPAENKDPYDKLKFWNVDLKEKFSLDLDQYPLGRKFLVQAGLRRKPTIGPRKRSAPSATTASKPAKRVRVRARK

Sequence 17(SEQ ID NO: 17):

MRPSDNTVYLPPPSVARVVNTDDYVTRTSIFYHAGSSRLLTVGNPYFRVVPNGAGNKQAVPKVSAYQYRVFRVQLPDPNKFGLPDTSIYNPETQRLVWACAGVEIGRGQPLGVGLSGHPFYNKLDDTESSHAATSNVSEDVRDNVSVDYKQTQLCILGCAPAIGEHWAKGTACKSRPLSQGDCPPLELKNTVLEDGDMVDTGYGAMDFSTLQDTKCEVPLDICQSICKYPDYLQMSADPYGDSMFFCLRREQLFARHFWNRAGTMGDTVPQSLYIKGTGMRASPGSCVYSPSPSGSIVTSDSQLFNKPYWLHKAQGHNNGVCWHNQLFVTVVDTTRSTNLTICASTTSSIPNVYTPTSFKQYSRHVEEYDLQFIFQLCTITLTADVMSYIHSMNSSILEDWNFGVPPPPTTSLVDTYRFVQSVAIACQKDAAPAENKDPYDKLKFWNVDLKEKFSLDLDQYPLGRKFLVQAGLRRKPTIGPRKRSAPSATTASKPAKRVRVRARK

sequence 18(SEQ ID NO: 18):

MRPSDNTVYLPPPSVARVVNTDDYVTRTSIFYHAGSSRLLTVGNPYFRVPAGGGNKQDIPKVSAYQYRVFRVQLPDPNKFGLPDTSIYNPETQRLVWACAGVEIGRGQPLGIGLSGHPFYNKLDDTESAHAATAVITQDVRDNVSVDYKQTQLCILGCAPAIGEHWAKGTACKSRPLSQGDCPPLELKNTVLEDGDMVDTGYGAMDFSTLQDTKCEVPLDICQSICKYPDYLQMSADPYGDSMFFCLRREQLFARHFWNRAGTMGDTVPQSLYIKGTGMRASPGSCVYSPSPSGSIVTSDSQLFNKPYWLHKAQGHNNGVCWHNQLFVTVVDTTRSTNLTICASTTSSIPNVYTPTSFKQYSRHVEEYDLQFIFQLCTITLTADVMSYIHSMNSSILEDWNFGVPPPPTTSLVDTYRFVQSVAIACQKDAAPAENKDPYDKLKFWNVDLKEKFSLDLDQYPLGRKFLVQAGLRRKPTIGPRKRSAPSATTASKPAKRVRVRARK

Sequence 19(SEQ ID NO: 19):

MRPSDNTVYLPPPSVARVVNTDDYVTRTSIFYHAGSSRLLTVGNPYFRVVPNGAGNKQAVPKVSAYQYRVFRVQLPDPNKFGLPDTSIYNPETQRLVWACAGVEIGRGQPLGVGLSGHPFYNKLDDTESSHAATSNVSEDVRDNVSVDYKQTQLCILGCAPAIGEHWAKGTLCKPAQLQPGDCPPLELKNTVLEDGDMVDTGYGAMDFSTLQDTKCEVPLDICQSICKYPDYLQMSADPYGDSMFFCLRREQLFARHFWNRAGTMGDTVPQSLYIKGTGMRASPGSCVYSPSPSGSIVTSDSQLFNKPYWLHKAQGHNNGVCWHNQLFVTVVDTTRSTNLTICASTTSSIPNVYTPTSFKQYSRHVEEYDLQFIFQLCTITLTADVMSYIHSMNSSILEDWNFGVPPPPTTSLVDTYRFVQSVAIACQKDAAPAENKDPYDKLKFWNVDLKEKFSLDLDQYPLGRKFLVQAGLRRKPTIGPRKRSAPSATTASKPAKRVRVRARK

sequence 20(SEQ ID NO: 20):

ATGTGCCTGTATACACGGGTCCTGATATTACATTACCATCTACTACCTCTGTATGGCCCATTGTATCACCCACAGCCCCTGCCTCTACACAGTATATTGGTATACATGGTACACATTATTATTTGTGGCCATTATATTATTTTATTCCTAAGAAACGTAAACGTGTTCCCTATTTTTTTGCAGATGGCTTTGTGGCGGCCTAGTGACAATACCGTATATCTTCCACCTCCTTCTGTGGCAAGAGTTGTAAATACCGATGATTACGTGACTCGCACAAGCATATTTTATCATGCTGGCAGCTCTAGATTATTAACTGTTGGTAATCCATATTTTAGGGTTCCTGCAGGTGGTGGCAATAAGCAGGATATTCCTAAGGTTTCTGCATACCAATATAGAGTATTTAGGGTGCAGTTACCTGACCCAAATAAATTTGGTTTACCTGATACTAGTATTTATAATCCTGAAACACAACGTTTAGTGTGGGCCTGTGCTGGAGTGGAAATTGGCCGTGGTCAGCCTTTAGGTGTTGGCCTTAGTGGGCATCCATTTTATAATAAATTAGATGACACTGAAAGTTCCCATGCCGCCACGTCTAATGTTTCTGAGGACGTTAGGGACAATGTGTCTGTAGATTATAAGCAGACACAGTTATGTATTTTGGGCTGTGCCCCTGCTATTGGGGAACACTGGGCTAAAGGCACTGCTTGTAAATCGCGTCCTTTATCACAGGGCGATTGCCCCCCTTTAGAACTTAAAAACACAGTTTTGGAAGATGGTGATATGGTAGATACTGGATATGGTGCCATGGACTTTAGTACATTGCAAGATACTAAATGTGAGGTACCATTGGATATTTGTCAGTCTATTTGTAAATATCCTGATTATTTACAAATGTCTGCAGATCCTTATGGGGATTCCATGTTTTTTTGCTTACGGCGTGAGCAGCTTTTTGCTAGGCATTTTTGGAATAGAGCAGGTACTATGGGTGACACTGTGCCTCAATCCTTATATATTAAAGGCACAGGTATGCGTGCTTCACCTGGCAGCTGTGTGTATTCTCCCTCTCCAAGTGGCTCTATTGTTACCTCTGACTCCCAGTTGTTTAATAAACCATATTGGTTACATAAGGCACAGGGTCATAACAATGGTGTTTGCTGGCATAATCAATTATTTGTTACTGTGGTAGATACCACTCGCAGTACCAATTTAACAATATGTGCTTCTACACAGTCTCCTGTACCTGGGCAATATGATGCTACCAAATTTAAGCAGTATAGCAGACATGTTGAGGAATATGATTTGCAGTTTATTTTTCAGTTGTGTACTATTACTTTAACTGCAGATGTTATGTCCTATATTCATAGTATGAATAGCAGTATTTTAGAGGATTGGAACTTTGGTGTTCCCCCCCCGCCAACTACTAGTTTGGTGGATACATATCGTTTTGTACAATCTGTTGCTATTGCCTGTCAAAAGGATGCTGCACCGGCTGAAAATAAGGATCCCTATGATAAGTTAAAGTTTTGGAATGTGGATTTAAAGGAAAAGTTTTCTTTAGACTTAGATCAATATCCCCTTGGACGTAAATTTTTGGTTCAGGCTGGATTGCGTCGCAAGCCCACCATAGGCCCTCGCAAACGTTCTGCTCCATCTGCCACTACGGCTTCTAAACCTGCCAAGCGTGTGCGTGTACGTGCCAGGAAGTAA

sequence 21(SEQ ID NO: 21):

ATGGCACACAATATTATTTATGGCCATGGTACTATTATTTTCCTAAAAAACGTAAACGTATTCCCTATTTTTTTGCAGATGGCCCTGTGGAGGCCCAGCGACAGCACCGTGTACCTGCCCCCCCCCAGCGTGGCCAGGGTGGTGAGCACCGACGACTACGTGAGCAGGACCAGCATCTTCTACCACGCCGGCAGCAGCAGGCTGCTGACCGTGGGCAACCCCTACTTCAGGGTGGTGCCCAACGGCGCCGGCAACAAGCAGGCCGTGCCCAAGGTGAGCGCCTACCAGTACAGGGTGTTCAGGGTGGCCCTGCCCGACCCCAACAAGTTCGGCCTGCCCGACAGCACCATCTACAACCCCGAGACCCAGAGGCTGGTGTGGGCCTGCGTGGGCATGGAGATCGGCAGGGGCCAGCCCCTGGGCATCGGCCTGAGCGGCCACCCCTTCTACAACAAGCTGGACGACACCGAGAGCGCCCACGCCGCCACCGCCGTGATCACCCAGGACGTGAGGGACAACGTGAGCGTGGACTACAAGCAGACCCAGCTGTGCATCCTGGGCTGCGTGCCCGCCATCGGCGAGCACTGGGCCAAGGGCACCCTGTGCAAGCCCGCCCAGCTGCAGCCCGGCGACTGCCCCCCCCTGGAGCTGAAGAACACCATCATCGAGGACGGCGACATGGTGGACACCGGCTACGGCGCCATGGACTTCAGCACCCTGCAGGACACCAAGTGCGAGGTGCCCCTGGACATCTGCCAGAGCATCTGCAAGTACCCCGACTACCTGCAGATGAGCGCCGACCCCTACGGCGACAGCATGTTCTTCTGCCTGAGGAGGGAGCAGCTGTTCGCCAGGCACTTCTGGAACAGGGCCGGCGTGATGGGCGACACCGTGCCCACCGACCTGTACATCAAGGGCACCAGCGCCAACATGAGGGAGACCCCCGGCAGCTGCGTGTACAGCCCCAGCCCCAGCGGCAGCATCATCACCAGCGACAGCCAGCTGTTCAACAAGCCCTACTGGCTGCACAAGGCCCAGGGCCACAACAACGGCATCTGCTGGCACAACCAGCTGTTCGTGACCGTGGTGGACACCACCAGGAGCACCAACCTGACCCTGTGCGCCAGCACCCAGAACCCCGTGCCCAGCACCTACGACCCCACCAAGTTCAAGCAGTACAGCAGGCACGTGGAGGAGTACGACCTGCAGTTCATCTTCCAGCTGTGCACCATCACCCTGACCGCCGAGGTGATGAGCTACATCCACAGCATGAACAGCAGCATCCTGGAGAACTGGAACTTCGGCGTGCCCCCCCCCCCCACCACCAGCCTGGTGGACACCTACAGGTTCGTGCAGAGCGTGGCCGTGACCTGCCAGAAGGACACCACCCCCCCCGAGAAGCAGGACCCCTACGACAAGCTGAAGTTCTGGACCGTGGACCTGAAGGAGAAGTTCAGCAGCGACCTGGACCAGTACCCCCTGGGCAGGAAGTTCCTGGTGCAGGCCGGCCTGAGGAGGAGGCCCACCATCGGCCCCAGGAAGAGGCCCGCCGCCAGCACCAGCACCGCCAGCACCGCCAGCAGGCCCGCCAAGAGGGTGAGGATCAGGAGCAAGAAGTGA

sequence 22(SEQ ID NO: 22):

ATGGCCCTGTGGAGGAGCAGCGACAACAAGGTGTACCTGCCCCCCCCCAGCGTGGCCAAGGTGGTGAGCACCGACGAGTACGTGACCAGGACCAGCATCTTCTACCACGCCGGCAGCAGCAGGCTGCTGACCGTGGGCCACCCCTACTTCAAGGTGCCCAAGGGCGGCAACGGCAGGCAGGACGTGCCCAAGGTGAGCGCCTACCAGTACAGGGTGTTCAGGGTGAAGCTGCCCGACCCCAACAAGTTCGGCCTGCCCGACAACACCGTGTACGACCCCAACAGCCAGAGGCTGGTGTGGGCCTGCGTGGGCGTGGAGATCGGCAGGGGCCAGCCCCTGGGCGTGGGCCTGAGCGGCCACCCCCTGTACAACAAGCTGGACGACACCGAGAACAGCCACGTGGCCAGCGCCGTGGACACCAAGGACACCAGGGACAACGTGAGCGTGGACTACAAGCAGACCCAGCTGTGCATCATCGGCTGCGTGCCCGCCATCGGCGAGCACTGGACCAAGGGCACCGCCTGCAAGCCCACCACCGTGGTGCAGGGCGACTGCCCCCCCCTGGAGCTGATCAACACCCCCATCGAGGACGGCGACATGGTGGACACCGGCTACGGCGCCATGGACTTCAAGCTGCTGCAGGACAACAAGAGCGAGGTGCCCCTGGACATCTGCCAGAGCATCTGCAAGTACCCCGACTACCTGCAGATGAGCGCCGACGCCTACGGCGACAGCATGTTCTTCTGCCTGAGGAGGGAGCAGGTGTTCGCCAGGCACTTCTGGAACAGGAGCGGCACCATGGGCGACCAGCTGCCCGAGAGCCTGTACATCAAGGGCACCGACATCAGGGCCAACCCCGGCAGCTACCTGTACAGCCCCAGCCCCAGCGGCAGCGTGGTGACCAGCGACAGCCAGCTGTTCAACAAGCCCTACTGGCTGCACAAGGCCCAGGGCCTGAACAACGGCATCTGCTGGCACAACCAGCTGTTCCTGACCGTGGTGGACACCACCAGGAGCACCAACCTGAGCGTGTGCGCCAGCACCACCAGCAGCATCCCCAACGTGTACACCCCCACCAGCTTCAAGGAGTACGCCAGGCACGTGGAGGAGTTCGACCTGCAGTTCATCTTCCAGCTGTGCAAGATCACCCTGACCACCGAGGTGATGAGCTACATCCACAACATGAACACCACCATCCTGGAGGACTGGAACTTCGGCGTGACCCCCCCCCCCACCGCCAGCCTGGTGGACACCTACAGGTTCGTGCAGAGCGCCGCCGTGACCTGCCAGAAGGACACCGCCCCCCCCGTGAAGCAGGACCCCTACGACAAGCTGAAGTTCTGGCCCGTGGACCTGAAGGAGAGGTTCAGCGCCGACCTGGACCAGTTCCCCCTGGGCAGGAAGTTCCTGCTGCAGCTGGGCGCCAGGCCCAAGCCCACCATCGGCCCCAGGAAGAGGGCCGCCCCCGCCCCCACCAGCACCCCCAGCCCCAAGAGGGTGAAGAGGAGGAAGAGCAGCAGGAAGTGA

Sequence 23(SEQ ID NO: 23):

ATGCGGCCTAGTGACAATACCGTATATCTTCCACCTCCTTCTGTGGCAAGAGTTGTAAATACCGATGATTACGTGACTAGGACCAGCATCTTCTACCACGCCGGCAGCAGCAGGCTGCTGACCGTGGGCAACCCCTACTTCAGGGTGGTGCCCAACGGCGCCGGCAACAAGCAGGCCGTGCCCAAGGTGAGCGCCTACCAGTACAGGGTGTTCAGGGTGCAGTTACCTGACCCAAATAAATTTGGTTTACCTGATACTAGTATTTATAATCCTGAAACACAACGTTTAGTGTGGGCCTGTGCTGGAGTGGAAATTGGCCGTGGTCAGCCTTTAGGTGTTGGCCTTAGTGGGCATCCATTTTATAATAAATTAGATGACACTGAAAGTTCCCATGCCGCCACGTCTAATGTTTCTGAGGACGTTAGGGACAATGTGTCTGTAGATTATAAGCAGACACAGTTATGTATTTTGGGCTGTGCCCCTGCTATTGGGGAACACTGGGCTAAAGGCACTGCTTGTAAATCGCGTCCTTTATCACAGGGCGATTGCCCCCCTTTAGAACTTAAAAACACAGTTTTGGAAGATGGTGATATGGTAGATACTGGATATGGTGCCATGGACTTTAGTACATTGCAAGATACTAAATGTGAGGTACCATTGGATATTTGTCAGTCTATTTGTAAATATCCTGATTATTTACAAATGTCTGCAGATCCTTATGGGGATTCCATGTTTTTTTGCTTACGGCGTGAGCAGCTTTTTGCTAGGCATTTTTGGAATAGAGCAGGTACTATGGGTGACACTGTGCCTCAATCCTTATATATTAAAGGCACAGGTATGCGTGCTTCACCTGGCAGCTGTGTGTATTCTCCCTCTCCAAGTGGCTCTATTGTTACCTCTGACTCCCAGTTGTTTAATAAACCATATTGGTTACATAAGGCACAGGGTCATAACAATGGTGTTTGCTGGCATAATCAATTATTTGTTACTGTGGTAGATACCACTCGCAGTACCAATTTAACAATATGTGCTTCTACACAGTCTCCTGTACCTGGGCAATATGATGCTACCAAATTTAAGCAGTATAGCAGACATGTTGAGGAATATGATTTGCAGTTTATTTTTCAGTTGTGTACTATTACTTTAACTGCAGATGTTATGTCCTATATTCATAGTATGAATAGCAGTATTTTAGAGGATTGGAACTTTGGTGTTCCCCCCCCGCCAACTACTAGTTTGGTGGATACATATCGTTTTGTACAATCTGTTGCTATTGCCTGTCAAAAGGATGCTGCACCGGCTGAAAATAAGGATCCCTATGATAAGTTAAAGTTTTGGAATGTGGATTTAAAGGAAAAGTTTTCTTTAGACTTAGATCAATATCCCCTTGGACGTAAATTTTTGGTTCAGGCTGGATTGCGTCGCAAGCCCACCATAGGCCCTCGCAAACGTTCTGCTCCATCTGCCACTACGGCTTCTAAACCTGCCAAGCGTGTGCGTGTACGTGCCAGGAAGTAA

Sequence 24(SEQ ID NO: 24):

ATGCGGCCTAGTGACAATACCGTATATCTTCCACCTCCTTCTGTGGCAAGAGTTGTAAATACCGATGATTACGTGACTCGCACAAGCATATTTTATCATGCTGGCAGCTCTAGATTATTAACTGTTGGTAATCCATATTTTAGGGTTCCTGCAGGTGGTGGCAATAAGCAGGATATTCCTAAGGTTTCTGCATACCAATATAGAGTATTTAGGGTGCAGTTACCTGACCCAAATAAATTTGGTTTACCTGATACTAGTATTTATAATCCTGAAACACAACGTTTAGTGTGGGCCTGTGCTGGAGTGGAGATCGGCAGGGGCCAGCCCCTGGGCATCGGCCTGAGCGGCCACCCCTTCTACAACAAGCTGGACGACACCGAGAGCGCCCACGCCGCCACCGCCGTGATCACCCAGGACGTGAGGGACAACGTGAGCGTGGACTACAAGCAGACCCAGCTGTGCATCCTGGGCTGCGCCCCTGCTATTGGGGAACACTGGGCTAAAGGCACTGCTTGTAAATCGCGTCCTTTATCACAGGGCGATTGCCCCCCTTTAGAACTTAAAAACACAGTTTTGGAAGATGGTGATATGGTAGATACTGGATATGGTGCCATGGACTTTAGTACATTGCAAGATACTAAATGTGAGGTACCATTGGATATTTGTCAGTCTATTTGTAAATATCCTGATTATTTACAAATGTCTGCAGATCCTTATGGGGATTCCATGTTTTTTTGCTTACGGCGTGAGCAGCTTTTTGCTAGGCATTTTTGGAATAGAGCAGGTACTATGGGTGACACTGTGCCTCAATCCTTATATATTAAAGGCACAGGTATGCGTGCTTCACCTGGCAGCTGTGTGTATTCTCCCTCTCCAAGTGGCTCTATTGTTACCTCTGACTCCCAGTTGTTTAATAAACCATATTGGTTACATAAGGCACAGGGTCATAACAATGGTGTTTGCTGGCATAATCAATTATTTGTTACTGTGGTAGATACCACTCGCAGTACCAATTTAACAATATGTGCTTCTACACAGTCTCCTGTACCTGGGCAATATGATGCTACCAAATTTAAGCAGTATAGCAGACATGTTGAGGAATATGATTTGCAGTTTATTTTTCAGTTGTGTACTATTACTTTAACTGCAGATGTTATGTCCTATATTCATAGTATGAATAGCAGTATTTTAGAGGATTGGAACTTTGGTGTTCCCCCCCCGCCAACTACTAGTTTGGTGGATACATATCGTTTTGTACAATCTGTTGCTATTGCCTGTCAAAAGGATGCTGCACCGGCTGAAAATAAGGATCCCTATGATAAGTTAAAGTTTTGGAATGTGGATTTAAAGGAAAAGTTTTCTTTAGACTTAGATCAATATCCCCTTGGACGTAAATTTTTGGTTCAGGCTGGATTGCGTCGCAAGCCCACCATAGGCCCTCGCAAACGTTCTGCTCCATCTGCCACTACGGCTTCTAAACCTGCCAAGCGTGTGCGTGTACGTGCCAGGAAGTAA

Sequence 25(SEQ ID NO: 25):

ATGCGGCCTAGTGACAATACCGTATATCTTCCACCTCCTTCTGTGGCAAGAGTTGTAAATACCGATGATTACGTGACTCGCACAAGCATATTTTATCATGCTGGCAGCTCTAGATTATTAACTGTTGGTAATCCATATTTTAGGGTTCCTGCAGGTGGTGGCAATAAGCAGGATATTCCTAAGGTTTCTGCATACCAATATAGAGTATTTAGGGTGCAGTTACCTGACCCAAATAAATTTGGTTTACCTGATACTAGTATTTATAATCCTGAAACACAACGTTTAGTGTGGGCCTGTGCTGGAGTGGAAATTGGCCGTGGTCAGCCTTTAGGTGTTGGCCTTAGTGGGCATCCATTTTATAATAAATTAGATGACACTGAAAGTTCCCATGCCGCCACGTCTAATGTTTCTGAGGACGTTAGGGACAATGTGTCTGTAGATTATAAGCAGACACAGTTATGTATTTTGGGCTGTGCCCCCGCCATCGGCGAGCACTGGGCCAAGGGCACCCTGTGCAAGCCCGCCCAGCTGCAGCCCGGCGACTGCCCCCCCCTGGAGCTGAAGAACACCGTTTTGGAAGATGGTGATATGGTAGATACTGGATATGGTGCCATGGACTTTAGTACATTGCAAGATACTAAATGTGAGGTACCATTGGATATTTGTCAGTCTATTTGTAAATATCCTGATTATTTACAAATGTCTGCAGATCCTTATGGGGATTCCATGTTTTTTTGCTTACGGCGTGAGCAGCTTTTTGCTAGGCATTTTTGGAATAGAGCAGGTACTATGGGTGACACTGTGCCTCAATCCTTATATATTAAAGGCACAGGTATGCGTGCTTCACCTGGCAGCTGTGTGTATTCTCCCTCTCCAAGTGGCTCTATTGTTACCTCTGACTCCCAGTTGTTTAATAAACCATATTGGTTACATAAGGCACAGGGTCATAACAATGGTGTTTGCTGGCATAATCAATTATTTGTTACTGTGGTAGATACCACTCGCAGTACCAATTTAACAATATGTGCTTCTACACAGTCTCCTGTACCTGGGCAATATGATGCTACCAAATTTAAGCAGTATAGCAGACATGTTGAGGAATATGATTTGCAGTTTATTTTTCAGTTGTGTACTATTACTTTAACTGCAGATGTTATGTCCTATATTCATAGTATGAATAGCAGTATTTTAGAGGATTGGAACTTTGGTGTTCCCCCCCCGCCAACTACTAGTTTGGTGGATACATATCGTTTTGTACAATCTGTTGCTATTGCCTGTCAAAAGGATGCTGCACCGGCTGAAAATAAGGATCCCTATGATAAGTTAAAGTTTTGGAATGTGGATTTAAAGGAAAAGTTTTCTTTAGACTTAGATCAATATCCCCTTGGACGTAAATTTTTGGTTCAGGCTGGATTGCGTCGCAAGCCCACCATAGGCCCTCGCAAACGTTCTGCTCCATCTGCCACTACGGCTTCTAAACCTGCCAAGCGTGTGCGTGTACGTGCCAGGAAGTAA

sequence 26(SEQ ID NO: 26):

ATGCGGCCTAGTGACAATACCGTATATCTTCCACCTCCTTCTGTGGCAAGAGTTGTAAATACCGATGATTACGTGACTCGCACAAGCATATTTTATCATGCTGGCAGCTCTAGATTATTAACTGTTGGTAATCCATATTTTAGGGTTCCTGCAGGTGGTGGCAATAAGCAGGATATTCCTAAGGTTTCTGCATACCAATATAGAGTATTTAGGGTGCAGTTACCTGACCCAAATAAATTTGGTTTACCTGATACTAGTATTTATAATCCTGAAACACAACGTTTAGTGTGGGCCTGTGCTGGAGTGGAAATTGGCCGTGGTCAGCCTTTAGGTGTTGGCCTTAGTGGGCATCCATTTTATAATAAATTAGATGACACTGAAAGTTCCCATGCCGCCACGTCTAATGTTTCTGAGGACGTTAGGGACAATGTGTCTGTAGATTATAAGCAGACACAGTTATGTATTTTGGGCTGTGCCCCTGCTATTGGGGAACACTGGGCTAAAGGCACTGCTTGTAAATCGCGTCCTTTATCACAGGGCGATTGCCCCCCTTTAGAACTTAAAAACACAGTTTTGGAAGATGGTGATATGGTAGATACTGGATACGGCGCCATGGACTTCAGCACCCTGCAGGACACCAAGTGCGAGGTGCCCCTGGACATCTGCCAGAGCATCTGCAAGTACCCCGACTACCTGCAGATGAGCGCCGACCCCTACGGCGACAGCATGTTCTTCTGCCTGAGGAGGGAGCAGCTGTTCGCCAGGCACTTCTGGAACAGGGCCGGCGTGATGGGCGACACCGTGCCCACCGACCTGTACATCAAGGGCACCAGCGCCAACATGAGGGAGACCCCCGGCAGCTGCGTGTACAGCCCCAGCCCCAGCGGCAGCATCGTTACCTCTGACTCCCAGTTGTTTAATAAACCATATTGGTTACATAAGGCACAGGGTCATAACAATGGTGTTTGCTGGCATAATCAATTATTTGTTACTGTGGTAGATACCACTCGCAGTACCAATTTAACAATATGTGCTTCTACACAGTCTCCTGTACCTGGGCAATATGATGCTACCAAATTTAAGCAGTATAGCAGACATGTTGAGGAATATGATTTGCAGTTTATTTTTCAGTTGTGTACTATTACTTTAACTGCAGATGTTATGTCCTATATTCATAGTATGAATAGCAGTATTTTAGAGGATTGGAACTTTGGTGTTCCCCCCCCGCCAACTACTAGTTTGGTGGATACATATCGTTTTGTACAATCTGTTGCTATTGCCTGTCAAAAGGATGCTGCACCGGCTGAAAATAAGGATCCCTATGATAAGTTAAAGTTTTGGAATGTGGATTTAAAGGAAAAGTTTTCTTTAGACTTAGATCAATATCCCCTTGGACGTAAATTTTTGGTTCAGGCTGGATTGCGTCGCAAGCCCACCATAGGCCCTCGCAAACGTTCTGCTCCATCTGCCACTACGGCTTCTAAACCTGCCAAGCGTGTGCGTGTACGTGCCAGGAAGTAA

sequence 27(SEQ ID NO: 27):

ATGCGGCCTAGTGACAATACCGTATATCTTCCACCTCCTTCTGTGGCAAGAGTTGTAAATACCGATGATTACGTGACTCGCACAAGCATATTTTATCATGCTGGCAGCTCTAGATTATTAACTGTTGGTAATCCATATTTTAGGGTTCCTGCAGGTGGTGGCAATAAGCAGGATATTCCTAAGGTTTCTGCATACCAATATAGAGTATTTAGGGTGCAGTTACCTGACCCAAATAAATTTGGTTTACCTGATACTAGTATTTATAATCCTGAAACACAACGTTTAGTGTGGGCCTGTGCTGGAGTGGAAATTGGCCGTGGTCAGCCTTTAGGTGTTGGCCTTAGTGGGCATCCATTTTATAATAAATTAGATGACACTGAAAGTTCCCATGCCGCCACGTCTAATGTTTCTGAGGACGTTAGGGACAATGTGTCTGTAGATTATAAGCAGACACAGTTATGTATTTTGGGCTGTGCCCCTGCTATTGGGGAACACTGGGCTAAAGGCACTGCTTGTAAATCGCGTCCTTTATCACAGGGCGATTGCCCCCCTTTAGAACTTAAAAACACAGTTTTGGAAGATGGTGATATGGTAGATACTGGATATGGTGCCATGGACTTTAGTACATTGCAAGATACTAAATGTGAGGTACCATTGGATATTTGTCAGTCTATTTGTAAATATCCTGATTATTTACAAATGTCTGCAGATCCTTATGGGGATTCCATGTTTTTTTGCTTACGGCGTGAGCAGCTTTTTGCTAGGCATTTTTGGAATAGAGCAGGTACTATGGGTGACACTGTGCCTCAATCCTTATATATTAAAGGCACAGGTATGCGTGCTTCACCTGGCAGCTGTGTGTATTCTCCCTCTCCAAGTGGCTCTATTGTTACCTCTGACTCCCAGTTGTTTAATAAACCATATTGGTTACATAAGGCACAGGGTCATAACAATGGTGTTTGCTGGCATAATCAATTATTTGTTACTGTGGTAGATACCACTCGCAGTACCAATTTAACAATATGCGCCAGCACCCAGAACCCCGTGCCCAGCACCTACGACCCCACCAAGTTCAAGCAGTACAGCAGGCACGTGGAGGAGTACGACCTGCAGTTCATCTTCCAGCTGTGCACCATCACCCTGACCGCCGATGTTATGTCCTATATTCATAGTATGAATAGCAGTATTTTAGAGGATTGGAACTTTGGTGTTCCCCCCCCGCCAACTACTAGTTTGGTGGATACATATCGTTTTGTACAATCTGTTGCTATTGCCTGTCAAAAGGATGCTGCACCGGCTGAAAATAAGGATCCCTATGATAAGTTAAAGTTTTGGAATGTGGATTTAAAGGAAAAGTTTTCTTTAGACTTAGATCAATATCCCCTTGGACGTAAATTTTTGGTTCAGGCTGGATTGCGTCGCAAGCCCACCATAGGCCCTCGCAAACGTTCTGCTCCATCTGCCACTACGGCTTCTAAACCTGCCAAGCGTGTGCGTGTACGTGCCAGGAAGTAA

sequence 28(SEQ ID NO: 28):

ATGCGGCCTAGTGACAATACCGTATATCTTCCACCTCCTTCTGTGGCAAGAGTTGTAAATACCGATGATTACGTGACTCGCACAAGCATATTTTATCATGCTGGCAGCTCTAGATTATTAACTGTTGGTAATCCATATTTCAAGGTGCCCAAGGGCGGCAACGGCAGGCAGGACGTGCCCAAGGTGAGCGCATACCAATATAGAGTATTTAGGGTGCAGTTACCTGACCCAAATAAATTTGGTTTACCTGATACTAGTATTTATAATCCTGAAACACAACGTTTAGTGTGGGCCTGTGCTGGAGTGGAAATTGGCCGTGGTCAGCCTTTAGGTGTTGGCCTTAGTGGGCATCCATTTTATAATAAATTAGATGACACTGAAAGTTCCCATGCCGCCACGTCTAATGTTTCTGAGGACGTTAGGGACAATGTGTCTGTAGATTATAAGCAGACACAGTTATGTATTTTGGGCTGTGCCCCCGCCATCGGCGAGCACTGGGCCAAGGGCACCCTGTGCAAGCCCGCCCAGCTGCAGCCCGGCGACTGCCCCCCCCTGGAGCTGAAGAACACCGTTTTGGAAGATGGTGATATGGTAGATACTGGATATGGTGCCATGGACTTTAGTACATTGCAAGATACTAAATGTGAGGTACCATTGGATATTTGTCAGTCTATTTGTAAATATCCTGATTATTTACAAATGTCTGCAGATCCTTATGGGGATTCCATGTTTTTTTGCTTACGGCGTGAGCAGCTTTTTGCTAGGCATTTTTGGAATAGAGCAGGTACTATGGGTGACACTGTGCCTCAATCCTTATATATTAAAGGCACAGGTATGCGTGCTTCACCTGGCAGCTGTGTGTATTCTCCCTCTCCAAGTGGCTCTATTGTTACCTCTGACTCCCAGTTGTTTAATAAACCATATTGGTTACATAAGGCACAGGGTCATAACAATGGTGTTTGCTGGCATAATCAATTATTTGTTACTGTGGTAGATACCACTCGCAGTACCAATTTAACAATATGTGCTTCTACACAGTCTCCTGTACCTGGGCAATATGATGCTACCAAATTTAAGCAGTATAGCAGACATGTTGAGGAATATGATTTGCAGTTTATTTTTCAGTTGTGTACTATTACTTTAACTGCAGATGTTATGTCCTATATTCATAGTATGAATAGCAGTATTTTAGAGGATTGGAACTTTGGTGTTCCCCCCCCGCCAACTACTAGTTTGGTGGATACATATCGTTTTGTACAATCTGTTGCTATTGCCTGTCAAAAGGATGCTGCACCGGCTGAAAATAAGGATCCCTATGATAAGTTAAAGTTTTGGAATGTGGATTTAAAGGAAAAGTTTTCTTTAGACTTAGATCAATATCCCCTTGGACGTAAATTTTTGGTTCAGGCTGGATTGCGTCGCAAGCCCACCATAGGCCCTCGCAAACGTTCTGCTCCATCTGCCACTACGGCTTCTAAACCTGCCAAGCGTGTGCGTGTACGTGCCAGGAAGTAA

sequence 29(SEQ ID NO: 29):

ATGCGGCCTAGTGACAATACCGTATATCTTCCACCTCCTTCTGTGGCAAGAGTTGTAAATACCGATGATTACGTGACTCGCACAAGCATATTTTATCATGCTGGCAGCTCTAGATTATTAACTGTTGGTAATCCATATTTTAGGGTTCCTGCAGGTGGTGGCAATAAGCAGGATATTCCTAAGGTTTCTGCATACCAATATAGAGTATTTAGGGTGCAGTTACCTGACCCAAATAAATTTGGTTTACCTGATACTAGTATTTATAATCCTGAAACACAACGTTTAGTGTGGGCCTGTGCTGGCGTGGAGATCGGCAGGGGCCAGCCCCTGGGCGTGGGCCTGAGCGGCCACCCCCTGTACAACAAGCTGGACGACACCGAGAACAGCCACGTGGCCAGCGCCGTGGACACCAAGGACACCAGGGACAACGTGAGCGTGGACTACAAGCAGACCCAGCTGTGCATCTTGGGCTGTGCCCCCGCCATCGGCGAGCACTGGGCCAAGGGCACCCTGTGCAAGCCCGCCCAGCTGCAGCCCGGCGACTGCCCCCCCCTGGAGCTGAAGAACACCGTTTTGGAAGATGGTGATATGGTAGATACTGGATATGGTGCCATGGACTTTAGTACATTGCAAGATACTAAATGTGAGGTACCATTGGATATTTGTCAGTCTATTTGTAAATATCCTGATTATTTACAAATGTCTGCAGATCCTTATGGGGATTCCATGTTTTTTTGCTTACGGCGTGAGCAGCTTTTTGCTAGGCATTTTTGGAATAGAGCAGGTACTATGGGTGACACTGTGCCTCAATCCTTATATATTAAAGGCACAGGTATGCGTGCTTCACCTGGCAGCTGTGTGTATTCTCCCTCTCCAAGTGGCTCTATTGTTACCTCTGACTCCCAGTTGTTTAATAAACCATATTGGTTACATAAGGCACAGGGTCATAACAATGGTGTTTGCTGGCATAATCAATTATTTGTTACTGTGGTAGATACCACTCGCAGTACCAATTTAACAATATGTGCTTCTACACAGTCTCCTGTACCTGGGCAATATGATGCTACCAAATTTAAGCAGTATAGCAGACATGTTGAGGAATATGATTTGCAGTTTATTTTTCAGTTGTGTACTATTACTTTAACTGCAGATGTTATGTCCTATATTCATAGTATGAATAGCAGTATTTTAGAGGATTGGAACTTTGGTGTTCCCCCCCCGCCAACTACTAGTTTGGTGGATACATATCGTTTTGTACAATCTGTTGCTATTGCCTGTCAAAAGGATGCTGCACCGGCTGAAAATAAGGATCCCTATGATAAGTTAAAGTTTTGGAATGTGGATTTAAAGGAAAAGTTTTCTTTAGACTTAGATCAATATCCCCTTGGACGTAAATTTTTGGTTCAGGCTGGATTGCGTCGCAAGCCCACCATAGGCCCTCGCAAACGTTCTGCTCCATCTGCCACTACGGCTTCTAAACCTGCCAAGCGTGTGCGTGTACGTGCCAGGAAGTAA

Sequence 30(SEQ ID NO: 30):

ATGCGGCCTAGTGACAATACCGTATATCTTCCACCTCCTTCTGTGGCAAGAGTTGTAAATACCGATGATTACGTGACTCGCACAAGCATATTTTATCATGCTGGCAGCTCTAGATTATTAACTGTTGGTAATCCATATTTTAGGGTTCCTGCAGGTGGTGGCAATAAGCAGGATATTCCTAAGGTTTCTGCATACCAATATAGAGTATTTAGGGTGCAGTTACCTGACCCAAATAAATTTGGTTTACCTGATACTAGTATTTATAATCCTGAAACACAACGTTTAGTGTGGGCCTGTGCTGGAGTGGAAATTGGCCGTGGTCAGCCTTTAGGTGTTGGCCTTAGTGGGCATCCATTTTATAATAAATTAGATGACACTGAAAGTTCCCATGCCGCCACGTCTAATGTTTCTGAGGACGTTAGGGACAATGTGTCTGTAGATTATAAGCAGACACAGTTATGTATTTTGGGCTGTGCCCCCGCCATCGGCGAGCACTGGGCCAAGGGCACCCTGTGCAAGCCCGCCCAGCTGCAGCCCGGCGACTGCCCCCCCCTGGAGCTGAAGAACACCGTTTTGGAAGATGGTGATATGGTAGATACTGGATATGGTGCCATGGACTTTAGTACATTGCAAGATACTAAATGTGAGGTACCATTGGATATTTGTCAGTCTATTTGTAAATATCCTGATTATTTACAAATGTCTGCAGATCCTTATGGGGATTCCATGTTTTTTTGCTTACGGCGTGAGCAGCTTTTCGCCAGGCACTTCTGGAACAGGAGCGGCACCATGGGCGACCAGCTGCCCGAGAGCCTGTACATCAAGGGCACCGACATCAGGGCCAACCCCGGCAGCTACCTGTACAGCCCCAGCCCCAGCGGCTCTATTGTTACCTCTGACTCCCAGTTGTTTAATAAACCATATTGGTTACATAAGGCACAGGGTCATAACAATGGTGTTTGCTGGCATAATCAATTATTTGTTACTGTGGTAGATACCACTCGCAGTACCAATTTAACAATATGTGCTTCTACACAGTCTCCTGTACCTGGGCAATATGATGCTACCAAATTTAAGCAGTATAGCAGACATGTTGAGGAATATGATTTGCAGTTTATTTTTCAGTTGTGTACTATTACTTTAACTGCAGATGTTATGTCCTATATTCATAGTATGAATAGCAGTATTTTAGAGGATTGGAACTTTGGTGTTCCCCCCCCGCCAACTACTAGTTTGGTGGATACATATCGTTTTGTACAATCTGTTGCTATTGCCTGTCAAAAGGATGCTGCACCGGCTGAAAATAAGGATCCCTATGATAAGTTAAAGTTTTGGAATGTGGATTTAAAGGAAAAGTTTTCTTTAGACTTAGATCAATATCCCCTTGGACGTAAATTTTTGGTTCAGGCTGGATTGCGTCGCAAGCCCACCATAGGCCCTCGCAAACGTTCTGCTCCATCTGCCACTACGGCTTCTAAACCTGCCAAGCGTGTGCGTGTACGTGCCAGGAAGTAA

sequence 31(SEQ ID NO: 31):

ATGCGGCCTAGTGACAATACCGTATATCTTCCACCTCCTTCTGTGGCAAGAGTTGTAAATACCGATGATTACGTGACTCGCACAAGCATATTTTATCATGCTGGCAGCTCTAGATTATTAACTGTTGGTAATCCATATTTTAGGGTTCCTGCAGGTGGTGGCAATAAGCAGGATATTCCTAAGGTTTCTGCATACCAATATAGAGTATTTAGGGTGCAGTTACCTGACCCAAATAAATTTGGTTTACCTGATACTAGTATTTATAATCCTGAAACACAACGTTTAGTGTGGGCCTGTGCTGGAGTGGAAATTGGCCGTGGTCAGCCTTTAGGTGTTGGCCTTAGTGGGCATCCATTTTATAATAAATTAGATGACACTGAAAGTTCCCATGCCGCCACGTCTAATGTTTCTGAGGACGTTAGGGACAATGTGTCTGTAGATTATAAGCAGACACAGTTATGTATTTTGGGCTGTGCCCCCGCCATCGGCGAGCACTGGGCCAAGGGCACCCTGTGCAAGCCCGCCCAGCTGCAGCCCGGCGACTGCCCCCCCCTGGAGCTGAAGAACACCGTTTTGGAAGATGGTGATATGGTAGATACTGGATATGGTGCCATGGACTTTAGTACATTGCAAGATACTAAATGTGAGGTACCATTGGATATTTGTCAGTCTATTTGTAAATATCCTGATTATTTACAAATGTCTGCAGATCCTTATGGGGATTCCATGTTTTTTTGCTTACGGCGTGAGCAGCTTTTTGCTAGGCATTTTTGGAATAGAGCAGGTACTATGGGTGACACTGTGCCTCAATCCTTATATATTAAAGGCACAGGTATGCGTGCTTCACCTGGCAGCTGTGTGTATTCTCCCTCTCCAAGTGGCTCTATTGTTACCTCTGACTCCCAGTTGTTTAATAAACCATATTGGTTACATAAGGCACAGGGTCATAACAATGGTGTTTGCTGGCATAATCAATTATTTGTTACTGTGGTAGATACCACTCGCAGTACCAATTTAACAATATGCGCCAGCACCACCAGCAGCATCCCCAACGTGTACACCCCCACCAGCTTCAAGCAGTATAGCAGACATGTTGAGGAATATGATTTGCAGTTTATTTTTCAGTTGTGTACTATTACTTTAACTGCAGATGTTATGTCCTATATTCATAGTATGAATAGCAGTATTTTAGAGGATTGGAACTTTGGTGTTCCCCCCCCGCCAACTACTAGTTTGGTGGATACATATCGTTTTGTACAATCTGTTGCTATTGCCTGTCAAAAGGATGCTGCACCGGCTGAAAATAAGGATCCCTATGATAAGTTAAAGTTTTGGAATGTGGATTTAAAGGAAAAGTTTTCTTTAGACTTAGATCAATATCCCCTTGGACGTAAATTTTTGGTTCAGGCTGGATTGCGTCGCAAGCCCACCATAGGCCCTCGCAAACGTTCTGCTCCATCTGCCACTACGGCTTCTAAACCTGCCAAGCGTGTGCGTGTACGTGCCAGGAAGTAA

Sequence 32(SEQ ID NO: 32):

ATGCGGCCTAGTGACAATACCGTATATCTTCCACCTCCTTCTGTGGCAAGAGTTGTAAATACCGATGATTACGTGACTCGCACAAGCATATTTTATCATGCTGGCAGCTCTAGATTATTAACTGTTGGTAATCCCTACTTCAAGGTGCCCAAGGGCGGCAACGGCAGGCAGGACGTGCCCAAGGTGAGCGCCTACCAGTACAGGGTATTTAGGGTGCAGTTACCTGACCCAAATAAATTTGGTTTACCTGATACTAGTATTTATAATCCTGAAACACAACGTTTAGTGTGGGCCTGTGCTGGAGTGGAAATTGGCCGTGGTCAGCCTTTAGGTGTTGGCCTTAGTGGGCATCCATTTTATAATAAATTAGATGACACTGAAAGTTCCCATGCCGCCACGTCTAATGTTTCTGAGGACGTTAGGGACAATGTGTCTGTAGATTATAAGCAGACACAGTTATGTATTTTGGGCTGTGCCCCTGCTATTGGGGAACACTGGGCTAAAGGCACTGCTTGTAAATCGCGTCCTTTATCACAGGGCGATTGCCCCCCTTTAGAACTTAAAAACACAGTTTTGGAAGATGGTGATATGGTAGATACTGGATACGGCGCCATGGACTTCAGCACCCTGCAGGACACCAAGTGCGAGGTGCCCCTGGACATCTGCCAGAGCATCTGCAAGTACCCCGACTACCTGCAGATGAGCGCCGACCCCTACGGCGACAGCATGTTCTTCTGCCTGAGGAGGGAGCAGCTGTTCGCCAGGCACTTCTGGAACAGGGCCGGCGTGATGGGCGACACCGTGCCCACCGACCTGTACATCAAGGGCACCAGCGCCAACATGAGGGAGACCCCCGGCAGCTGCGTGTACAGCCCCAGCCCCAGCGGCAGCATCGTTACCTCTGACTCCCAGTTGTTTAATAAACCATATTGGTTACATAAGGCACAGGGTCATAACAATGGTGTTTGCTGGCATAATCAATTATTTGTTACTGTGGTAGATACCACTCGCAGTACCAATTTAACAATATGTGCTTCTACACAGTCTCCTGTACCTGGGCAATATGATGCTACCAAATTTAAGCAGTATAGCAGACATGTTGAGGAATATGATTTGCAGTTTATTTTTCAGTTGTGTACTATTACTTTAACTGCAGATGTTATGTCCTATATTCATAGTATGAATAGCAGTATTTTAGAGGATTGGAACTTTGGTGTTCCCCCCCCGCCAACTACTAGTTTGGTGGATACATATCGTTTTGTACAATCTGTTGCTATTGCCTGTCAAAAGGATGCTGCACCGGCTGAAAATAAGGATCCCTATGATAAGTTAAAGTTTTGGAATGTGGATTTAAAGGAAAAGTTTTCTTTAGACTTAGATCAATATCCCCTTGGACGTAAATTTTTGGTTCAGGCTGGATTGCGTCGCAAGCCCACCATAGGCCCTCGCAAACGTTCTGCTCCATCTGCCACTACGGCTTCTAAACCTGCCAAGCGTGTGCGTGTACGTGCCAGGAAGTAA

sequence 33(SEQ ID NO: 33):

ATGCGGCCTAGTGACAATACCGTATATCTTCCACCTCCTTCTGTGGCAAGAGTTGTAAATACCGATGATTACGTGACTCGCACAAGCATATTTTATCATGCTGGCAGCTCTAGATTATTAACTGTTGGTAATCCATATTTTAGGGTTCCTGCAGGTGGTGGCAATAAGCAGGATATTCCTAAGGTTTCTGCATACCAATATAGAGTATTTAGGGTGCAGTTACCTGACCCAAATAAATTTGGTTTACCTGATACTAGTATTTATAATCCTGAAACACAACGTTTAGTGTGGGCCTGTGCTGGCGTGGAGATCGGCAGGGGCCAGCCCCTGGGCGTGGGCCTGAGCGGCCACCCCCTGTACAACAAGCTGGACGACACCGAGAACAGCCACGTGGCCAGCGCCGTGGACACCAAGGACACCAGGGACAACGTGAGCGTGGACTACAAGCAGACCCAGCTGTGCATCTTGGGCTGTGCCCCTGCTATTGGGGAACACTGGGCTAAAGGCACTGCTTGTAAATCGCGTCCTTTATCACAGGGCGATTGCCCCCCTTTAGAACTTAAAAACACAGTTTTGGAAGATGGTGATATGGTAGATACTGGATACGGCGCCATGGACTTCAGCACCCTGCAGGACACCAAGTGCGAGGTGCCCCTGGACATCTGCCAGAGCATCTGCAAGTACCCCGACTACCTGCAGATGAGCGCCGAcCCCTACGGCGACAGCATGTTCTTCTGCCTGAGGAGGGAGCAGCTGTTCGCCAGGCACTTCTGGAACAGGGCCGGCGTGATGGGCGACACCGTGCCCACCGACCTGTACATCAAGGGCACCAGCGCCAACATGAGGGAGACCCCCGGCAGCTGCGTGTACAGCCCCAGCCCCAGCGGCAGCATCGTTACCTCTGACTCCCAGTTGTTTAATAAACCATATTGGTTACATAAGGCACAGGGTCATAACAATGGTGTTTGCTGGCATAATCAATTATTTGTTACTGTGGTAGATACCACTCGCAGTACCAATTTAACAATATGTGCTTCTACACAGTCTCCTGTACCTGGGCAATATGATGCTACCAAATTTAAGCAGTATAGCAGACATGTTGAGGAATATGATTTGCAGTTTATTTTTCAGTTGTGTACTATTACTTTAACTGCAGATGTTATGTCCTATATTCATAGTATGAATAGCAGTATTTTAGAGGATTGGAACTTTGGTGTTCCCCCCCCGCCAACTACTAGTTTGGTGGATACATATCGTTTTGTACAATCTGTTGCTATTGCCTGTCAAAAGGATGCTGCACCGGCTGAAAATAAGGATCCCTATGATAAGTTAAAGTTTTGGAATGTGGATTTAAAGGAAAAGTTTTCTTTAGACTTAGATCAATATCCCCTTGGACGTAAATTTTTGGTTCAGGCTGGATTGCGTCGCAAGCCCACCATAGGCCCTCGCAAACGTTCTGCTCCATCTGCCACTACGGCTTCTAAACCTGCCAAGCGTGTGCGTGTACGTGCCAGGAAGTAA

sequence 34(SEQ ID NO: 34):

ATGCGGCCTAGTGACAATACCGTATATCTTCCACCTCCTTCTGTGGCAAGAGTTGTAAATACCGATGATTACGTGACTCGCACAAGCATATTTTATCATGCTGGCAGCTCTAGATTATTAACTGTTGGTAATCCATATTTTAGGGTTCCTGCAGGTGGTGGCAATAAGCAGGATATTCCTAAGGTTTCTGCATACCAATATAGAGTATTTAGGGTGCAGTTACCTGACCCAAATAAATTTGGTTTACCTGATACTAGTATTTATAATCCTGAAACACAACGTTTAGTGTGGGCCTGTGCTGGAGTGGAAATTGGCCGTGGTCAGCCTTTAGGTGTTGGCCTTAGTGGGCATCCATTTTATAATAAATTAGATGACACTGAAAGTTCCCATGCCGCCACGTCTAATGTTTCTGAGGACGTTAGGGACAATGTGTCTGTAGATTATAAGCAGACACAGTTATGTATTTTGGGCTGTGCCCCCGCCATCGGCGAGCACTGGACCAAGGGCACCGCCTGCAAGCCCACCACCGTGGTGCAGGGCGACTGCCCCCCCCTGGAGCTGAAAAACACAGTTTTGGAAGATGGTGATATGGTAGATACTGGATACGGCGCCATGGACTTCAGCACCCTGCAGGACACCAAGTGCGAGGTGCCCCTGGACATCTGCCAGAGCATCTGCAAGTACCCCGACTACCTGCAGATGAGCGCCGACCCCTACGGCGACAGCATGTTCTTCTGCCTGAGGAGGGAGCAGCTGTTCGCCAGGCACTTCTGGAACAGGGCCGGCGTGATGGGCGACACCGTGCCCACCGACCTGTACATCAAGGGCACCAGCGCCAACATGAGGGAGACCCCCGGCAGCTGCGTGTACAGCCCCAGCCCCAGCGGCAGCATCGTTACCTCTGACTCCCAGTTGTTTAATAAACCATATTGGTTACATAAGGCACAGGGTCATAACAATGGTGTTTGCTGGCATAATCAATTATTTGTTACTGTGGTAGATACCACTCGCAGTACCAATTTAACAATATGTGCTTCTACACAGTCTCCTGTACCTGGGCAATATGATGCTACCAAATTTAAGCAGTATAGCAGACATGTTGAGGAATATGATTTGCAGTTTATTTTTCAGTTGTGTACTATTACTTTAACTGCAGATGTTATGTCCTATATTCATAGTATGAATAGCAGTATTTTAGAGGATTGGAACTTTGGTGTTCCCCCCCCGCCAACTACTAGTTTGGTGGATACATATCGTTTTGTACAATCTGTTGCTATTGCCTGTCAAAAGGATGCTGCACCGGCTGAAAATAAGGATCCCTATGATAAGTTAAAGTTTTGGAATGTGGATTTAAAGGAAAAGTTTTCTTTAGACTTAGATCAATATCCCCTTGGACGTAAATTTTTGGTTCAGGCTGGATTGCGTCGCAAGCCCACCATAGGCCCTCGCAAACGTTCTGCTCCATCTGCCACTACGGCTTCTAAACCTGCCAAGCGTGTGCGTGTACGTGCCAGGAAGTAA

sequence 35(SEQ ID NO: 35):

ATGCGGCCTAGTGACAATACCGTATATCTTCCACCTCCTTCTGTGGCAAGAGTTGTAAATACCGATGATTACGTGACTCGCACAAGCATATTTTATCATGCTGGCAGCTCTAGATTATTAACTGTTGGTAATCCATATTTTAGGGTTCCTGCAGGTGGTGGCAATAAGCAGGATATTCCTAAGGTTTCTGCATACCAATATAGAGTATTTAGGGTGCAGTTACCTGACCCAAATAAATTTGGTTTACCTGATACTAGTATTTATAATCCTGAAACACAACGTTTAGTGTGGGCCTGTGCTGGAGTGGAAATTGGCCGTGGTCAGCCTTTAGGTGTTGGCCTTAGTGGGCATCCATTTTATAATAAATTAGATGACACTGAAAGTTCCCATGCCGCCACGTCTAATGTTTCTGAGGACGTTAGGGACAATGTGTCTGTAGATTATAAGCAGACACAGTTATGTATTTTGGGCTGTGCCCCTGCTATTGGGGAACACTGGGCTAAAGGCACTGCTTGTAAATCGCGTCCTTTATCACAGGGCGATTGCCCCCCTTTAGAACTTAAAAACACAGTTTTGGAAGATGGTGATATGGTAGATACTGGATACGGCGCCATGGACTTCAGCACCCTGCAGGACACCAAGTGCGAGGTGCCCCTGGACATCTGCCAGAGCATCTGCAAGTACCCCGACTACCTGCAGATGAGCGCCGAcCCCTACGGCGACAGCATGTTCTTCTGCCTGAGGAGGGAGCAGCTGTTCGCCAGGCACTTCTGGAACAGGGCCGGCGTGATGGGCGACACCGTGCCCACCGACCTGTACATCAAGGGCACCAGCGCCAACATGAGGGAGACCCCCGGCAGCTGCGTGTACAGCCCCAGCCCCAGCGGCAGCATCGTTACCTCTGACTCCCAGTTGTTTAATAAACCATATTGGTTACATAAGGCACAGGGTCATAACAATGGTGTTTGCTGGCATAATCAATTATTTGTTACTGTGGTAGATACCACTCGCAGTACCAATTTAACAATATGCGCCAGCACCACCAGCAGCATCCCCAACGTGTACACCCCCACCAGCTTCAAGCAGTATAGCAGACATGTTGAGGAATATGATTTGCAGTTTATTTTTCAGTTGTGTACTATTACTTTAACTGCAGATGTTATGTCCTATATTCATAGTATGAATAGCAGTATTTTAGAGGATTGGAACTTTGGTGTTCCCCCCCCGCCAACTACTAGTTTGGTGGATACATATCGTTTTGTACAATCTGTTGCTATTGCCTGTCAAAAGGATGCTGCACCGGCTGAAAATAAGGATCCCTATGATAAGTTAAAGTTTTGGAATGTGGATTTAAAGGAAAAGTTTTCTTTAGACTTAGATCAATATCCCCTTGGACGTAAATTTTTGGTTCAGGCTGGATTGCGTCGCAAGCCCACCATAGGCCCTCGCAAACGTTCTGCTCCATCTGCCACTACGGCTTCTAAACCTGCCAAGCGTGTGCGTGTACGTGCCAGGAAGTAA

Sequence 36(SEQ ID NO: 36):

ATGCGGCCTAGTGACAATACCGTATATCTTCCACCTCCTTCTGTGGCAAGAGTTGTAAATACCGATGATTACGTGACTAGGACCAGCATCTTCTACCACGCCGGCAGCAGCAGGCTGCTGACCGTGGGCAACCCCTACTTCAGGGTGGTGCCCAACGGCGCCGGCAACAAGCAGGCCGTGCCCAAGGTGAGCGCCTACCAGTACAGGGTGTTCAGGGTGCAGTTACCTGACCCAAATAAATTTGGTTTACCTGATACTAGTATTTATAATCCTGAAACACAACGTTTAGTGTGGGCCTGTGCTGGAGTGGAAATTGGCCGTGGTCAGCCTTTAGGTGTTGGCCTTAGTGGGCATCCATTTTATAATAAATTAGATGACACTGAAAGTTCCCATGCCGCCACGTCTAATGTTTCTGAGGACGTTAGGGACAATGTGTCTGTAGATTATAAGCAGACACAGTTATGTATTTTGGGCTGTGCCCCTGCTATTGGGGAACACTGGGCTAAAGGCACTGCTTGTAAATCGCGTCCTTTATCACAGGGCGATTGCCCCCCTTTAGAACTTAAAAACACAGTTTTGGAAGATGGTGATATGGTAGATACTGGATATGGTGCCATGGACTTTAGTACATTGCAAGATACTAAATGTGAGGTACCATTGGATATTTGTCAGTCTATTTGTAAATATCCTGATTATTTACAAATGTCTGCAGATCCTTATGGGGATTCCATGTTTTTTTGCTTACGGCGTGAGCAGCTTTTTGCTAGGCATTTTTGGAATAGAGCAGGTACTATGGGTGACACTGTGCCTCAATCCTTATATATTAAAGGCACAGGTATGCGTGCTTCACCTGGCAGCTGTGTGTATTCTCCCTCTCCAAGTGGCTCTATTGTTACCTCTGACTCCCAGTTGTTTAATAAACCATATTGGTTACATAAGGCACAGGGTCATAACAATGGTGTTTGCTGGCATAATCAATTATTTGTTACTGTGGTAGATACCACTCGCAGTACCAATTTAACAATATGCGCCAGCACCACCAGCAGCATCCCCAACGTGTACACCCCCACCAGCTTCAAGCAGTATAGCAGACATGTTGAGGAATATGATTTGCAGTTTATTTTTCAGTTGTGTACTATTACTTTAACTGCAGATGTTATGTCCTATATTCATAGTATGAATAGCAGTATTTTAGAGGATTGGAACTTTGGTGTTCCCCCCCCGCCAACTACTAGTTTGGTGGATACATATCGTTTTGTACAATCTGTTGCTATTGCCTGTCAAAAGGATGCTGCACCGGCTGAAAATAAGGATCCCTATGATAAGTTAAAGTTTTGGAATGTGGATTTAAAGGAAAAGTTTTCTTTAGACTTAGATCAATATCCCCTTGGACGTAAATTTTTGGTTCAGGCTGGATTGCGTCGCAAGCCCACCATAGGCCCTCGCAAACGTTCTGCTCCATCTGCCACTACGGCTTCTAAACCTGCCAAGCGTGTGCGTGTACGTGCCAGGAAGTAA

Sequence 37(SEQ ID NO: 37):

ATGCGGCCTAGTGACAATACCGTATATCTTCCACCTCCTTCTGTGGCAAGAGTTGTAAATACCGATGATTACGTGACTCGCACAAGCATATTTTATCATGCTGGCAGCTCTAGATTATTAACTGTTGGTAATCCATATTTTAGGGTTCCTGCAGGTGGTGGCAATAAGCAGGATATTCCTAAGGTTTCTGCATACCAATATAGAGTATTTAGGGTGCAGTTACCTGACCCAAATAAATTTGGTTTACCTGATACTAGTATTTATAATCCTGAAACACAACGTTTAGTGTGGGCCTGTGCTGGAGTGGAGATCGGCAGGGGCCAGCCCCTGGGCATCGGCCTGAGCGGCCACCCCTTCTACAACAAGCTGGACGACACCGAGAGCGCCCACGCCGCCACCGCCGTGATCACCCAGGACGTGAGGGACAACGTGAGCGTGGACTACAAGCAGACCCAGCTGTGCATCCTGGGCTGCGCCCCTGCTATTGGGGAACACTGGGCTAAAGGCACTGCTTGTAAATCGCGTCCTTTATCACAGGGCGATTGCCCCCCTTTAGAACTTAAAAACACAGTTTTGGAAGATGGTGATATGGTAGATACTGGATATGGTGCCATGGACTTTAGTACATTGCAAGATACTAAATGTGAGGTACCATTGGATATTTGTCAGTCTATTTGTAAATATCCTGATTATTTACAAATGTCTGCAGATCCTTATGGGGATTCCATGTTTTTTTGCTTACGGCGTGAGCAGCTTTTTGCTAGGCATTTTTGGAATAGAGCAGGTACTATGGGTGACACTGTGCCTCAATCCTTATATATTAAAGGCACAGGTATGCGTGCTTCACCTGGCAGCTGTGTGTATTCTCCCTCTCCAAGTGGCTCTATTGTTACCTCTGACTCCCAGTTGTTTAATAAACCATATTGGTTACATAAGGCACAGGGTCATAACAATGGTGTTTGCTGGCATAATCAATTATTTGTTACTGTGGTAGATACCACTCGCAGTACCAATTTAACAATATGCGCCAGCACCACCAGCAGCATCCCCAACGTGTACACCCCCACCAGCTTCAAGCAGTATAGCAGACATGTTGAGGAATATGATTTGCAGTTTATTTTTCAGTTGTGTACTATTACTTTAACTGCAGATGTTATGTCCTATATTCATAGTATGAATAGCAGTATTTTAGAGGATTGGAACTTTGGTGTTCCCCCCCCGCCAACTACTAGTTTGGTGGATACATATCGTTTTGTACAATCTGTTGCTATTGCCTGTCAAAAGGATGCTGCACCGGCTGAAAATAAGGATCCCTATGATAAGTTAAAGTTTTGGAATGTGGATTTAAAGGAAAAGTTTTCTTTAGACTTAGATCAATATCCCCTTGGACGTAAATTTTTGGTTCAGGCTGGATTGCGTCGCAAGCCCACCATAGGCCCTCGCAAACGTTCTGCTCCATCTGCCACTACGGCTTCTAAACCTGCCAAGCGTGTGCGTGTACGTGCCAGGAAGTAA

Sequence 38(SEQ ID NO: 38):

ATGCGGCCTAGTGACAATACCGTATATCTTCCACCTCCTTCTGTGGCAAGAGTTGTAAATACCGATGATTACGTGACTAGGACCAGCATCTTCTACCACGCCGGCAGCAGCAGGCTGCTGACCGTGGGCAACCCCTACTTCAGGGTGGTGCCCAACGGCGCCGGCAACAAGCAGGCCGTGCCCAAGGTGAGCGCCTACCAGTACAGGGTGTTCAGGGTGCAGTTACCTGACCCAAATAAATTTGGTTTACCTGATACTAGTATTTATAATCCTGAAACACAACGTTTAGTGTGGGCCTGTGCTGGAGTGGAAATTGGCCGTGGTCAGCCTTTAGGTGTTGGCCTTAGTGGGCATCCATTTTATAATAAATTAGATGACACTGAAAGTTCCCATGCCGCCACGTCTAATGTTTCTGAGGACGTTAGGGACAATGTGTCTGTAGATTATAAGCAGACACAGTTATGTATTTTGGGCTGTGCCCCCGCCATCGGCGAGCACTGGGCCAAGGGCACCCTGTGCAAGCCCGCCCAGCTGCAGCCCGGCGACTGCCCCCCCCTGGAGCTGAAGAACACCGTTTTGGAAGATGGTGATATGGTAGATACTGGATATGGTGCCATGGACTTTAGTACATTGCAAGATACTAAATGTGAGGTACCATTGGATATTTGTCAGTCTATTTGTAAATATCCTGATTATTTACAAATGTCTGCAGATCCTTATGGGGATTCCATGTTTTTTTGCTTACGGCGTGAGCAGCTTTTTGCTAGGCATTTTTGGAATAGAGCAGGTACTATGGGTGACACTGTGCCTCAATCCTTATATATTAAAGGCACAGGTATGCGTGCTTCACCTGGCAGCTGTGTGTATTCTCCCTCTCCAAGTGGCTCTATTGTTACCTCTGACTCCCAGTTGTTTAATAAACCATATTGGTTACATAAGGCACAGGGTCATAACAATGGTGTTTGCTGGCATAATCAATTATTTGTTACTGTGGTAGATACCACTCGCAGTACCAATTTAACAATATGCGCCAGCACCACCAGCAGCATCCCCAACGTGTACACCCCCACCAGCTTCAAGCAGTATAGCAGACATGTTGAGGAATATGATTTGCAGTTTATTTTTCAGTTGTGTACTATTACTTTAACTGCAGATGTTATGTCCTATATTCATAGTATGAATAGCAGTATTTTAGAGGATTGGAACTTTGGTGTTCCCCCCCCGCCAACTACTAGTTTGGTGGATACATATCGTTTTGTACAATCTGTTGCTATTGCCTGTCAAAAGGATGCTGCACCGGCTGAAAATAAGGATCCCTATGATAAGTTAAAGTTTTGGAATGTGGATTTAAAGGAAAAGTTTTCTTTAGACTTAGATCAATATCCCCTTGGACGTAAATTTTTGGTTCAGGCTGGATTGCGTCGCAAGCCCACCATAGGCCCTCGCAAACGTTCTGCTCCATCTGCCACTACGGCTTCTAAACCTGCCAAGCGTGTGCGTGTACGTGCCAGGAAGTAA

sequence 39(SEQ ID NO: 39):

VPNGAGNKQAV

Sequence 40(SEQ ID NO: 40):

IGLSGHPFYNKLDDTESAHAATAVITQ

Sequence 41(SEQ ID NO: 41):

LCKPAQLQP

Sequence 42(SEQ ID NO: 42):

VMGDTVPTDLYIKGTSANMRET

Sequence 43(SEQ ID NO: 43):

KVPKGGNGRQDV

Sequence 44(SEQ ID NO: 44):

TSSIPNVYTPTS

Sequence 109(SEQ ID NO: 109):

NPVPSTYDP

Sequence 110(SEQ ID NO: 110):

LYNKLDDTENSHVASAVDTKDT

sequence 111(SEQ ID NO: 111):

SGTMGDQLPESLYIKGTDIRANPGSYL

Sequence 112(SEQ ID NO: 112):

TKGTACKPTTVV

sequence 113(SEQ ID NO: 113):

MRPSDNTVYLPPPSVARVVNTDDYVTRTSIFYHAGSSRLLTVGNPYFRVPAGGGNKQDIPKVSAYQYRVFRVQLPDPNKFGLPDTSIYNPETQRLVWACAGVEIGRGQPLGVGLSGHPFYNKLDDTESSHAATSNVSEDVRDNVSVDYKQTQLCILGCAPAIGEHWAKGTACKSRPLSQGDCPPLELKNTVLEDGDMVDTGYGAMDFSTLQDTKCEVPLDICQSICKYPDYLQMSADPYGDSMFFCLRREQLFARHFWNRAGTMGDTVPQSLYIKGTGMRASPGSCVYSPSPSGSIVTSDSQLFNKPYWLHKAQGHNNGVCWHNQLFVTVVDTTRSTNLTICASTQSPVPGQYDATKFKQYSRHVEEYDLQFIFQLCTITLTADVMSYIHSMNSSILEDWNFGVPPPPTTSLVDTYRFVQSVAIACQKDAAPAENKDPYDKLKFWNVDLKEKFSLDLDQYPLGRKFLVQAGLRRKPTIGPRKRSAPSATTASKPAKRVRVRARK

Sequence 114(SEQ ID NO: 114):

ATGCGGCCTAGTGACAATACCGTATATCTTCCACCTCCTTCTGTGGCAAGAGTTGTAAATACCGATGATTACGTGACTCGCACAAGCATATTTTATCATGCTGGCAGCTCTAGATTATTAACTGTTGGTAATCCATATTTTAGGGTTCCTGCAGGTGGTGGCAATAAGCAGGATATTCCTAAGGTTTCTGCATACCAATATAGAGTATTTAGGGTGCAGTTACCTGACCCAAATAAATTTGGTTTACCTGATACTAGTATTTATAATCCTGAAACACAACGTTTAGTGTGGGCCTGTGCTGGAGTGGAAATTGGCCGTGGTCAGCCTTTAGGTGTTGGCCTTAGTGGGCATCCATTTTATAATAAATTAGATGACACTGAAAGTTCCCATGCCGCCACGTCTAATGTTTCTGAGGACGTTAGGGACAATGTGTCTGTAGATTATAAGCAGACACAGTTATGTATTTTGGGCTGTGCCCCTGCTATTGGGGAACACTGGGCTAAAGGCACTGCTTGTAAATCGCGTCCTTTATCACAGGGCGATTGCCCCCCTTTAGAACTTAAAAACACAGTTTTGGAAGATGGTGATATGGTAGATACTGGATATGGTGCCATGGACTTTAGTACATTGCAAGATACTAAATGTGAGGTACCATTGGATATTTGTCAGTCTATTTGTAAATATCCTGATTATTTACAAATGTCTGCAGATCCTTATGGGGATTCCATGTTTTTTTGCTTACGGCGTGAGCAGCTTTTTGCTAGGCATTTTTGGAATAGAGCAGGTACTATGGGTGACACTGTGCCTCAATCcTTATATATTAAAGGCACAGGTATGCGTGCTTCACCTGGCAGCTGTGTGTATTCTCCCTCTCCAAGTGGCTCTATTGTTACCTCTGACTCCCAGTTGTTTAATAAACCATATTGGTTACATAAGGCACAGGGTCATAACAATGGTGTTTGCTGGCATAATCAATTATTTGTTACTGTGGTAGATACCACTCGCAGTACCAATTTAACAATATGTGCTTCTACACAGTCTCCTGTACCTGGGCAATATGATGCTACCAAATTTAAGCAGTATAGCAGACATGTTGAGGAATATGATTTGCAGTTTATTTTTCAGTTGTGTACTATTACTTTAACTGCAGATGTTATGTCCTATATTCATAGTATGAATAGCAGTATTTTAGAGGATTGGAACTTTGGTGTTCCCCCCCCGCCAACTACTAGTTTGGTGGATACATATCGTTTTGTACAATCTGTTGCTATTGCCTGTCAAAAGGATGCTGCACCGGCTGAAAATAAGGATCCCTATGATAAGTTAAAGTTTTGGAATGTGGATTTAAAGGAAAAGTTTTCTTTAGACTTAGATCAATATCCCCTTGGACGTAAATTTTTGGTTCAGGCTGGATTGCGTCGCAAGCCCACCATAGGCCCTCGCAAACGTTCTGCTCCATCTGCCACTACGGCTTCTAAACCTGCCAAGCGTGTGCGTGTACGTGCCAGGAAGTAA

sequence 115(SEQ ID NO: 115):

MALWRPSDSTVYLPPPSVARVVSTDDYVSRTSIFYHAGSSRLLTVGNPYFRVVPNGAGNKQAVPKVSAYQYRVFRVALPDPNKFGLPDSTIYNPETQRLVWACVGMEIGRGQPLGIGLSGHPFYNKLDDTESAHAATAVITQDVRDNVSVDYKQTQLCILGCVPAIGEHWAKGTLCKPAQLQPGDCPPLELKNTIIEDGDMVDTGYGAMDFSTLQDTKCEVPLDICQSICKYPDYLQMSADPYGDSMFFCLRREQLFARHFWNRAGVMGDTVPTDLYIKGTSANMRETPGSCVYSPSPSGSIITSDSQLFNKPYWLHKAQGHNNGICWHNQLFVTVVDTTRSTNLTLCASTQNPVPSTYDPTKFKQYSRHVEEYDLQFIFQLCTITLTAEVMSYIHSMNSSILENWNFGVPPPPTTSLVDTYRFVQSVAVTCQKDTTPPEKQDPYDKLKFWTVDLKEKFSSDLDQYPLGRKFLVQAGLRRRPTIGPRKRPAASTSTASTASRPAKRVRIRSKK

Sequence 116(SEQ ID NO: 116):

ATGGCCCTGTGGAGGCCCAGCGACAGCACCGTGTACCTGCCCCCCCCCAGCGTGGCCAGGGTGGTGAGCACCGACGACTACGTGAGCAGGACCAGCATCTTCTACCACGCCGGCAGCAGCAGGCTGCTGACCGTGGGCAACCCCTACTTCAGGGTGGTGCCCAACGGCGCCGGCAACAAGCAGGCCGTGCCCAAGGTGAGCGCCTACCAGTACAGGGTGTTCAGGGTGGCCCTGCCCGACCCCAACAAGTTCGGCCTGCCCGACAGCACCATCTACAACCCCGAGACCCAGAGGCTGGTGTGGGCCTGCGTGGGCATGGAGATCGGCAGGGGCCAGCCCCTGGGCATCGGCCTGAGCGGCCACCCCTTCTACAACAAGCTGGACGACACCGAGAGCGCCCACGCCGCCACCGCCGTGATCACCCAGGACGTGAGGGACAACGTGAGCGTGGACTACAAGCAGACCCAGCTGTGCATCCTGGGCTGCGTGCCCGCCATCGGCGAGCACTGGGCCAAGGGCACCCTGTGCAAGCCCGCCCAGCTGCAGCCCGGCGACTGCCCCCCCCTGGAGCTGAAGAACACCATCATCGAGGACGGCGACATGGTGGACACCGGCTACGGCGCCATGGACTTCAGCACCCTGCAGGACACCAAGTGCGAGGTGCCCCTGGACATCTGCCAGAGCATCTGCAAGTACCCCGACTACCTGCAGATGAGCGCCGACCCCTACGGCGACAGCATGTTCTTCTGCCTGAGGAGGGAGCAGCTGTTCGCCAGGCACTTCTGGAACAGGGCCGGCGTGATGGGCGACACCGTGCCCACCGACCTGTACATCAAGGGCACCAGCGCCAACATGAGGGAGACCCCCGGCAGCTGCGTGTACAGCCCCAGCCCCAGCGGCAGCATCATCACCAGCGACAGCCAGCTGTTCAACAAGCCCTACTGGCTGCACAAGGCCCAGGGCCACAACAACGGCATCTGCTGGCACAACCAGCTGTTCGTGACCGTGGTGGACACCACCAGGAGCACCAACCTGACCCTGTGCGCCAGCACCCAGAACCCCGTGCCCAGCACCTACGACCCCACCAAGTTCAAGCAGTACAGCAGGCACGTGGAGGAGTACGACCTGCAGTTCATCTTCCAGCTGTGCACCATCACCCTGACCGCCGAGGTGATGAGCTACATCCACAGCATGAACAGCAGCATCCTGGAGAACTGGAACTTCGGCGTGCCCCCCCCCCCCACCACCAGCCTGGTGGACACCTACAGGTTCGTGCAGAGCGTGGCCGTGACCTGCCAGAAGGACACCACCCCCCCCGAGAAGCAGGACCCCTACGACAAGCTGAAGTTCTGGACCGTGGACCTGAAGGAGAAGTTCAGCAGCGACCTGGACCAGTACCCCCTGGGCAGGAAGTTCCTGGTGCAGGCCGGCCTGAGGAGGAGGCCCACCATCGGCCCCAGGAAGAGGCCCGCCGCCAGCACCAGCACCGCCAGCACCGCCAGCAGGCCCGCCAAGAGGGTGAGGATCAGGAGCAAGAAGTGA

Detailed Description

The invention will now be described with reference to the following examples, which are intended to illustrate the invention, but not to limit it.

Unless otherwise indicated, the molecular biological experimental methods and immunoassay methods used in the present invention are essentially described by reference to j.sambrook et al, molecular cloning: a laboratory manual, 2 nd edition, cold spring harbor laboratory Press, 1989, and F.M. Ausubel et al, eds. molecular biology laboratory Manual, 3 rd edition, John Wiley & Sons, Inc., 1995; the use of restriction enzymes follows the conditions recommended by the product manufacturer. The examples are given by way of illustration and are not intended to limit the scope of the invention as claimed.