Antigenic variants of varicella zoster virus and uses thereof

Abstract

The present invention relates to an antigen variant and its use, wherein the antigen variant is a protein in the surface protein (gE) of varicella zoster virus, and the protein exhibits high expression level and high immunogenicity. Therefore, when the antigen variant is used as a vaccine composition, the vaccine composition has more excellent safety compared with a live virus vaccine, and the expression level of the antigen variant in host cells is higher than that of other antigens, so it can be used as a vaccine for preventing or treating chickenpox or herpes zoster caused by varicella zoster virus.

Description

Technical Field

[0001] The present invention relates to varicella-zoster virus antigen variants and uses thereof, and more specifically, to varicella-zoster virus antigen variants with high expression levels and high immunogenicity, which are selected from varicella-zoster virus surface protein (gE) antigen variants, and a vaccine composition for preventing or treating chickenpox or herpes zoster, which comprises the varicella-zoster virus antigen variant as an active ingredient. Background Art

[0002] Varicella zoster virus (VZV) is a virus that causes chickenpox, primarily in children and adolescents. Once infection occurs, VZV remains dormant in sensory root and cranial nerve ganglion cells for several years and reactivates when immunity is reduced to cause herpes zoster in adulthood. Varicella is highly contagious. Once infection occurs, it causes a vesicular rash over the body, accompanied by fever and malaise. In most normal children, varicella rarely develops into a serious condition and ultimately becomes a self-limited illness. However, a number of cases of varicella developing into severe symptoms are known to occur in patients undergoing organ transplantation or chemotherapy (Adriana Weinberg et al., Journal of Infectious Diseases, 200(7):1068, 2009; Judith Breuer et al., Vaccine Expert Reviews, 2017, DOI: 10.1080 / 14760584.2017.1394843).

[0003] The initial symptoms of shingles are general aches and pains, such as pain all over the body, or severe itching, tingling and burning sensations, accompanied by sharp pain sensations as if being stabbed by a knife. Shingles is a disease in which blisters appear after a few days, and the pain increases as the skin damage increases, and elderly patients often complain of more severe pain. Even in cases where shingles is cured, neuralgia may be left as a sequelae. It is well known that in people aged 60 or above, neuralgia may prevent them from sleeping, make them complain of chronic fatigue, feel severe pain even at the slightest touch or friction, and even lead to depression, although this neuralgia is relatively rare in adults aged 40 or below.

[0004] Representative preventive vaccines against varicella include products such as VARIVAX (Merck) and VARILRIX (GlaxoSmithKline Biologicals), which are developed using the Oka strain, an attenuated strain developed in 1970. In South Korea, products such as Suduvax (Green Cross), produced using the MAV / 06 strain developed in 1980, are commercially available. The commercially available live vaccines in question show an average protective efficacy of 80%, which means that even after vaccination, 20% of vaccine recipients develop an infection. Stability problems are constantly pointed out, such as the occurrence of varicella and herpes zoster caused by the live virus contained in the vaccine.

[0005] ZOSTAVAX (Merck) is a live attenuated vaccine produced using the Oka strain and developed as a preventive vaccine against herpes zoster. Because the vaccine contains a large amount of virus, it has been approved and marketed in the United States and South Korea for use in adults 50 years of age or older, not in children or adolescents. More recently, GlaxoSmithKline Biologicals has developed a vaccine consisting of the viral surface protein (gE) and an adjuvant that is intended for use in adults 50 years of age or older and has been shown to be preventive in clinical trials (U.S. Patent No. 7,939,084, granted January 7, 2011).

[0006] In the early stage of vaccine development, attenuated live cells or dead cells were mainly used as antigens. However, due to safety issues and immunosuppressive substances present in pathogens, the development of such antigens is turning to the development of protein antigens, which have clear structures and compositions and can establish the necessary immunity for disease defense.

[0007] Therefore, the inventors have developed a protein antigen with a high expression level in host cells through in-depth efforts, which increases the yield of varicella zoster virus surface protein (gE) antigen without affecting immunogenicity. As a result, the inventors produced varicella zoster virus surface protein antigen variants of different lengths and measured their expression levels, thereby identifying specific amino acid sequences with high expression levels in the antigen variants. Thus, the present invention was completed.

[0008] Invention Disclosure

[0009] Technical issues

[0010] The object of the present invention is to provide a specific antigen variant with high expression level and high immunogenicity, which is selected from varicella zoster virus surface protein (gE) antigen variants.

[0011] Another object of the present invention is to provide a gene encoding an antigen variant, a recombinant vector comprising the gene, and a host cell transformed with the recombinant vector.

[0012] Another object of the present invention is to provide a vaccine composition for preventing or treating varicella or herpes zoster, which comprises the antigen variant as an active ingredient.

[0013] Still another object of the present invention is to provide a method for preventing or treating varicella or herpes zoster using a vaccine composition comprising the antigen variant as an active ingredient.

[0014] Another object of the present invention is to provide use of a vaccine composition comprising the antigen variant as an active ingredient for preventing or treating varicella or herpes zoster.

[0015] Another object of the present invention is to provide use of a vaccine composition comprising an antigen variant as an active ingredient for preparing a medicament for preventing or treating varicella or herpes zoster.

[0016] Solutions to technical problems

[0017] In order to achieve the above-mentioned purpose, the present invention provides a varicella-zoster virus surface protein antigen variant, characterized in that the antigen variant includes a variant with a carboxyl-terminal truncation of any amino acid residue selected from the following groups: an amino acid sequence consisting of amino acid residues 525 to 543 in the varicella-zoster virus surface protein (gE) antigen shown in SEQ ID NO: 1.

[0018] In addition, the present invention provides a gene encoding an antigen variant, a recombinant vector comprising the gene, and a host cell transformed with the recombinant vector.

[0019] In addition, the present invention provides a vaccine composition for preventing or treating varicella or herpes zoster, which comprises an antigen variant as an active ingredient.

[0020] In addition, in the present invention, a method for preventing or treating varicella or herpes zoster using a vaccine composition comprising the antigen variant as an active ingredient is provided.

[0021] In addition, the present invention provides use of a vaccine composition comprising the antigen variant as an active ingredient for preventing or treating varicella or herpes zoster.

[0022] In addition, the present invention provides the use of a vaccine composition comprising an antigen variant as an active ingredient for preparing a medicament for preventing or treating varicella or herpes zoster. <110> Makigan Life Science Research Institute <120> Antigenic variants of varicella zoster virus and uses thereof <130> PP-B2204 <150> KR 2018-0058219 <151> 2018-05-23 <160> 24 <170> KoPatentIn 3.0 <210> 1 <211> 623 <212> PRT <213> Artificial Sequence <220> <223> Artificial Sequence <400> 1 Met Gly Thr Val Asn Lys Pro Val Val Gly Val Leu Met Gly Phe Gly 1 5 10 15 Ile Ile Thr Gly Thr Leu Arg Ile Thr Asn Pro Val Arg Ala Ser Val 20 25 30 Leu Arg Tyr Asp Asp Phe His Xaa Asp Glu Asp Lys Leu Asp Thr Asn 35 40 45 Ser Val Tyr Glu Pro Tyr Tyr His Ser Asp His Ala Glu Ser Ser Trp 50 55 60 Val Asn Arg Gly Glu Ser Ser Arg Lys Ala Tyr Asp His Asn Ser Pro 65 70 75 80 Tyr Ile Trp Pro Arg Asn Asp Tyr Asp Gly Phe Leu Glu Asn Ala His 85 90 95 Glu His His Gly Val Tyr Asn Gln Gly Arg Gly Ile Asp Ser Gly Glu 100 105 110 Arg Leu Met Gln Pro Thr Gln Met Ser Ala Gln Glu Asp Leu Gly Asp 115 120 125 Asp Thr Gly Ile His Val Ile Pro Thr Leu Asn Gly Asp Asp Arg His 130 135 140 Lys Ile Val Asn Val Asp Gln Arg Gln Tyr Gly Asp Val Phe Lys Gly 145 150 155 160 Asp Leu Asn Pro Lys Pro Gln Gly Gln Arg Leu Ile Glu Val Ser Val 165 170 175 Glu Glu Asn His Pro Phe Thr Leu Arg Ala Pro Ile Gln Arg Ile Tyr 180 185 190 Gly Val Arg Tyr Thr Glu Thr Trp Ser Phe Leu Pro Ser Leu Thr Cys 195 200 205 Thr Gly Asp Ala Ala Pro Ala Ile Gln His Ile Cys Leu Lys His Thr 210 215 220 Thr Cys Phe Gln Asp Val Val Val Asp Val Asp Cys Ala Glu Asn Thr 225 230 235 240 Lys Glu Asp Gln Leu Ala Glu Ile Ser Tyr Arg Phe Gln Gly Lys Lys 245 250 255 Glu Ala Asp Gln Pro Trp Ile Val Val Asn Thr Ser Thr Leu Phe Asp 260 265 270 Glu Leu Glu Leu Asp Pro Pro Glu Ile Glu Pro Gly Val Leu Lys Val 275 280 285 Leu Arg Thr Glu Lys Gln Tyr Leu Gly Val Tyr Ile Trp Asn Met Arg 290 295 300 Gly Ser Asp Gly Thr Ser Thr Tyr Ala Thr Phe Leu Val Thr Trp Lys 305 310 315 320 Gly Asp Glu Lys Thr Arg Asn Pro Thr Pro Ala Val Thr Pro Gln Pro 325 330 335 Arg Gly Ala Glu Phe His Met Trp Asn Tyr His Ser His Val Phe Ser 340 345 350 Val Gly Asp Thr Phe Ser Leu Ala Met His Leu Gln Tyr Lys Ile His 355 360 365 Glu Ala Pro Phe Asp Leu Leu Leu Glu Trp Leu Tyr Val Pro Ile Asp 370 375 380 Pro Thr Cys Gln Pro Met Arg Leu Tyr Ser Thr Cys Leu Tyr His Pro 385 390 395 400 Asn Ala Pro Gln Cys Leu Ser His Met Asn Ser Gly Cys Thr Phe Thr 405 410 415 Ser Pro His Leu Ala Gln Arg Val Ala Ser Thr Val Tyr Gln Asn Cys 420 425 430 Glu His Ala Asp Asn Tyr Thr Ala Tyr Cys Leu Gly Ile Ser His Met 435 440 445 Glu Pro Ser Phe Gly Leu Ile Leu His Asp Gly Gly Thr Thr Leu Lys 450 455 460 Phe Val Asp Thr Pro Glu Ser Leu Ser Gly Leu Tyr Val Phe Val Val 465 470 475 480 Tyr Phe Asn Gly His Val Glu Ala Val Ala Tyr Thr Val Val Ser Thr 485 490 495 Val Asp His Phe Val Asn Ala Ile Glu Glu Arg Gly Phe Pro Pro Thr 500 505 510 Ala Gly Gln Pro Pro Ala Thr Thr Lys Pro Lys Glu Ile Thr Pro Val 515 520 525 Asn Pro Gly Thr Ser Pro Leu Xaa Arg Tyr Ala Ala Trp Thr Gly Gly 530 535 540 Leu Ala Ala Val Val Leu Leu Cys Leu Val Ile Phe Leu Ile Cys Thr 545 550 555 560 Ala Lys Arg Met Arg Val Lys Ala Tyr Arg Val Asp Lys Ser Pro Tyr 565 570 575 Asn Gln Ser Met Tyr Tyr Ala Gly Leu Pro Val Asp Asp Phe Glu Asp 580 585 590 Ser Glu Ser Thr Asp Thr Glu Glu Glu Phe Gly Asn Ala Ile Gly Gly 595 600 605 Ser His Gly Gly Ser Ser Tyr Thr Val Tyr Ile Asp Lys Thr Arg 610 615 620 <210> 2 <211> 534 <212> PRT <213> Artificial Sequence <220> <223> Artificial Sequence <400> 2 Met Gly Thr Val Asn Lys Pro Val Val Gly Val Leu Met Gly Phe Gly 1 5 10 15 Ile Ile Thr Gly Thr Leu Arg Ile Thr Asn Pro Val Arg Ala Ser Val 20 25 30 Leu Arg Tyr Asp Asp Phe His Thr Asp Glu Asp Lys Leu Asp Thr Asn 35 40 45 Ser Val Tyr Glu Pro Tyr Tyr His Ser Asp His Ala Glu Ser Ser Trp 50 55 60 Val Asn Arg Gly Glu Ser Ser Arg Lys Ala Tyr Asp His Asn Ser Pro 65 70 75 80 Tyr Ile Trp Pro Arg Asn Asp Tyr Asp Gly Phe Leu Glu Asn Ala His 85 90 95 Glu His His Gly Val Tyr Asn Gln Gly Arg Gly Ile Asp Ser Gly Glu 100 105 110 Arg Leu Met Gln Pro Thr Gln Met Ser Ala Gln Glu Asp Leu Gly Asp 115 120 125 Asp Thr Gly Ile His Val Ile Pro Thr Leu Asn Gly Asp Asp Arg His 130 135 140 Lys Ile Val Asn Val Asp Gln Arg Gln Tyr Gly Asp Val Phe Lys Gly 145 150 155 160 Asp Leu Asn Pro Lys Pro Gln Gly Gln Arg Leu Ile Glu Val Ser Val 165 170 175 Glu Glu Asn His Pro Phe Thr Leu Arg Ala Pro Ile Gln Arg Ile Tyr 180 185 190 Gly Val Arg Tyr Thr Glu Thr Trp Ser Phe Leu Pro Ser Leu Thr Cys 195 200 205 Thr Gly Asp Ala Ala Pro Ala Ile Gln His Ile Cys Leu Lys His Thr 210 215 220 Thr Cys Phe Gln Asp Val Val Val Asp Val Asp Cys Ala Glu Asn Thr 225 230 235 240 Lys Glu Asp Gln Leu Ala Glu Ile Ser Tyr Arg Phe Gln Gly Lys Lys 245 250 255 Glu Ala Asp Gln Pro Trp Ile Val Val Asn Thr Ser Thr Leu Phe Asp 260 265 270 Glu Leu Glu Leu Asp Pro Pro Glu Ile Glu Pro Gly Val Leu Lys Val 275 280 285 Leu Arg Thr Glu Lys Gln Tyr Leu Gly Val Tyr Ile Trp Asn Met Arg 290 295 300 Gly Ser Asp Gly Thr Ser Thr Tyr Ala Thr Phe Leu Val Thr Trp Lys 305 310 315 320 Gly Asp Glu Lys Thr Arg Asn Pro Thr Pro Ala Val Thr Pro Gln Pro 325 330 335 Arg Gly Ala Glu Phe His Met Trp Asn Tyr His Ser His Val Phe Ser 340 345 350 Val Gly Asp Thr Phe Ser Leu Ala Met His Leu Gln Tyr Lys Ile His 355 360 365 Glu Ala Pro Phe Asp Leu Leu Leu Glu Trp Leu Tyr Val Pro Ile Asp 370 375 380 Pro Thr Cys Gln Pro Met Arg Leu Tyr Ser Thr Cys Leu Tyr His Pro 385 390 395 400 Asn Ala Pro Gln Cys Leu Ser His Met Asn Ser Gly Cys Thr Phe Thr 405 410 415 Ser Pro His Leu Ala Gln Arg Val Ala Ser Thr Val Tyr Gln Asn Cys 420 425 430 Glu His Ala Asp Asn Tyr Thr Ala Tyr Cys Leu Gly Ile Ser His Met 435 440 445 Glu Pro Ser Phe Gly Leu Ile Leu His Asp Gly Gly Thr Thr Leu Lys 450 455 460 Phe Val Asp Thr Pro Glu Ser Leu Ser Gly Leu Tyr Val Phe Val Val 465 470 475 480 Tyr Phe Asn Gly His Val Glu Ala Val Ala Tyr Thr Val Val Ser Thr 485 490 495 Val Asp His Phe Val Asn Ala Ile Glu Glu Arg Gly Phe Pro Pro Thr 500 505 510 Ala Gly Gln Pro Pro Ala Thr Thr Lys Pro Lys Glu Ile Thr Pro Val 515 520 525 Asn Pro Gly Thr Ser Pro 530 <210> 3 <211> 535 <212> PRT <213> Artificial Sequence <220> <223> Artificial Sequence <400> 3 Met Gly Thr Val Asn Lys Pro Val Val Gly Val Leu Met Gly Phe Gly 1 5 10 15 Ile Ile Thr Gly Thr Leu Arg Ile Thr Asn Pro Val Arg Ala Ser Val 20 25 30 Leu Arg Tyr Asp Asp Phe His Thr Asp Glu Asp Lys Leu Asp Thr Asn 35 40 45 Ser Val Tyr Glu Pro Tyr Tyr His Ser Asp His Ala Glu Ser Ser Trp 50 55 60 Val Asn Arg Gly Glu Ser Ser Arg Lys Ala Tyr Asp His Asn Ser Pro 65 70 75 80 Tyr Ile Trp Pro Arg Asn Asp Tyr Asp Gly Phe Leu Glu Asn Ala His 85 90 95 Glu His His Gly Val Tyr Asn Gln Gly Arg Gly Ile Asp Ser Gly Glu 100 105 110 Arg Leu Met Gln Pro Thr Gln Met Ser Ala Gln Glu Asp Leu Gly Asp 115 120 125 Asp Thr Gly Ile His Val Ile Pro Thr Leu Asn Gly Asp Asp Arg His 130 135 140 Lys Ile Val Asn Val Asp Gln Arg Gln Tyr Gly Asp Val Phe Lys Gly 145 150 155 160 Asp Leu Asn Pro Lys Pro Gln Gly Gln Arg Leu Ile Glu Val Ser Val 165 170 175 Glu Glu Asn His Pro Phe Thr Leu Arg Ala Pro Ile Gln Arg Ile Tyr 180 185 190 Gly Val Arg Tyr Thr Glu Thr Trp Ser Phe Leu Pro Ser Leu Thr Cys 195 200 205 Thr Gly Asp Ala Ala Pro Ala Ile Gln His Ile Cys Leu Lys His Thr 210 215 220 Thr Cys Phe Gln Asp Val Val Val Asp Val Asp Cys Ala Glu Asn Thr 225 230 235 240 Lys Glu Asp Gln Leu Ala Glu Ile Ser Tyr Arg Phe Gln Gly Lys Lys 245 250 255 Glu Ala Asp Gln Pro Trp Ile Val Val Asn Thr Ser Thr Leu Phe Asp 260 265 270 Glu Leu Glu Leu Asp Pro Pro Glu Ile Glu Pro Gly Val Leu Lys Val 275 280 285 Leu Arg Thr Glu Lys Gln Tyr Leu Gly Val Tyr Ile Trp Asn Met Arg 290 295 300 Gly Ser Asp Gly Thr Ser Thr Tyr Ala Thr Phe Leu Val Thr Trp Lys 305 310 315 320 Gly Asp Glu Lys Thr Arg Asn Pro Thr Pro Ala Val Thr Pro Gln Pro 325 330 335 Arg Gly Ala Glu Phe His Met Trp Asn Tyr His Ser His Val Phe Ser 340 345 350 Val Gly Asp Thr Phe Ser Leu Ala Met His Leu Gln Tyr Lys Ile His 355 360 365 Glu Ala Pro Phe Asp Leu Leu Leu Glu Trp Leu Tyr Val Pro Ile Asp 370 375 380 Pro Thr Cys Gln Pro Met Arg Leu Tyr Ser Thr Cys Leu Tyr His Pro 385 390 395 400 Asn Ala Pro Gln Cys Leu Ser His Met Asn Ser Gly Cys Thr Phe Thr 405 410 415 Ser Pro His Leu Ala Gln Arg Val Ala Ser Thr Val Tyr Gln Asn Cys 420 425 430 Glu His Ala Asp Asn Tyr Thr Ala Tyr Cys Leu Gly Ile Ser His Met 435 440 445 Glu Pro Ser Phe Gly Leu Ile Leu His Asp Gly Gly Thr Thr Leu Lys 450 455 460 Phe Val Asp Thr Pro Glu Ser Leu Ser Gly Leu Tyr Val Phe Val Val 465 470 475 480 Tyr Phe Asn Gly His Val Glu Ala Val Ala Tyr Thr Val Val Ser Thr 485 490 495 Val Asp His Phe Val Asn Ala Ile Glu Glu Arg Gly Phe Pro Pro Thr 500 505 510 Ala Gly Gln Pro Pro Ala Thr Thr Lys Pro Lys Glu Ile Thr Pro Val 515 520 525 Asn Pro Gly Thr Ser Pro Leu 530 535 <210> 4 <211> 536 <212> PRT <213> Artificial Sequence <220> <223> Artificial Sequence <400> 4 Met Gly Thr Val Asn Lys Pro Val Val Gly Val Leu Met Gly Phe Gly 1 5 10 15 Ile Ile Thr Gly Thr Leu Arg Ile Thr Asn Pro Val Arg Ala Ser Val 20 25 30 Leu Arg Tyr Asp Asp Phe His Thr Asp Glu Asp Lys Leu Asp Thr Asn 35 40 45 Ser Val Tyr Glu Pro Tyr Tyr His Ser Asp His Ala Glu Ser Ser Trp 50 55 60 Val Asn Arg Gly Glu Ser Ser Arg Lys Ala Tyr Asp His Asn Ser Pro 65 70 75 80 Tyr Ile Trp Pro Arg Asn Asp Tyr Asp Gly Phe Leu Glu Asn Ala His 85 90 95 Glu His His Gly Val Tyr Asn Gln Gly Arg Gly Ile Asp Ser Gly Glu 100 105 110 Arg Leu Met Gln Pro Thr Gln Met Ser Ala Gln Glu Asp Leu Gly Asp 115 120 125 Asp Thr Gly Ile His Val Ile Pro Thr Leu Asn Gly Asp Asp Arg His 130 135 140 Lys Ile Val Asn Val Asp Gln Arg Gln Tyr Gly Asp Val Phe Lys Gly 145 150 155 160 Asp Leu Asn Pro Lys Pro Gln Gly Gln Arg Leu Ile Glu Val Ser Val 165 170 175 Glu Glu Asn His Pro Phe Thr Leu Arg Ala Pro Ile Gln Arg Ile Tyr 180 185 190 Gly Val Arg Tyr Thr Glu Thr Trp Ser Phe Leu Pro Ser Leu Thr Cys 195 200 205 Thr Gly Asp Ala Ala Pro Ala Ile Gln His Ile Cys Leu Lys His Thr 210 215 220 Thr Cys Phe Gln Asp Val Val Val Asp Val Asp Cys Ala Glu Asn Thr 225 230 235 240 Lys Glu Asp Gln Leu Ala Glu Ile Ser Tyr Arg Phe Gln Gly Lys Lys 245 250 255 Glu Ala Asp Gln Pro Trp Ile Val Val Asn Thr Ser Thr Leu Phe Asp 260 265 270 Glu Leu Glu Leu Asp Pro Pro Glu Ile Glu Pro Gly Val Leu Lys Val 275 280 285 Leu Arg Thr Glu Lys Gln Tyr Leu Gly Val Tyr Ile Trp Asn Met Arg 290 295 300 Gly Ser Asp Gly Thr Ser Thr Tyr Ala Thr Phe Leu Val Thr Trp Lys 305 310 315 320 Gly Asp Glu Lys Thr Arg Asn Pro Thr Pro Ala Val Thr Pro Gln Pro 325 330 335 Arg Gly Ala Glu Phe His Met Trp Asn Tyr His Ser His Val Phe Ser 340 345 350 Val Gly Asp Thr Phe Ser Leu Ala Met His Leu Gln Tyr Lys Ile His 355 360 365 Glu Ala Pro Phe Asp Leu Leu Leu Glu Trp Leu Tyr Val Pro Ile Asp 370 375 380 Pro Thr Cys Gln Pro Met Arg Leu Tyr Ser Thr Cys Leu Tyr His Pro 385 390 395 400 Asn Ala Pro Gln Cys Leu Ser His Met Asn Ser Gly Cys Thr Phe Thr 405 410 415 Ser Pro His Leu Ala Gln Arg Val Ala Ser Thr Val Tyr Gln Asn Cys 420 425 430 Glu His Ala Asp Asn Tyr Thr Ala Tyr Cys Leu Gly Ile Ser His Met 435 440 445 Glu Pro Ser Phe Gly Leu Ile Leu His Asp Gly Gly Thr Thr Leu Lys 450 455 460 Phe Val Asp Thr Pro Glu Ser Leu Ser Gly Leu Tyr Val Phe Val Val 465 470 475 480 Tyr Phe Asn Gly His Val Glu Ala Val Ala Tyr Thr Val Val Ser Thr 485 490 495 Val Asp His Phe Val Asn Ala Ile Glu Glu Arg Gly Phe Pro Pro Thr 500 505 510 Ala Gly Gln Pro Pro Ala Thr Thr Lys Pro Lys Glu Ile Thr Pro Val 515 520 525 Asn Pro Gly Thr Ser Pro Leu Leu 530 535 <210> 5 <211> 537 <212> PRT <213> Artificial Sequence <220> <223> Artificial Sequence <400> 5 Met Gly Thr Val Asn Lys Pro Val Val Gly Val Leu Met Gly Phe Gly 1 5 10 15 Ile Ile Thr Gly Thr Leu Arg Ile Thr Asn Pro Val Arg Ala Ser Val 20 25 30 Leu Arg Tyr Asp Asp Phe His Thr Asp Glu Asp Lys Leu Asp Thr Asn 35 40 45 Ser Val Tyr Glu Pro Tyr Tyr His Ser Asp His Ala Glu Ser Ser Trp 50 55 60 Val Asn Arg Gly Glu Ser Ser Arg Lys Ala Tyr Asp His Asn Ser Pro 65 70 75 80 Tyr Ile Trp Pro Arg Asn Asp Tyr Asp Gly Phe Leu Glu Asn Ala His 85 90 95 Glu His His Gly Val Tyr Asn Gln Gly Arg Gly Ile Asp Ser Gly Glu 100 105 110 Arg Leu Met Gln Pro Thr Gln Met Ser Ala Gln Glu Asp Leu Gly Asp 115 120 125 Asp Thr Gly Ile His Val Ile Pro Thr Leu Asn Gly Asp Asp Arg His 130 135 140 Lys Ile Val Asn Val Asp Gln Arg Gln Tyr Gly Asp Val Phe Lys Gly 145 150 155 160 Asp Leu Asn Pro Lys Pro Gln Gly Gln Arg Leu Ile Glu Val Ser Val 165 170 175 Glu Glu Asn His Pro Phe Thr Leu Arg Ala Pro Ile Gln Arg Ile Tyr 180 185 190 Gly Val Arg Tyr Thr Glu Thr Trp Ser Phe Leu Pro Ser Leu Thr Cys 195 200 205 Thr Gly Asp Ala Ala Pro Ala Ile Gln His Ile Cys Leu Lys His Thr 210 215 220 Thr Cys Phe Gln Asp Val Val Val Asp Val Asp Cys Ala Glu Asn Thr 225 230 235 240 Lys Glu Asp Gln Leu Ala Glu Ile Ser Tyr Arg Phe Gln Gly Lys Lys 245 250 255 Glu Ala Asp Gln Pro Trp Ile Val Val Asn Thr Ser Thr Leu Phe Asp 260 265 270 Glu Leu Glu Leu Asp Pro Pro Glu Ile Glu Pro Gly Val Leu Lys Val 275 280 285 Leu Arg Thr Glu Lys Gln Tyr Leu Gly Val Tyr Ile Trp Asn Met Arg 290 295 300 Gly Ser Asp Gly Thr Ser Thr Tyr Ala Thr Phe Leu Val Thr Trp Lys 305 310 315 320 Gly Asp Glu Lys Thr Arg Asn Pro Thr Pro Ala Val Thr Pro Gln Pro 325 330 335 Arg Gly Ala Glu Phe His Met Trp Asn Tyr His Ser His Val Phe Ser 340 345 350 Val Gly Asp Thr Phe Ser Leu Ala Met His Leu Gln Tyr Lys Ile His 355 360 365 Glu Ala Pro Phe Asp Leu Leu Leu Glu Trp Leu Tyr Val Pro Ile Asp 370 375 380 Pro Thr Cys Gln Pro Met Arg Leu Tyr Ser Thr Cys Leu Tyr His Pro 385 390 395 400 Asn Ala Pro Gln Cys Leu Ser His Met Asn Ser Gly Cys Thr Phe Thr 405 410 415 Ser Pro His Leu Ala Gln Arg Val Ala Ser Thr Val Tyr Gln Asn Cys 420 425 430 Glu His Ala Asp Asn Tyr Thr Ala Tyr Cys Leu Gly Ile Ser His Met 435 440 445 Glu Pro Ser Phe Gly Leu Ile Leu His Asp Gly Gly Thr Thr Leu Lys 450 455 460 Phe Val Asp Thr Pro Glu Ser Leu Ser Gly Leu Tyr Val Phe Val Val 465 470 475 480 Tyr Phe Asn Gly His Val Glu Ala Val Ala Tyr Thr Val Val Ser Thr 485 490 495 Val Asp His Phe Val Asn Ala Ile Glu Glu Arg Gly Phe Pro Pro Thr 500 505 510 Ala Gly Gln Pro Pro Ala Thr Thr Lys Pro Lys Glu Ile Thr Pro Val 515 520 525 Asn Pro Gly Thr Ser Pro Leu Leu Arg 530 535 <210> 6 <211> 538 <212> PRT <213> Artificial Sequence <220> <223> Artificial Sequence <400> 6 Met Gly Thr Val Asn Lys Pro Val Val Gly Val Leu Met Gly Phe Gly 1 5 10 15 Ile Ile Thr Gly Thr Leu Arg Ile Thr Asn Pro Val Arg Ala Ser Val 20 25 30 Leu Arg Tyr Asp Asp Phe His Thr Asp Glu Asp Lys Leu Asp Thr Asn 35 40 45 Ser Val Tyr Glu Pro Tyr Tyr His Ser Asp His Ala Glu Ser Ser Trp 50 55 60 Val Asn Arg Gly Glu Ser Ser Arg Lys Ala Tyr Asp His Asn Ser Pro 65 70 75 80 Tyr Ile Trp Pro Arg Asn Asp Tyr Asp Gly Phe Leu Glu Asn Ala His 85 90 95 Glu His His Gly Val Tyr Asn Gln Gly Arg Gly Ile Asp Ser Gly Glu 100 105 110 Arg Leu Met Gln Pro Thr Gln Met Ser Ala Gln Glu Asp Leu Gly Asp 115 120 125 Asp Thr Gly Ile His Val Ile Pro Thr Leu Asn Gly Asp Asp Arg His 130 135 140 Lys Ile Val Asn Val Asp Gln Arg Gln Tyr Gly Asp Val Phe Lys Gly 145 150 155 160 Asp Leu Asn Pro Lys Pro Gln Gly Gln Arg Leu Ile Glu Val Ser Val 165 170 175 Glu Glu Asn His Pro Phe Thr Leu Arg Ala Pro Ile Gln Arg Ile Tyr 180 185 190 Gly Val Arg Tyr Thr Glu Thr Trp Ser Phe Leu Pro Ser Leu Thr Cys 195 200 205 Thr Gly Asp Ala Ala Pro Ala Ile Gln His Ile Cys Leu Lys His Thr 210 215 220 Thr Cys Phe Gln Asp Val Val Val Asp Val Asp Cys Ala Glu Asn Thr 225 230 235 240 Lys Glu Asp Gln Leu Ala Glu Ile Ser Tyr Arg Phe Gln Gly Lys Lys 245 250 255 Glu Ala Asp Gln Pro Trp Ile Val Val Asn Thr Ser Thr Leu Phe Asp 260 265 270 Glu Leu Glu Leu Asp Pro Pro Glu Ile Glu Pro Gly Val Leu Lys Val 275 280 285 Leu Arg Thr Glu Lys Gln Tyr Leu Gly Val Tyr Ile Trp Asn Met Arg 290 295 300 Gly Ser Asp Gly Thr Ser Thr Tyr Ala Thr Phe Leu Val Thr Trp Lys 305 310 315 320 Gly Asp Glu Lys Thr Arg Asn Pro Thr Pro Ala Val Thr Pro Gln Pro 325 330 335 Arg Gly Ala Glu Phe His Met Trp Asn Tyr His Ser His Val Phe Ser 340 345 350 Val Gly Asp Thr Phe Ser Leu Ala Met His Leu Gln Tyr Lys Ile His 355 360 365 Glu Ala Pro Phe Asp Leu Leu Leu Glu Trp Leu Tyr Val Pro Ile Asp 370 375 380 Pro Thr Cys Gln Pro Met Arg Leu Tyr Ser Thr Cys Leu Tyr His Pro 385 390 395 400 Asn Ala Pro Gln Cys Leu Ser His Met Asn Ser Gly Cys Thr Phe Thr 405 410 415 Ser Pro His Leu Ala Gln Arg Val Ala Ser Thr Val Tyr Gln Asn Cys 420 425 430 Glu His Ala Asp Asn Tyr Thr Ala Tyr Cys Leu Gly Ile Ser His Met 435 440 445 Glu Pro Ser Phe Gly Leu Ile Leu His Asp Gly Gly Thr Thr Leu Lys 450 455 460 Phe Val Asp Thr Pro Glu Ser Leu Ser Gly Leu Tyr Val Phe Val Val 465 470 475 480 Tyr Phe Asn Gly His Val Glu Ala Val Ala Tyr Thr Val Val Ser Thr 485 490 495 Val Asp His Phe Val Asn Ala Ile Glu Glu Arg Gly Phe Pro Pro Thr 500 505 510 Ala Gly Gln Pro Pro Ala Thr Thr Lys Pro Lys Glu Ile Thr Pro Val 515 520 525 Asn Pro Gly Thr Ser Pro Leu Leu Arg Tyr 530 535 <210> 7 <211> 539 <212> PRT <213> Artificial Sequence <220> <223> Artificial Sequence <400> 7 Met Gly Thr Val Asn Lys Pro Val Val Gly Val Leu Met Gly Phe Gly 1 5 10 15 Ile Ile Thr Gly Thr Leu Arg Ile Thr Asn Pro Val Arg Ala Ser Val 20 25 30 Leu Arg Tyr Asp Asp Phe His Thr Asp Glu Asp Lys Leu Asp Thr Asn 35 40 45 Ser Val Tyr Glu Pro Tyr Tyr His Ser Asp His Ala Glu Ser Ser Trp 50 55 60 Val Asn Arg Gly Glu Ser Ser Arg Lys Ala Tyr Asp His Asn Ser Pro 65 70 75 80 Tyr Ile Trp Pro Arg Asn Asp Tyr Asp Gly Phe Leu Glu Asn Ala His 85 90 95 Glu His His Gly Val Tyr Asn Gln Gly Arg Gly Ile Asp Ser Gly Glu 100 105 110 Arg Leu Met Gln Pro Thr Gln Met Ser Ala Gln Glu Asp Leu Gly Asp 115 120 125 Asp Thr Gly Ile His Val Ile Pro Thr Leu Asn Gly Asp Asp Arg His 130 135 140 Lys Ile Val Asn Val Asp Gln Arg Gln Tyr Gly Asp Val Phe Lys Gly 145 150 155 160 Asp Leu Asn Pro Lys Pro Gln Gly Gln Arg Leu Ile Glu Val Ser Val 165 170 175 Glu Glu Asn His Pro Phe Thr Leu Arg Ala Pro Ile Gln Arg Ile Tyr 180 185 190 Gly Val Arg Tyr Thr Glu Thr Trp Ser Phe Leu Pro Ser Leu Thr Cys 195 200 205 Thr Gly Asp Ala Ala Pro Ala Ile Gln His Ile Cys Leu Lys His Thr 210 215 220 Thr Cys Phe Gln Asp Val Val Val Asp Val Asp Cys Ala Glu Asn Thr 225 230 235 240 Lys Glu Asp Gln Leu Ala Glu Ile Ser Tyr Arg Phe Gln Gly Lys Lys 245 250 255 Glu Ala Asp Gln Pro Trp Ile Val Val Asn Thr Ser Thr Leu Phe Asp 260 265 270 Glu Leu Glu Leu Asp Pro Pro Glu Ile Glu Pro Gly Val Leu Lys Val 275 280 285 Leu Arg Thr Glu Lys Gln Tyr Leu Gly Val Tyr Ile Trp Asn Met Arg 290 295 300 Gly Ser Asp Gly Thr Ser Thr Tyr Ala Thr Phe Leu Val Thr Trp Lys 305 310 315 320 Gly Asp Glu Lys Thr Arg Asn Pro Thr Pro Ala Val Thr Pro Gln Pro 325 330 335 Arg Gly Ala Glu Phe His Met Trp Asn Tyr His Ser His Val Phe Ser 340 345 350 Val Gly Asp Thr Phe Ser Leu Ala Met His Leu Gln Tyr Lys Ile His 355 360 365 Glu Ala Pro Phe Asp Leu Leu Leu Glu Trp Leu Tyr Val Pro Ile Asp 370 375 380 Pro Thr Cys Gln Pro Met Arg Leu Tyr Ser Thr Cys Leu Tyr His Pro 385 390 395 400 Asn Ala Pro Gln Cys Leu Ser His Met Asn Ser Gly Cys Thr Phe Thr 405 410 415 Ser Pro His Leu Ala Gln Arg Val Ala Ser Thr Val Tyr Gln Asn Cys 420 425 430 Glu His Ala Asp Asn Tyr Thr Ala Tyr Cys Leu Gly Ile Ser His Met 435 440 445 Glu Pro Ser Phe Gly Leu Ile Leu His Asp Gly Gly Thr Thr Leu Lys 450 455 460 Phe Val Asp Thr Pro Glu Ser Leu Ser Gly Leu Tyr Val Phe Val Val 465 470 475 480 Tyr Phe Asn Gly His Val Glu Ala Val Ala Tyr Thr Val Val Ser Thr 485 490 495 Val Asp His Phe Val Asn Ala Ile Glu Glu Arg Gly Phe Pro Pro Thr 500 505 510 Ala Gly Gln Pro Pro Ala Thr Thr Lys Pro Lys Glu Ile Thr Pro Val 515 520 525 Asn Pro Gly Thr Ser Pro Leu Leu Arg Tyr Ala 530 535 <210> 8 <211> 540 <212> PRT <213> Artificial Sequence <220> <223> Artificial Sequence <400> 8 Met Gly Thr Val Asn Lys Pro Val Val Gly Val Leu Met Gly Phe Gly 1 5 10 15 Ile Ile Thr Gly Thr Leu Arg Ile Thr Asn Pro Val Arg Ala Ser Val 20 25 30 Leu Arg Tyr Asp Asp Phe His Thr Asp Glu Asp Lys Leu Asp Thr Asn 35 40 45 Ser Val Tyr Glu Pro Tyr Tyr His Ser Asp His Ala Glu Ser Ser Trp 50 55 60 Val Asn Arg Gly Glu Ser Ser Arg Lys Ala Tyr Asp His Asn Ser Pro 65 70 75 80 Tyr Ile Trp Pro Arg Asn Asp Tyr Asp Gly Phe Leu Glu Asn Ala His 85 90 95 Glu His His Gly Val Tyr Asn Gln Gly Arg Gly Ile Asp Ser Gly Glu 100 105 110 Arg Leu Met Gln Pro Thr Gln Met Ser Ala Gln Glu Asp Leu Gly Asp 115 120 125 Asp Thr Gly Ile His Val Ile Pro Thr Leu Asn Gly Asp Asp Arg His 130 135 140 Lys Ile Val Asn Val Asp Gln Arg Gln Tyr Gly Asp Val Phe Lys Gly 145 150 155 160 Asp Leu Asn Pro Lys Pro Gln Gly Gln Arg Leu Ile Glu Val Ser Val 165 170 175 Glu Glu Asn His Pro Phe Thr Leu Arg Ala Pro Ile Gln Arg Ile Tyr 180 185 190 Gly Val Arg Tyr Thr Glu Thr Trp Ser Phe Leu Pro Ser Leu Thr Cys 195 200 205 Thr Gly Asp Ala Ala Pro Ala Ile Gln His Ile Cys Leu Lys His Thr 210 215 220 Thr Cys Phe Gln Asp Val Val Val Asp Val Asp Cys Ala Glu Asn Thr 225 230 235 240 Lys Glu Asp Gln Leu Ala Glu Ile Ser Tyr Arg Phe Gln Gly Lys Lys 245 250 255 Glu Ala Asp Gln Pro Trp Ile Val Val Asn Thr Ser Thr Leu Phe Asp 260 265 270 Glu Leu Glu Leu Asp Pro Pro Glu Ile Glu Pro Gly Val Leu Lys Val 275 280 285 Leu Arg Thr Glu Lys Gln Tyr Leu Gly Val Tyr Ile Trp Asn Met Arg 290 295 300 Gly Ser Asp Gly Thr Ser Thr Tyr Ala Thr Phe Leu Val Thr Trp Lys 305 310 315 320 Gly Asp Glu Lys Thr Arg Asn Pro Thr Pro Ala Val Thr Pro Gln Pro 325 330 335 Arg Gly Ala Glu Phe His Met Trp Asn Tyr His Ser His Val Phe Ser 340 345 350 Val Gly Asp Thr Phe Ser Leu Ala Met His Leu Gln Tyr Lys Ile His 355 360 365 Glu Ala Pro Phe Asp Leu Leu Leu Glu Trp Leu Tyr Val Pro Ile Asp 370 375 380 Pro Thr Cys Gln Pro Met Arg Leu Tyr Ser Thr Cys Leu Tyr His Pro 385 390 395 400 Asn Ala Pro Gln Cys Leu Ser His Met Asn Ser Gly Cys Thr Phe Thr 405 410 415 Ser Pro His Leu Ala Gln Arg Val Ala Ser Thr Val Tyr Gln Asn Cys 420 425 430 Glu His Ala Asp Asn Tyr Thr Ala Tyr Cys Leu Gly Ile Ser His Met 435 440 445 Glu Pro Ser Phe Gly Leu Ile Leu His Asp Gly Gly Thr Thr Leu Lys 450 455 460 Phe Val Asp Thr Pro Glu Ser Leu Ser Gly Leu Tyr Val Phe Val Val 465 470 475 480 Tyr Phe Asn Gly His Val Glu Ala Val Ala Tyr Thr Val Val Ser Thr 485 490 495 Val Asp His Phe Val Asn Ala Ile Glu Glu Arg Gly Phe Pro Pro Thr 500 505 510 Ala Gly Gln Pro Pro Ala Thr Thr Lys Pro Lys Glu Ile Thr Pro Val 515 520 525 Asn Pro Gly Thr Ser Pro Leu Leu Arg Tyr Ala Ala 530 535 540 <210> 9 <211> 1602 <212> DNA <213> Artificial Sequence <220> <223> Artificial Sequence <400> 9 atgggaacag tcaacaaacc agtcgtcggc gtgctgatgg gcttcggtat tattacagga 60 actctgagga ttactaaccc cgtgcgcgcc tctgtgctgc ggtacgacga tttccacaca 120 gacgaggata agctggacac caattccgtg tatgagccct actatcactc tgatcacgcc 180 gagagctcct gggtgaaccg gggcgagtct agcaggaagg cttacgacca caacagccct 240 tatatctggc cacggaatga ctacgatggc tttctggaga acgcccacga gcaccacggc 300 gtgtataatc agggcagagg catcgactct ggcgagcggc tgatgcagcc cacccagatg 360 agcgcccagg aggatctggg cgacgataca ggcatccacg tgatccctac cctgaatggc 420 gacgataggc acaagatcgt gaacgtggat cagagacagt acggcgacgt gttcaagggc 480 gatctgaatc ccaagcctca gggccagagg ctgatcgagg tgtccgtgga ggagaaccac 540 cccttcaccc tgagagcccc tatccagcgg atctacggcg tgaggtatac cgagacatgg 600 agctttctgc catccctgac atgcaccggc gacgctgctc ctgctatcca gcacatctgc 660 ctgaagcaca ccacatgttt tcaggacgtg gtggtggacg tggattgtgc cgagaataca 720 aaggaggatc agctggctga gatctcctac cggttccagg ccaagaagga ggccgatcag 780 ccttggatcg tggtgaacac ctctacactg tttgacgagc tggagctgga tccccctgag 840 atcgagccag gcgtgctgaa ggtgctgaga accgagaagc agtacctggg cgtgtatatc 900 tggaacatgc ggggctctga cggcaccagc acatacgcta ccttcctggt cacatggaag 960 ggcgatgaga agaccccggaa tccaacacct gctgtgaccc ctcagccaag gggagctgag 1020 tttcacatgt ggaactatca ctcccacgtg ttctctgtgg gcgacacctt tagcctggcc 1080 atgcacctgc aatataagat ccacgaggct cctttcgacc tgctgctgga gtggctgtat 1140 gtgcccatcg atcctacatg ccagccaatg aggctgtact ccacctgtct gtatcaccca 1200 aatgcccccc aatgcctgag ccacatgaac tccggctgta cctttacaag cccccacctg 1260 gcccagagag tggcttccac agtgtaccag aactgcgagc acgccgacaa ttacaccgct 1320 tattgtctgg gcatctctca catggagccc agcttcggcc tgatcctgca cgacggcggc 1380 accacactga agtttgtgga tacacccgag tccctgtctg gcctctacgt gttcgtggtg 1440 tacttcaacg gccacgtgga ggccgtggct tatacagtgg tgtctaccgt ggatcacttc 1500 gtgaacgcca tcgaggagag aggatttcca cctaccgctg gacagcctcc agctaccaca 1560 aagcctaagg aaatcacccc tgtcaatcct ggaacttcac ct 1602 <210> 10 <211> 1605 <212> DNA <213> Artificial Sequence <220> <223> Artificial Sequence <400> 10 atgggaacag tcaacaaacc agtcgtcggc gtgctgatgg gcttcggtat tattacagga 60 actctgagga ttactaaccc cgtgcgcgcc tctgtgctgc ggtacgacga tttccacaca 120 gacgaggata agctggacac caattccgtg tatgagccct actatcactc tgatcacgcc 180 gagagctcct gggtgaaccg gggcgagtct agcaggaagg cttacgacca caacagccct 240 tatatctggc cacggaatga ctacgatggc tttctggaga acgcccacga gcaccacggc 300 gtgtataatc agggcagagg catcgactct ggcgagcggc tgatgcagcc cacccagatg 360 agcgcccagg aggatctggg cgacgataca ggcatccacg tgatccctac cctgaatggc 420 gacgataggc acaagatcgt gaacgtggat cagagacagt acggcgacgt gttcaagggc 480 gatctgaatc ccaagcctca gggccagagg ctgatcgagg tgtccgtgga ggagaaccac 540 cccttcaccc tgagagcccc tatccagcgg atctacggcg tgaggtatac cgagacatgg 600 agctttctgc catccctgac atgcaccggc gacgctgctc ctgctatcca gcacatctgc 660 ctgaagcaca ccacatgttt tcaggacgtg gtggtggacg tggattgtgc cgagaataca 720 aaggaggatc agctggctga gatctcctac cggttccagg ccaagaagga ggccgatcag 780 ccttggatcg tggtgaacac ctctacactg tttgacgagc tggagctgga tccccctgag 840 atcgagccag gcgtgctgaa ggtgctgaga accgagaagc agtacctggg cgtgtatatc 900 tggaacatgc ggggctctga cggcaccagc acatacgcta ccttcctggt cacatggaag 960 ggcgatgaga agaccccggaa tccaacacct gctgtgaccc ctcagccaag gggagctgag 1020 tttcacatgt ggaactatca ctcccacgtg ttctctgtgg gcgacacctt tagcctggcc 1080 atgcacctgc aataataagat ccacgaggct cctttcgacc tgctgctgga gtggctgtat 1140 gtgcccatcg atcctacatg ccagccaatg aggctgtact ccacctgtct gtatcaccca 1200 aatgcccccc aatgcctgag ccacatgaac tccggctga cctttacaag ccccacctg 1260 gcccagagag tggcttccac agtgtaccag aactgcgagc acgccgacaa ttacaccgct 1320 tattgtctgg gcatctctca catggagccc agcttcggcc tgatcctgca cgacggcggc 1380 accacactga agtttgtgga tacaccccgag tccctgtctg gcctctacgt gttcgtggtg 1440 tacttcaacg gccacgtgga ggccgtggct tatacagtgg tgtctaccgt ggatcacttc 1500 gtgaacgcca tcgaggagag aggatttcca cctaccgctg gacagcctcc agctaccaca 1560 aagcctaagg aaatcacccc tgtcaatcct ggaacttcac ctctg 1605 <210> 11 <211> 1608 <212> DNA <213> Artificial Sequence <220> <223> Artificial Sequence <400> 11 atgggaacag tcaacaaacc agtcgtcggc gtgctgatgg gcttcggtat tattacagga 60 actctgagga ttactaaccc cgtgcgcgcc tctgtgctgc ggtacgacga tttccacaca 120 gacgaggata agctggacac caattccgtg tatgagccct actatcactc tgatcacgcc 180 gagagctcct gggtgaaccg gggcgagtct agcaggaagg cttacgacca caacagccct 240 tatatctggc cacggaatga ctacgatggc tttctggaga acgcccacga gcaccacggc 300 gtgtataatc agggcagagg catcgactct ggcgagcggc tgatgcagcc cacccagatg 360 agcgcccagg aggatctggg cgacgataca ggcatccacg tgatccctac cctgaatggc 420 gacgataggc acaagatcgt gaacgtggat cagagacagt acggcgacgt gttcaagggc 480 gatctgaatc ccaagcctca gggccagagg ctgatcgagg tgtccgtgga ggagaaccac 540 cccttcaccc tgagagcccc tatccagcgg atctacggcg tgaggtatac cgagacatgg 600 agctttctgc catccctgac atgcaccggc gacgctgctc ctgctatcca gcacatctgc 660 ctgaagcaca ccacatgttt tcaggacgtg gtggtggacg tggattgtgc cgagaataca 720 aaggaggatc agctggctga gatctcctac cggttccagg ccaagaagga ggccgatcag 780 ccttggatcg tggtgaacac ctctacactg tttgacgagc tggagctgga tccccctgag 840 atcgagccag gcgtgctgaa ggtgctgaga accgagaagc agtacctggg cgtgtatatc 900 tggaacatgc ggggctctga cggcaccagc acatacgcta ccttcctggt cacatggaag 960 ggcgatgaga agaccccggaa tccaacacct gctgtgaccc ctcagccaag gggagctgag 1020 tttcacatgt ggaactatca ctcccacgtg ttctctgtgg gcgacacctt tagcctggcc 1080 atgcacctgc aataataagat ccacgaggct cctttcgacc tgctgctgga gtggctgtat 1140 gtgcccatcg atcctacatg ccagccaatg aggctgtact ccacctgtct gtatcaccca 1200 aatgcccccc aatgcctgag ccacatgaac tccggctgta cctttacaag cccccacctg 1260 gcccagagag tggcttccac agtgtaccag aactgcgagc acgccgacaa ttacaccgct 1320 tattgtctgg gcatctctca catggagccc agcttcggcc tgatcctgca cgacggcggc 1380 accacactga agtttgtgga tacacccgag tccctgtctg gcctctacgt gttcgtggtg 1440 tacttcaacg gccacgtgga ggccgtggct tatacagtgg tgtctaccgt ggatcacttc 1500 gtgaacgcca tcgaggagag aggatttcca cctaccgctg gacagcctcc agctaccaca 1560 aagcctaagg aaatcacccc tgtcaatcct ggaacttcac ctctgctg 1608 <210> 12 <211> 1611 <212> DNA <213> Artificial Sequence <220> <223> Artificial Sequence <400> 12 atgggaacag tcaacaaacc agtcgtcggc gtgctgatgg gcttcggtat tattacagga 60 actctgagga ttactaaccc cgtgcgcgcc tctgtgctgc ggtacgacga tttccacaca 120 gacgaggata agctggacac caattccgtg tatgagccct actatcactc tgatcacgcc 180 gagagctcct gggtgaaccg gggcgagtct agcaggaagg cttacgacca caacagccct 240 300 gtgtataatc agggcagagg catcgactct ggcgagcggc tgatgcagcc cacccagatg 360 agcgcccagg aggatctggg cgacgataca ggcatccacg tgatccctac cctgaatggc 420 gacgataggc acaagatcgt gaacgtggat cagagacagt acggcgacgt gttcaagggc 480 gatctgaatc ccaagcctca gggccagagg ctgatcgagg tgtccgtgga ggagaaccac 540 cccttcaccc tgagagcccc tatccagcgg atctacggcg tgaggtatac cgagacatgg 600 agctttctgc catccctgac atgcaccggc gacgctgctc ctgctatcca gcacatctgc 660 ctgaagcaca ccacatgttt tcaggacgtg gtggtggacg tggattgtgc cgagaataca 720 aaggaggatc agctggctga gatctcctac cggttccagg ccaagaagga ggccgatcag 780 ccttggatcg tggtgaacac ctctacactg tttgacgagc tggagctgga tccccctgag 840 atcgagccag gcgtgctgaa ggtgctgaga accgagaagc agtacctggg cgtgtatatc 900 tggaacatgc ggggctctga cggcaccagc acatacgcta ccttcctggt cacatggaag 960 ggcgatgaga agacccggaa tccaacacct gctgtgaccc ctcagccaag gggagctgag 1020 tttcacatgt ggaactatca ctcccacgtg ttctctgtgg gcgacacctt tagcctggcc 1080 atgcacctgc aatataagat ccacgaggct cctttcgacc tgctgctgga gtggctgtat 1140 gtgcccatcg atcctacatg ccagccaatg aggctgtact ccacctgtct gtatcaccca 1200 aatgcccccc aatgcctgag ccacatgaac tccggctgta cctttacaag cccccacctg 1260 gcccagagag tggcttccac agtgtaccag aactgcgagc acgccgacaa ttacaccgct 1320 tattgtctgg gcatctctca catggagccc agcttcggcc tgatcctgca cgacggcggc 1380 accacactga agtttgtgga tacacccgag tccctgtctg gcctctacgt gttcgtggtg 1440 tacttcaacg gccacgtgga ggccgtggct tatacagtgg tgtctaccgt ggatcacttc 1500 gtgaacgcca tcgaggagag aggatttcca cctaccgctg gacagcctcc agctaccaca 1560 aagcctaagg aaatcacccc tgtcaatcct ggaacttcac ctctgctgcg c 1611 <210> 13 <211> 1614 <212> DNA <213> Artificial Sequence <220> <223> Artificial Sequence <400> 13 atgggaacag tcaacaaacc agtcgtcggc gtgctgatgg gcttcggtat tattacagga 60 actctgagga ttactaaccc cgtgcgcgcc tctgtgctgc ggtacgacga tttccacaca 120 gacgaggata agctggacac caattccgtg tatgagccct actatcactc tgatcacgcc 180 gagagctcct gggtgaaccg gggcgagtct agcaggaagg cttacgacca caacagccct 240 tatatctggc cacggaatga ctacgatggc tttctggaga acgcccacga gcaccacggc 300 gtgtataatc agggcagagg catcgactct ggcgagcggc tgatgcagcc cacccagatg 360 agcgcccagg aggatctggg cgacgataca ggcatccacg tgatccctac cctgaatggc 420 gacgataggc acaagatcgt gaacgtggat cagagacagt acggcgacgt gttcaagggc 480 gatctgaatc ccaagcctca gggccagagg ctgatcgagg tgtccgtgga ggagaaccac 540 cccttcaccc tgagagcccc tatccagcgg atctacggcg tgaggtatac cgagacatgg 600 agctttctgc catccctgac atgcaccggc gacgctgctc ctgctatcca gcacatctgc 660 ctgaagcaca ccacatgttt tcaggacgtg gtggtggacg tggattgtgc cgagaataca 720 aaggaggatc agctggctga gatctcctac cggttccagg ccaagaagga ggccgatcag 780 ccttggatcg tggtgaacac ctctacactg tttgacgagc tggagctgga tccccctgag 840 atcgagccag gcgtgctgaa ggtgctgaga accgagaagc agtacctggg cgtgtatatc 900 tggaacatgc ggggctctga cggcaccagc acatacgcta ccttcctggt cacatggaag 960 ggcgatgaga agaccccggaa tccaacacct gctgtgaccc ctcagccaag gggagctgag 1020 tttcacatgt ggaactatca ctcccacgtg ttctctgtgg gcgacacctt tagcctggcc 1080 atgcacctgc aataataagat ccacgaggct cctttcgacc tgctgctgga gtggctgtat 1140 gtgcccatcg atcctacatg ccagccaatg aggctgtact ccacctgtct gtatcaccca 1200 aatgcccccc aatgcctgag ccacatgaac tccggctga cctttacaag ccccacctg 1260 gcccagagag tggcttccac agtgtaccag aactgcgagc acgccgacaa ttacaccgct 1320 tattgtctgg gcatctctca catggagccc agcttcggcc tgatcctgca cgacggcggc 1380 accacactga agtttgtgga tacacccgag tccctgtctg gcctctacgt gttcgtggtg 1440 tacttcaacg gccacgtgga ggccgtggct tatacagtgg tgtctaccgt ggatcacttc 1500 gtgaacgcca tcgaggagag aggatttcca cctaccgctg gacagcctcc agctaccaca 1560 aagcctaagg aaatcacccc tgtcaatcct ggaacttcac ctctgctgcg ctac 1614 <210> 14 <211> 1617 <212> DNA <213> Artificial Sequence <220> <223> Artificial Sequence <400> 14 atgggaacag tcaacaaacc agtcgtcggc gtgctgatgg gcttcggtat tattacagga 60 actctgagga ttactaaccc cgtgcgcgcc tctgtgctgc ggtacgacga tttccacaca 120 gacgaggata agctggacac caattccgtg tatgagccct actatcactc tgatcacgcc 180 gagagctcct gggtgaaccg gggcgagtct agcaggaagg cttacgacca caacagccct 240 300 gtgtataatc agggcagagg catcgactct ggcgagcggc tgatgcagcc cacccagatg 360 agcgcccagg aggatctggg cgacgataca ggcatccacg tgatccctac cctgaatggc 420 gacgataggc acaagatcgt gaacgtggat cagagacagt acggcgacgt gttcaagggc 480 gatctgaatc ccaagcctca gggccagagg ctgatcgagg tgtccgtgga ggagaaccac 540 cccttcaccc tgagagcccc tatccagcgg atctacggcg tgaggtatac cgagacatgg 600 agctttctgc catccctgac atgcaccggc gacgctgctc ctgctatcca gcacatctgc 660 ctgaagcaca ccacatgttt tcaggacgtg gtggtggacg tggattgtgc cgagaataca 720 aaggaggatc agctggctga gatctcctac cggttccagg ccaagaagga ggccgatcag 780 ccttggatcg tggtgaacac ctctacactg tttgacgagc tggagctgga tccccctgag 840 atcgagccag gcgtgctgaa ggtgctgaga accgagaagc agtacctggg cgtgtatatc 900 tggaacatgc ggggctctga cggcaccagc acatacgcta ccttcctggt cacatggaag 960 ggcgatgaga agacccggaa tccaacacct gctgtgaccc ctcagccaag gggagctgag 1020 tttcacatgt ggaactatca ctcccacgtg ttctctgtgg gcgacacctt tagcctggcc 1080 atgcacctgc aatataagat ccacgaggct cctttcgacc tgctgctgga gtggctgtat 1140 gtgcccatcg atcctacatg ccagccaatg aggctgtact ccacctgtct gtatcaccca 1200 aatgcccccc aatgcctgag ccacatgaac tccggctgta cctttacaag cccccacctg 1260 gcccagagag tggcttccac agtgtaccag aactgcgagc acgccgacaa ttacaccgct 1320 tattgtctgg gcatctctca catggagccc agcttcggcc tgatcctgca cgacggcggc 1380 accacactga agtttgtgga tacacccgag tccctgtctg gcctctacgt gttcgtggtg 1440 tacttcaacg gccacgtgga ggccgtggct tatacagtgg tgtctaccgt ggatcacttc 1500 gtgaacgcca tcgaggagag aggatttcca cctaccgctg gacagcctcc agctaccaca 1560 aagcctaagg aaatcacccc tgtcaatcct ggaacttcac ctctgctgcg ctacgca 1617 <210> 15 <211> 1620 <212> DNA <213> Artificial Sequence <220> <223> Artificial Sequence <400> 15 atgggaacag tcaacaaacc agtcgtcggc gtgctgatgg gcttcggtat tattacagga 60 actctgagga ttactaaccc cgtgcgcgcc tctgtgctgc ggtacgacga tttccacaca 120 gacgaggata agctggacac caattccgtg tatgagccct actatcactc tgatcacgcc 180 gagagctcct gggtgaaccg gggcgagtct agcaggaagg cttacgacca caacagccct 240 tatatctggc cacggaatga ctacgatggc tttctggaga acgcccacga gcaccacggc 300 gtgtataatc agggcagagg catcgactct ggcgagcggc tgatgcagcc cacccagatg 360 agcgcccagg aggatctggg cgacgataca ggcatccacg tgatccctac cctgaatggc 420 gacgataggc acaagatcgt gaacgtggat cagagacagt acggcgacgt gttcaagggc 480 gatctgaatc ccaagcctca gggccagagg ctgatcgagg tgtccgtgga ggagaaccac 540 cccttcaccc tgagagcccc tatccagcgg atctacggcg tgaggtatac cgagacatgg 600 agctttctgc catccctgac atgcaccggc gacgctgctc ctgctatcca gcacatctgc 660 ctgaagcaca ccacatgttt tcaggacgtg gtggtggacg tggattgtgc cgagaataca 720 aaggaggatc agctggctga gatctcctac cggttccagg ccaagaagga ggccgatcag 780 ccttggatcg tggtgaacac ctctacactg tttgacgagc tggagctgga tccccctgag 840 atcgagccag gcgtgctgaa ggtgctgaga accgagaagc agtacctggg cgtgtatatc 900 tggaacatgc ggggctctga cggcaccagc acatacgcta ccttcctggt cacatggaag 960 ggcgatgaga agaccccggaa tccaacacct gctgtgaccc ctcagccaag gggagctgag 1020 tttcacatgt ggaactatca ctcccacgtg ttctctgtgg gcgacacctt tagcctggcc 1080 atgcacctgc aataataagat ccacgaggct cctttcgacc tgctgctgga gtggctgtat 1140 gtgcccatcg atcctacatg ccagccaatg aggctgtact ccacctgtct gtatcaccca 1200 aatgcccccc aatgcctgag ccacatgaac tccggctgta cctttacaag ccccacctg 1260 gcccagagag tggcttccac agtgtaccag aactgcgagc acgccgacaa ttacaccgct 1320 tattgtctgg gcatctctca catggagccc agcttcggcc tgatcctgca cgacggcggc 1380 accacactga agtttgtgga tacacccgag tccctgtctg gcctctacgt gttcgtggtg 1440 tacttcaacg gccacgtgga ggccgtggct tatacagtgg tgtctaccgt ggatcacttc 1500 gtgaacgcca tcgaggagag aggatttcca cctaccgctg gacagcctcc agctaccaca 1560 aagcctaagg aaatcacccc tgtcaatcct ggaacttcac ctctgctgcg ctacgcagcc 1620 1620 <210> 16 <211> 500 <212> PRT <213> Artificial Sequence <220> <223> Artificial Sequence <400> 16 Met Gly Thr Val Asn Lys Pro Val Val Gly Val Leu Met Gly Phe Gly 1 5 10 15 Ile Ile Thr Gly Thr Leu Arg Ile Thr Asn Pro Val Arg Ala Ser Val 20 25 30 Leu Arg Tyr Asp Asp Phe His Thr Asp Glu Asp Lys Leu Asp Thr Asn 35 40 45 Ser Val Tyr Glu Pro Tyr Tyr His Ser Asp His Ala Glu Ser Ser Trp 50 55 60 Val Asn Arg Gly Glu Ser Ser Arg Lys Ala Tyr Asp His Asn Ser Pro 65 70 75 80 Tyr Ile Trp Pro Arg Asn Asp Tyr Asp Gly Phe Leu Glu Asn Ala His 85 90 95 Glu His His Gly Val Tyr Asn Gln Gly Arg Gly Ile Asp Ser Gly Glu 100 105 110 Arg Leu Met Gln Pro Thr Gln Met Ser Ala Gln Glu Asp Leu Gly Asp 115 120 125 Asp Thr Gly Ile His Val Ile Pro Thr Leu Asn Gly Asp Asp Arg His 130 135 140 Lys Ile Val Asn Val Asp Gln Arg Gln Tyr Gly Asp Val Phe Lys Gly 145 150 155 160 Asp Leu Asn Pro Lys Pro Gln Gly Gln Arg Leu Ile Glu Val Ser Val 165 170 175 Glu Glu Asn His Pro Phe Thr Leu Arg Ala Pro Ile Gln Arg Ile Tyr 180 185 190 Gly Val Arg Tyr Thr Glu Thr Trp Ser Phe Leu Pro Ser Leu Thr Cys 195 200 205 Thr Gly Asp Ala Ala Pro Ala Ile Gln His Ile Cys Leu Lys His Thr 210 215 220 Thr Cys Phe Gln Asp Val Val Val Asp Val Asp Cys Ala Glu Asn Thr 225 230 235 240 Lys Glu Asp Gln Leu Ala Glu Ile Ser Tyr Arg Phe Gln Gly Lys Lys 245 250 255 Glu Ala Asp Gln Pro Trp Ile Val Val Asn Thr Ser Thr Leu Phe Asp 260 265 270 Glu Leu Glu Leu Asp Pro Pro Glu Ile Glu Pro Gly Val Leu Lys Val 275 280 285 Leu Arg Thr Glu Lys Gln Tyr Leu Gly Val Tyr Ile Trp Asn Met Arg 290 295 300 Gly Ser Asp Gly Thr Ser Thr Tyr Ala Thr Phe Leu Val Thr Trp Lys 305 310 315 320 Gly Asp Glu Lys Thr Arg Asn Pro Thr Pro Ala Val Thr Pro Gln Pro 325 330 335 Arg Gly Ala Glu Phe His Met Trp Asn Tyr His Ser His Val Phe Ser 340 345 350 Val Gly Asp Thr Phe Ser Leu Ala Met His Leu Gln Tyr Lys Ile His 355 360 365 Glu Ala Pro Phe Asp Leu Leu Leu Glu Trp Leu Tyr Val Pro Ile Asp 370 375 380 Pro Thr Cys Gln Pro Met Arg Leu Tyr Ser Thr Cys Leu Tyr His Pro 385 390 395 400 Asn Ala Pro Gln Cys Leu Ser His Met Asn Ser Gly Cys Thr Phe Thr 405 410 415 Ser Pro His Leu Ala Gln Arg Val Ala Ser Thr Val Tyr Gln Asn Cys 420 425 430 Glu His Ala Asp Asn Tyr Thr Ala Tyr Cys Leu Gly Ile Ser His Met 435 440 445 Glu Pro Ser Phe Gly Leu Ile Leu His Asp Gly Gly Thr Thr Leu Lys 450 455 460 Phe Val Asp Thr Pro Glu Ser Leu Ser Gly Leu Tyr Val Phe Val Val 465 470 475 480 Tyr Phe Asn Gly His Val Glu Ala Val Ala Tyr Thr Val Val Ser Thr 485 490 495 Val Asp His Phe 500 <210> 17 <211> 505 <212> PRT <213> Artificial Sequence <220> <223> Artificial Sequence <400> 17 Met Gly Thr Val Asn Lys Pro Val Val Gly Val Leu Met Gly Phe Gly 1 5 10 15 Ile Ile Thr Gly Thr Leu Arg Ile Thr Asn Pro Val Arg Ala Ser Val 20 25 30 Leu Arg Tyr Asp Asp Phe His Thr Asp Glu Asp Lys Leu Asp Thr Asn 35 40 45 Ser Val Tyr Glu Pro Tyr Tyr His Ser Asp His Ala Glu Ser Ser Trp 50 55 60 Val Asn Arg Gly Glu Ser Ser Arg Lys Ala Tyr Asp His Asn Ser Pro 65 70 75 80 Tyr Ile Trp Pro Arg Asn Asp Tyr Asp Gly Phe Leu Glu Asn Ala His 85 90 95 Glu His His Gly Val Tyr Asn Gln Gly Arg Gly Ile Asp Ser Gly Glu 100 105 110 Arg Leu Met Gln Pro Thr Gln Met Ser Ala Gln Glu Asp Leu Gly Asp 115 120 125 Asp Thr Gly Ile His Val Ile Pro Thr Leu Asn Gly Asp Asp Arg His 130 135 140 Lys Ile Val Asn Val Asp Gln Arg Gln Tyr Gly Asp Val Phe Lys Gly 145 150 155 160 Asp Leu Asn Pro Lys Pro Gln Gly Gln Arg Leu Ile Glu Val Ser Val 165 170 175 Glu Glu Asn His Pro Phe Thr Leu Arg Ala Pro Ile Gln Arg Ile Tyr 180 185 190 Gly Val Arg Tyr Thr Glu Thr Trp Ser Phe Leu Pro Ser Leu Thr Cys 195 200 205 Thr Gly Asp Ala Ala Pro Ala Ile Gln His Ile Cys Leu Lys His Thr 210 215 220 Thr Cys Phe Gln Asp Val Val Val Asp Val Asp Cys Ala Glu Asn Thr 225 230 235 240 Lys Glu Asp Gln Leu Ala Glu Ile Ser Tyr Arg Phe Gln Gly Lys Lys 245 250 255 Glu Ala Asp Gln Pro Trp Ile Val Val Asn Thr Ser Thr Leu Phe Asp 260 265 270 Glu Leu Glu Leu Asp Pro Pro Glu Ile Glu Pro Gly Val Leu Lys Val 275 280 285 Leu Arg Thr Glu Lys Gln Tyr Leu Gly Val Tyr Ile Trp Asn Met Arg 290 295 300 Gly Ser Asp Gly Thr Ser Thr Tyr Ala Thr Phe Leu Val Thr Trp Lys 305 310 315 320 Gly Asp Glu Lys Thr Arg Asn Pro Thr Pro Ala Val Thr Pro Gln Pro 325 330 335 Arg Gly Ala Glu Phe His Met Trp Asn Tyr His Ser His Val Phe Ser 340 345 350 Val Gly Asp Thr Phe Ser Leu Ala Met His Leu Gln Tyr Lys Ile His 355 360 365 Glu Ala Pro Phe Asp Leu Leu Leu Glu Trp Leu Tyr Val Pro Ile Asp 370 375 380 Pro Thr Cys Gln Pro Met Arg Leu Tyr Ser Thr Cys Leu Tyr His Pro 385 390 395 400 Asn Ala Pro Gln Cys Leu Ser His Met Asn Ser Gly Cys Thr Phe Thr 405 410 415 Ser Pro His Leu Ala Gln Arg Val Ala Ser Thr Val Tyr Gln Asn Cys 420 425 430 Glu His Ala Asp Asn Tyr Thr Ala Tyr Cys Leu Gly Ile Ser His Met 435 440 445 Glu Pro Ser Phe Gly Leu Ile Leu His Asp Gly Gly Thr Thr Leu Lys 450 455 460 Phe Val Asp Thr Pro Glu Ser Leu Ser Gly Leu Tyr Val Phe Val Val 465 470 475 480 Tyr Phe Asn Gly His Val Glu Ala Val Ala Tyr Thr Val Val Ser Thr 485 490 495 Val Asp His Phe Val Asn Ala Ile Glu 500 505 <210> 18 <211> 510 <212> PRT <213> Artificial Sequence <220> <223> Artificial Sequence <400> 18 Met Gly Thr Val Asn Lys Pro Val Val Gly Val Leu Met Gly Phe Gly 1 5 10 15 Ile Ile Thr Gly Thr Leu Arg Ile Thr Asn Pro Val Arg Ala Ser Val 20 25 30 Leu Arg Tyr Asp Asp Phe His Thr Asp Glu Asp Lys Leu Asp Thr Asn 35 40 45 Ser Val Tyr Glu Pro Tyr Tyr His Ser Asp His Ala Glu Ser Ser Trp 50 55 60 Val Asn Arg Gly Glu Ser Ser Arg Lys Ala Tyr Asp His Asn Ser Pro 65 70 75 80 Tyr Ile Trp Pro Arg Asn Asp Tyr Asp Gly Phe Leu Glu Asn Ala His 85 90 95 Glu His His Gly Val Tyr Asn Gln Gly Arg Gly Ile Asp Ser Gly Glu 100 105 110 Arg Leu Met Gln Pro Thr Gln Met Ser Ala Gln Glu Asp Leu Gly Asp 115 120 125 Asp Thr Gly Ile His Val Ile Pro Thr Leu Asn Gly Asp Asp Arg His 130 135 140 Lys Ile Val Asn Val Asp Gln Arg Gln Tyr Gly Asp Val Phe Lys Gly 145 150 155 160 Asp Leu Asn Pro Lys Pro Gln Gly Gln Arg Leu Ile Glu Val Ser Val 165 170 175 Glu Glu Asn His Pro Phe Thr Leu Arg Ala Pro Ile Gln Arg Ile Tyr 180 185 190 Gly Val Arg Tyr Thr Glu Thr Trp Ser Phe Leu Pro Ser Leu Thr Cys 195 200 205 Thr Gly Asp Ala Ala Pro Ala Ile Gln His Ile Cys Leu Lys His Thr 210 215 220 Thr Cys Phe Gln Asp Val Val Val Asp Val Asp Cys Ala Glu Asn Thr 225 230 235 240 Lys Glu Asp Gln Leu Ala Glu Ile Ser Tyr Arg Phe Gln Gly Lys Lys 245 250 255 Glu Ala Asp Gln Pro Trp Ile Val Val Asn Thr Ser Thr Leu Phe Asp 260 265 270 Glu Leu Glu Leu Asp Pro Pro Glu Ile Glu Pro Gly Val Leu Lys Val 275 280 285 Leu Arg Thr Glu Lys Gln Tyr Leu Gly Val Tyr Ile Trp Asn Met Arg 290 295 300 Gly Ser Asp Gly Thr Ser Thr Tyr Ala Thr Phe Leu Val Thr Trp Lys 305 310 315 320 Gly Asp Glu Lys Thr Arg Asn Pro Thr Pro Ala Val Thr Pro Gln Pro 325 330 335 Arg Gly Ala Glu Phe His Met Trp Asn Tyr His Ser His Val Phe Ser 340 345 350 Val Gly Asp Thr Phe Ser Leu Ala Met His Leu Gln Tyr Lys Ile His 355 360 365 Glu Ala Pro Phe Asp Leu Leu Leu Glu Trp Leu Tyr Val Pro Ile Asp 370 375 380 Pro Thr Cys Gln Pro Met Arg Leu Tyr Ser Thr Cys Leu Tyr His Pro 385 390 395 400 Asn Ala Pro Gln Cys Leu Ser His Met Asn Ser Gly Cys Thr Phe Thr 405 410 415 Ser Pro His Leu Ala Gln Arg Val Ala Ser Thr Val Tyr Gln Asn Cys 420 425 430 Glu His Ala Asp Asn Tyr Thr Ala Tyr Cys Leu Gly Ile Ser His Met 435 440 445 Glu Pro Ser Phe Gly Leu Ile Leu His Asp Gly Gly Thr Thr Leu Lys 450 455 460 Phe Val Asp Thr Pro Glu Ser Leu Ser Gly Leu Tyr Val Phe Val Val 465 470 475 480 Tyr Phe Asn Gly His Val Glu Ala Val Ala Tyr Thr Val Val Ser Thr 485 490 495 Val Asp His Phe Val Asn Ala Ile Glu Glu Arg Gly Phe Pro 500 505 510 <210> 19 <211> 515 <212> PRT <213> Artificial Sequence <220> <223> Artificial Sequence <400> 19 Met Gly Thr Val Asn Lys Pro Val Val Gly Val Leu Met Gly Phe Gly 1 5 10 15 Ile Ile Thr Gly Thr Leu Arg Ile Thr Asn Pro Val Arg Ala Ser Val 20 25 30 Leu Arg Tyr Asp Asp Phe His Thr Asp Glu Asp Lys Leu Asp Thr Asn 35 40 45 Ser Val Tyr Glu Pro Tyr Tyr His Ser Asp His Ala Glu Ser Ser Trp 50 55 60 Val Asn Arg Gly Glu Ser Ser Arg Lys Ala Tyr Asp His Asn Ser Pro 65 70 75 80 Tyr Ile Trp Pro Arg Asn Asp Tyr Asp Gly Phe Leu Glu Asn Ala His 85 90 95 Glu His His Gly Val Tyr Asn Gln Gly Arg Gly Ile Asp Ser Gly Glu 100 105 110 Arg Leu Met Gln Pro Thr Gln Met Ser Ala Gln Glu Asp Leu Gly Asp 115 120 125 Asp Thr Gly Ile His Val Ile Pro Thr Leu Asn Gly Asp Asp Arg His 130 135 140 Lys Ile Val Asn Val Asp Gln Arg Gln Tyr Gly Asp Val Phe Lys Gly 145 150 155 160 Asp Leu Asn Pro Lys Pro Gln Gly Gln Arg Leu Ile Glu Val Ser Val 165 170 175 Glu Glu Asn His Pro Phe Thr Leu Arg Ala Pro Ile Gln Arg Ile Tyr 180 185 190 Gly Val Arg Tyr Thr Glu Thr Trp Ser Phe Leu Pro Ser Leu Thr Cys 195 200 205 Thr Gly Asp Ala Ala Pro Ala Ile Gln His Ile Cys Leu Lys His Thr 210 215 220 Thr Cys Phe Gln Asp Val Val Val Asp Val Asp Cys Ala Glu Asn Thr 225 230 235 240 Lys Glu Asp Gln Leu Ala Glu Ile Ser Tyr Arg Phe Gln Gly Lys Lys 245 250 255 Glu Ala Asp Gln Pro Trp Ile Val Val Asn Thr Ser Thr Leu Phe Asp 260 265 270 Glu Leu Glu Leu Asp Pro Pro Glu Ile Glu Pro Gly Val Leu Lys Val 275 280 285 Leu Arg Thr Glu Lys Gln Tyr Leu Gly Val Tyr Ile Trp Asn Met Arg 290 295 300 Gly Ser Asp Gly Thr Ser Thr Tyr Ala Thr Phe Leu Val Thr Trp Lys 305 310 315 320 Gly Asp Glu Lys Thr Arg Asn Pro Thr Pro Ala Val Thr Pro Gln Pro 325 330 335 Arg Gly Ala Glu Phe His Met Trp Asn Tyr His Ser His Val Phe Ser 340 345 350 Val Gly Asp Thr Phe Ser Leu Ala Met His Leu Gln Tyr Lys Ile His 355 360 365 Glu Ala Pro Phe Asp Leu Leu Leu Glu Trp Leu Tyr Val Pro Ile Asp 370 375 380 Pro Thr Cys Gln Pro Met Arg Leu Tyr Ser Thr Cys Leu Tyr His Pro 385 390 395 400 Asn Ala Pro Gln Cys Leu Ser His Met Asn Ser Gly Cys Thr Phe Thr 405 410 415 Ser Pro His Leu Ala Gln Arg Val Ala Ser Thr Val Tyr Gln Asn Cys 420 425 430 Glu His Ala Asp Asn Tyr Thr Ala Tyr Cys Leu Gly Ile Ser His Met 435 440 445 Glu Pro Ser Phe Gly Leu Ile Leu His Asp Gly Gly Thr Thr Leu Lys 450 455 460 Phe Val Asp Thr Pro Glu Ser Leu Ser Gly Leu Tyr Val Phe Val Val 465 470 475 480 Tyr Phe Asn Gly His Val Glu Ala Val Ala Tyr Thr Val Val Ser Thr 485 490 495 Val Asp His Phe Val Asn Ala Ile Glu Glu Arg Gly Phe Pro Pro Thr 500 505 510 Ala Gly Gln 515 <210> 20 <211> 520 <212> PRT <213> Artificial Sequence <220> <223> Artificial Sequence <400> 20 Met Gly Thr Val Asn Lys Pro Val Val Gly Val Leu Met Gly Phe Gly 1 5 10 15 Ile Ile Thr Gly Thr Leu Arg Ile Thr Asn Pro Val Arg Ala Ser Val 20 25 30 Leu Arg Tyr Asp Asp Phe His Thr Asp Glu Asp Lys Leu Asp Thr Asn 35 40 45 Ser Val Tyr Glu Pro Tyr Tyr His Ser Asp His Ala Glu Ser Ser Trp 50 55 60 Val Asn Arg Gly Glu Ser Ser Arg Lys Ala Tyr Asp His Asn Ser Pro 65 70 75 80 Tyr Ile Trp Pro Arg Asn Asp Tyr Asp Gly Phe Leu Glu Asn Ala His 85 90 95 Glu His His Gly Val Tyr Asn Gln Gly Arg Gly Ile Asp Ser Gly Glu 100 105 110 Arg Leu Met Gln Pro Thr Gln Met Ser Ala Gln Glu Asp Leu Gly Asp 115 120 125 Asp Thr Gly Ile His Val Ile Pro Thr Leu Asn Gly Asp Asp Arg His 130 135 140 Lys Ile Val Asn Val Asp Gln Arg Gln Tyr Gly Asp Val Phe Lys Gly 145 150 155 160 Asp Leu Asn Pro Lys Pro Gln Gly Gln Arg Leu Ile Glu Val Ser Val 165 170 175 Glu Glu Asn His Pro Phe Thr Leu Arg Ala Pro Ile Gln Arg Ile Tyr 180 185 190 Gly Val Arg Tyr Thr Glu Thr Trp Ser Phe Leu Pro Ser Leu Thr Cys 195 200 205 Thr Gly Asp Ala Ala Pro Ala Ile Gln His Ile Cys Leu Lys His Thr 210 215 220 Thr Cys Phe Gln Asp Val Val Val Asp Val Asp Cys Ala Glu Asn Thr 225 230 235 240 Lys Glu Asp Gln Leu Ala Glu Ile Ser Tyr Arg Phe Gln Gly Lys Lys 245 250 255 Glu Ala Asp Gln Pro Trp Ile Val Val Asn Thr Ser Thr Leu Phe Asp 260 265 270 Glu Leu Glu Leu Asp Pro Pro Glu Ile Glu Pro Gly Val Leu Lys Val 275 280 285 Leu Arg Thr Glu Lys Gln Tyr Leu Gly Val Tyr Ile Trp Asn Met Arg 290 295 300 Gly Ser Asp Gly Thr Ser Thr Tyr Ala Thr Phe Leu Val Thr Trp Lys 305 310 315 320 Gly Asp Glu Lys Thr Arg Asn Pro Thr Pro Ala Val Thr Pro Gln Pro 325 330 335 Arg Gly Ala Glu Phe His Met Trp Asn Tyr His Ser His Val Phe Ser 340 345 350 Val Gly Asp Thr Phe Ser Leu Ala Met His Leu Gln Tyr Lys Ile His 355 360 365 Glu Ala Pro Phe Asp Leu Leu Leu Glu Trp Leu Tyr Val Pro Ile Asp 370 375 380 Pro Thr Cys Gln Pro Met Arg Leu Tyr Ser Thr Cys Leu Tyr His Pro 385 390 395 400 Asn Ala Pro Gln Cys Leu Ser His Met Asn Ser Gly Cys Thr Phe Thr 405 410 415 Ser Pro His Leu Ala Gln Arg Val Ala Ser Thr Val Tyr Gln Asn Cys 420 425 430 Glu His Ala Asp Asn Tyr Thr Ala Tyr Cys Leu Gly Ile Ser His Met 435 440 445 Glu Pro Ser Phe Gly Leu Ile Leu His Asp Gly Gly Thr Thr Leu Lys 450 455 460 Phe Val Asp Thr Pro Glu Ser Leu Ser Gly Leu Tyr Val Phe Val Val 465 470 475 480 Tyr Phe Asn Gly His Val Glu Ala Val Ala Tyr Thr Val Val Ser Thr 485 490 495 Val Asp His Phe Val Asn Ala Ile Glu Glu Arg Gly Phe Pro Pro Thr 500 505 510 Ala Gly Gln Pro Pro Ala Thr Thr 515 520 <210> 21 <211> 525 <212> PRT <213> Artificial Sequence <220> <223> Artificial Sequence <400> 21 Met Gly Thr Val Asn Lys Pro Val Val Gly Val Leu Met Gly Phe Gly 1 5 10 15 Ile Ile Thr Gly Thr Leu Arg Ile Thr Asn Pro Val Arg Ala Ser Val 20 25 30 Leu Arg Tyr Asp Asp Phe His Thr Asp Glu Asp Lys Leu Asp Thr Asn 35 40 45 Ser Val Tyr Glu Pro Tyr Tyr His Ser Asp His Ala Glu Ser Ser Trp 50 55 60 Val Asn Arg Gly Glu Ser Ser Arg Lys Ala Tyr Asp His Asn Ser Pro 65 70 75 80 Tyr Ile Trp Pro Arg Asn Asp Tyr Asp Gly Phe Leu Glu Asn Ala His 85 90 95 Glu His His Gly Val Tyr Asn Gln Gly Arg Gly Ile Asp Ser Gly Glu 100 105 110 Arg Leu Met Gln Pro Thr Gln Met Ser Ala Gln Glu Asp Leu Gly Asp 115 120 125 Asp Thr Gly Ile His Val Ile Pro Thr Leu Asn Gly Asp Asp Arg His 130 135 140 Lys Ile Val Asn Val Asp Gln Arg Gln Tyr Gly Asp Val Phe Lys Gly 145 150 155 160 Asp Leu Asn Pro Lys Pro Gln Gly Gln Arg Leu Ile Glu Val Ser Val 165 170 175 Glu Glu Asn His Pro Phe Thr Leu Arg Ala Pro Ile Gln Arg Ile Tyr 180 185 190 Gly Val Arg Tyr Thr Glu Thr Trp Ser Phe Leu Pro Ser Leu Thr Cys 195 200 205 Thr Gly Asp Ala Ala Pro Ala Ile Gln His Ile Cys Leu Lys His Thr 210 215 220 Thr Cys Phe Gln Asp Val Val Val Asp Val Asp Cys Ala Glu Asn Thr 225 230 235 240 Lys Glu Asp Gln Leu Ala Glu Ile Ser Tyr Arg Phe Gln Gly Lys Lys 245 250 255 Glu Ala Asp Gln Pro Trp Ile Val Val Asn Thr Ser Thr Leu Phe Asp 260 265 270 Glu Leu Glu Leu Asp Pro Pro Glu Ile Glu Pro Gly Val Leu Lys Val 275 280 285 Leu Arg Thr Glu Lys Gln Tyr Leu Gly Val Tyr Ile Trp Asn Met Arg 290 295 300 Gly Ser Asp Gly Thr Ser Thr Tyr Ala Thr Phe Leu Val Thr Trp Lys 305 310 315 320 Gly Asp Glu Lys Thr Arg Asn Pro Thr Pro Ala Val Thr Pro Gln Pro 325 330 335 Arg Gly Ala Glu Phe His Met Trp Asn Tyr His Ser His Val Phe Ser 340 345 350 Val Gly Asp Thr Phe Ser Leu Ala Met His Leu Gln Tyr Lys Ile His 355 360 365 Glu Ala Pro Phe Asp Leu Leu Leu Glu Trp Leu Tyr Val Pro Ile Asp 370 375 380 Pro Thr Cys Gln Pro Met Arg Leu Tyr Ser Thr Cys Leu Tyr His Pro 385 390 395 400 Asn Ala Pro Gln Cys Leu Ser His Met Asn Ser Gly Cys Thr Phe Thr 405 410 415 Ser Pro His Leu Ala Gln Arg Val Ala Ser Thr Val Tyr Gln Asn Cys 420 425 430 Glu His Ala Asp Asn Tyr Thr Ala Tyr Cys Leu Gly Ile Ser His Met 435 440 445 Glu Pro Ser Phe Gly Leu Ile Leu His Asp Gly Gly Thr Thr Leu Lys 450 455 460 Phe Val Asp Thr Pro Glu Ser Leu Ser Gly Leu Tyr Val Phe Val Val 465 470 475 480 Tyr Phe Asn Gly His Val Glu Ala Val Ala Tyr Thr Val Val Ser Thr 485 490 495 Val Asp His Phe Val Asn Ala Ile Glu Glu Arg Gly Phe Pro Pro Thr 500 505 510 Ala Gly Gln Pro Pro Ala Thr Thr Lys Pro Lys Glu Ile 515 520 525 <210> 22 <211> 530 <212> PRT <213> Artificial Sequence <220> <223> Artificial Sequence <400> 22 Met Gly Thr Val Asn Lys Pro Val Val Gly Val Leu Met Gly Phe Gly 1 5 10 15 Ile Ile Thr Gly Thr Leu Arg Ile Thr Asn Pro Val Arg Ala Ser Val 20 25 30 Leu Arg Tyr Asp Asp Phe His Thr Asp Glu Asp Lys Leu Asp Thr Asn 35 40 45 Ser Val Tyr Glu Pro Tyr Tyr His Ser Asp His Ala Glu Ser Ser Trp 50 55 60 Val Asn Arg Gly Glu Ser Ser Arg Lys Ala Tyr Asp His Asn Ser Pro 65 70 75 80 Tyr Ile Trp Pro Arg Asn Asp Tyr Asp Gly Phe Leu Glu Asn Ala His 85 90 95 Glu His His Gly Val Tyr Asn Gln Gly Arg Gly Ile Asp Ser Gly Glu 100 105 110 Arg Leu Met Gln Pro Thr Gln Met Ser Ala Gln Glu Asp Leu Gly Asp 115 120 125 Asp Thr Gly Ile His Val Ile Pro Thr Leu Asn Gly Asp Asp Arg His 130 135 140 Lys Ile Val Asn Val Asp Gln Arg Gln Tyr Gly Asp Val Phe Lys Gly 145 150 155 160 Asp Leu Asn Pro Lys Pro Gln Gly Gln Arg Leu Ile Glu Val Ser Val 165 170 175 Glu Glu Asn His Pro Phe Thr Leu Arg Ala Pro Ile Gln Arg Ile Tyr 180 185 190 Gly Val Arg Tyr Thr Glu Thr Trp Ser Phe Leu Pro Ser Leu Thr Cys 195 200 205 Thr Gly Asp Ala Ala Pro Ala Ile Gln His Ile Cys Leu Lys His Thr 210 215 220 Thr Cys Phe Gln Asp Val Val Val Asp Val Asp Cys Ala Glu Asn Thr 225 230 235 240 Lys Glu Asp Gln Leu Ala Glu Ile Ser Tyr Arg Phe Gln Gly Lys Lys 245 250 255 Glu Ala Asp Gln Pro Trp Ile Val Val Asn Thr Ser Thr Leu Phe Asp 260 265 270 Glu Leu Glu Leu Asp Pro Pro Glu Ile Glu Pro Gly Val Leu Lys Val 275 280 285 Leu Arg Thr Glu Lys Gln Tyr Leu Gly Val Tyr Ile Trp Asn Met Arg 290 295 300 Gly Ser Asp Gly Thr Ser Thr Tyr Ala Thr Phe Leu Val Thr Trp Lys 305 310 315 320 Gly Asp Glu Lys Thr Arg Asn Pro Thr Pro Ala Val Thr Pro Gln Pro 325 330 335 Arg Gly Ala Glu Phe His Met Trp Asn Tyr His Ser His Val Phe Ser 340 345 350 Val Gly Asp Thr Phe Ser Leu Ala Met His Leu Gln Tyr Lys Ile His 355 360 365 Glu Ala Pro Phe Asp Leu Leu Leu Glu Trp Leu Tyr Val Pro Ile Asp 370 375 380 Pro Thr Cys Gln Pro Met Arg Leu Tyr Ser Thr Cys Leu Tyr His Pro 385 390 395 400 Asn Ala Pro Gln Cys Leu Ser His Met Asn Ser Gly Cys Thr Phe Thr 405 410 415 Ser Pro His Leu Ala Gln Arg Val Ala Ser Thr Val Tyr Gln Asn Cys 420 425 430 Glu His Ala Asp Asn Tyr Thr Ala Tyr Cys Leu Gly Ile Ser His Met 435 440 445 Glu Pro Ser Phe Gly Leu Ile Leu His Asp Gly Gly Thr Thr Leu Lys 450 455 460 Phe Val Asp Thr Pro Glu Ser Leu Ser Gly Leu Tyr Val Phe Val Val 465 470 475 480 Tyr Phe Asn Gly His Val Glu Ala Val Ala Tyr Thr Val Val Ser Thr 485 490 495 Val Asp His Phe Val Asn Ala Ile Glu Glu Arg Gly Phe Pro Pro Thr 500 505 510 Ala Gly Gln Pro Pro Ala Thr Thr Lys Pro Lys Glu Ile Thr Pro Val 515 520 525 Asn Pro 530 <210> 23 <211> 543 <212> PRT <213> Artificial Sequence <220> <223> Artificial Sequence <400> 23 Met Gly Thr Val Asn Lys Pro Val Val Gly Val Leu Met Gly Phe Gly 1 5 10 15 Ile Ile Thr Gly Thr Leu Arg Ile Thr Asn Pro Val Arg Ala Ser Val 20 25 30 Leu Arg Tyr Asp Asp Phe His Thr Asp Glu Asp Lys Leu Asp Thr Asn 35 40 45 Ser Val Tyr Glu Pro Tyr Tyr His Ser Asp His Ala Glu Ser Ser Trp 50 55 60 Val Asn Arg Gly Glu Ser Ser Arg Lys Ala Tyr Asp His Asn Ser Pro 65 70 75 80 Tyr Ile Trp Pro Arg Asn Asp Tyr Asp Gly Phe Leu Glu Asn Ala His 85 90 95 Glu His His Gly Val Tyr Asn Gln Gly Arg Gly Ile Asp Ser Gly Glu 100 105 110 Arg Leu Met Gln Pro Thr Gln Met Ser Ala Gln Glu Asp Leu Gly Asp 115 120 125 Asp Thr Gly Ile His Val Ile Pro Thr Leu Asn Gly Asp Asp Arg His 130 135 140 Lys Ile Val Asn Val Asp Gln Arg Gln Tyr Gly Asp Val Phe Lys Gly 145 150 155 160 Asp Leu Asn Pro Lys Pro Gln Gly Gln Arg Leu Ile Glu Val Ser Val 165 170 175 Glu Glu Asn His Pro Phe Thr Leu Arg Ala Pro Ile Gln Arg Ile Tyr 180 185 190 Gly Val Arg Tyr Thr Glu Thr Trp Ser Phe Leu Pro Ser Leu Thr Cys 195 200 205 Thr Gly Asp Ala Ala Pro Ala Ile Gln His Ile Cys Leu Lys His Thr 210 215 220 Thr Cys Phe Gln Asp Val Val Val Asp Val Asp Cys Ala Glu Asn Thr 225 230 235 240 Lys Glu Asp Gln Leu Ala Glu Ile Ser Tyr Arg Phe Gln Gly Lys Lys 245 250 255 Glu Ala Asp Gln Pro Trp Ile Val Val Asn Thr Ser Thr Leu Phe Asp 260 265 270 Glu Leu Glu Leu Asp Pro Pro Glu Ile Glu Pro Gly Val Leu Lys Val 275 280 285 Leu Arg Thr Glu Lys Gln Tyr Leu Gly Val Tyr Ile Trp Asn Met Arg 290 295 300 Gly Ser Asp Gly Thr Ser Thr Tyr Ala Thr Phe Leu Val Thr Trp Lys 305 310 315 320 Gly Asp Glu Lys Thr Arg Asn Pro Thr Pro Ala Val Thr Pro Gln Pro 325 330 335 Arg Gly Ala Glu Phe His Met Trp Asn Tyr His Ser His Val Phe Ser 340 345 350 Val Gly Asp Thr Phe Ser Leu Ala Met His Leu Gln Tyr Lys Ile His 355 360 365 Glu Ala Pro Phe Asp Leu Leu Leu Glu Trp Leu Tyr Val Pro Ile Asp 370 375 380 Pro Thr Cys Gln Pro Met Arg Leu Tyr Ser Thr Cys Leu Tyr His Pro 385 390 395 400 Asn Ala Pro Gln Cys Leu Ser His Met Asn Ser Gly Cys Thr Phe Thr 405 410 415 Ser Pro His Leu Ala Gln Arg Val Ala Ser Thr Val Tyr Gln Asn Cys 420 425 430 Glu His Ala Asp Asn Tyr Thr Ala Tyr Cys Leu Gly Ile Ser His Met 435 440 445 Glu Pro Ser Phe Gly Leu Ile Leu His Asp Gly Gly Thr Thr Leu Lys 450 455 460 Phe Val Asp Thr Pro Glu Ser Leu Ser Gly Leu Tyr Val Phe Val Val 465 470 475 480 Tyr Phe Asn Gly His Val Glu Ala Val Ala Tyr Thr Val Val Ser Thr 485 490 495 Val Asp His Phe Val Asn Ala Ile Glu Glu Arg Gly Phe Pro Pro Thr 500 505 510 Ala Gly Gln Pro Pro Ala Thr Thr Lys Pro Lys Glu Ile Thr Pro Val 515 520 525 Asn Pro Gly Thr Ser Pro Leu Leu Arg Tyr Ala Ala Trp Thr Gly 530 535 540 <210> 24 <211> 546 <212> PRT <213> Artificial Sequence <220> <223> Artificial Sequence <400> 24 Met Gly Thr Val Asn Lys Pro Val Val Gly Val Leu Met Gly Phe Gly 1 5 10 15 Ile Ile Thr Gly Thr Leu Arg Ile Thr Asn Pro Val Arg Ala Ser Val 20 25 30 Leu Arg Tyr Asp Asp Phe His Thr Asp Glu Asp Lys Leu Asp Thr Asn 35 40 45 Ser Val Tyr Glu Pro Tyr Tyr His Ser Asp His Ala Glu Ser Ser Trp 50 55 60 Val Asn Arg Gly Glu Ser Ser Arg Lys Ala Tyr Asp His Asn Ser Pro 65 70 75 80 Tyr Ile Trp Pro Arg Asn Asp Tyr Asp Gly Phe Leu Glu Asn Ala His 85 90 95 Glu His His Gly Val Tyr Asn Gln Gly Arg Gly Ile Asp Ser Gly Glu 100 105 110 Arg Leu Met Gln Pro Thr Gln Met Ser Ala Gln Glu Asp Leu Gly Asp 115 120 125 Asp Thr Gly Ile His Val Ile Pro Thr Leu Asn Gly Asp Asp Arg His 130 135 140 Lys Ile Val Asn Val Asp Gln Arg Gln Tyr Gly Asp Val Phe Lys Gly 145 150 155 160 Asp Leu Asn Pro Lys Pro Gln Gly Gln Arg Leu Ile Glu Val Ser Val 165 170 175 Glu Glu Asn His Pro Phe Thr Leu Arg Ala Pro Ile Gln Arg Ile Tyr 180 185 190 Gly Val Arg Tyr Thr Glu Thr Trp Ser Phe Leu Pro Ser Leu Thr Cys 195 200 205 Thr Gly Asp Ala Ala Pro Ala Ile Gln His Ile Cys Leu Lys His Thr 210 215 220 Thr Cys Phe Gln Asp Val Val Val Asp Val Asp Cys Ala Glu Asn Thr 225 230 235 240 Lys Glu Asp Gln Leu Ala Glu Ile Ser Tyr Arg Phe Gln Gly Lys Lys 245 250 255 Glu Ala Asp Gln Pro Trp Ile Val Val Asn Thr Ser Thr Leu Phe Asp 260 265 270 Glu Leu Glu Leu Asp Pro Pro Glu Ile Glu Pro Gly Val Leu Lys Val 275 280 285 Leu Arg Thr Glu Lys Gln Tyr Leu Gly Val Tyr Ile Trp Asn Met Arg 290 295 300 Gly Ser Asp Gly Thr Ser Thr Tyr Ala Thr Phe Leu Val Thr Trp Lys 305 310 315 320 Gly Asp Glu Lys Thr Arg Asn Pro Thr Pro Ala Val Thr Pro Gln Pro 325 330 335 Arg Gly Ala Glu Phe His Met Trp Asn Tyr His Ser His Val Phe Ser 340 345 350 Val Gly Asp Thr Phe Ser Leu Ala Met His Leu Gln Tyr Lys Ile His 355 360 365 Glu Ala Pro Phe Asp Leu Leu Leu Glu Trp Leu Tyr Val Pro Ile Asp 370 375 380 Pro Thr Cys Gln Pro Met Arg Leu Tyr Ser Thr Cys Leu Tyr His Pro 385 390 395 400 Asn Ala Pro Gln Cys Leu Ser His Met Asn Ser Gly Cys Thr Phe Thr 405 410 415 Ser Pro His Leu Ala Gln Arg Val Ala Ser Thr Val Tyr Gln Asn Cys 420 425 430 Glu His Ala Asp Asn Tyr Thr Ala Tyr Cys Leu Gly Ile Ser His Met 435 440 445 Glu Pro Ser Phe Gly Leu Ile Leu His Asp Gly Gly Thr Thr Leu Lys 450 455 460 Phe Val Asp Thr Pro Glu Ser Leu Ser Gly Leu Tyr Val Phe Val Val 465 470 475 480 Tyr Phe Asn Gly His Val Glu Ala Val Ala Tyr Thr Val Val Ser Thr 485 490 495 Val Asp His Phe Val Asn Ala Ile Glu Glu Arg Gly Phe Pro Pro Thr 500 505 510 Ala Gly Gln Pro Pro Ala Thr Thr Lys Pro Lys Glu Ile Thr Pro Val 515 520 525 Asn Pro Gly Thr Ser Pro Leu Leu Arg Tyr Ala Ala Trp Thr Gly Gly 530 535 540 Leu Ala 545 Description of the Drawings

[0023] Figure 1 The experimental process of the present invention is schematically shown.

[0024] Figure 2 Amino acid sequences of various lengths of the varicella zoster virus surface protein (gE) antigen are shown.

[0025] Figure 3 Shown are Western blot results comparing the expression levels of the gE fragment of varicella zoster virus antigen.

[0026] Figure 4 Shown are the results of comparing the expression levels of the surface protein antigen (GSK gE 546 amino acids) contained in the varicella zoster virus vaccine Shingrix produced by GlaxoSmithKline Biologicals, and the antigen variant (Mogam gE 537 amino acids) produced by the present inventors by Western blotting.

[0027] Figure 5 Shown are the results of humoral immune responses comparing the surface protein antigen (GSK gE 546 amino acids) contained in the varicella zoster virus vaccine Shingrix produced by GlaxoSmithKline Biologicals, and the antigen variant (Mogam gE 537 amino acids) produced by the present inventors by anti-gE-specific IgG ELISA.

[0028] Figure 6 Shown are the results of comparing the cell-mediated immune response (CMI) of the surface protein antigen contained in the varicella-zoster virus vaccine Shingrix produced by GlaxoSmithKline Biologicals (GSK gE 546 amino acids) and the antigen variant produced by the present inventors (Mogam gE 537 amino acids) by mouse IFN-γ ELISA.

[0029] Figure 7 The results of comparing humoral immune responses to the gE fragment of varicella zoster virus antigen by anti-gE-specific IgG ELISA are presented.

[0030] Figure 8 Shown are the results of a comparison of humoral immune responses to gE fragments as varicella zoster virus antigens by anti-gE-specific IgG ELISA, followed by a summary of the number of responders showing antigen-specific antibody responses.

[0031] Best Mode for Carrying Out the Invention

[0032] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the invention belongs. Generally, the terms used herein are well known and commonly used in the art.

[0033] In the present invention, in order to improve productivity without affecting immunogenicity, an antigen with a high expression level in host cells was developed from a varicella zoster virus protein antigen, and an experiment was conducted by generating surface protein (gE) antigen variants of different lengths and then measuring their expression levels. In the results, among the varicella zoster virus surface protein antigen variants produced by the present inventors, the antigen represented by a specific amino acid sequence showed a higher expression level than other antigens ( Figure 3 ).

[0034] In addition, antibody titer was measured ( Figure 7 ) and antigen-specific responder measurements ( Figure 8 ) to select antigen variants with high immunogenicity. In the results, it was determined that Mogam gE 534 amino acids to gE 540 amino acids had higher immunogenicity.

[0035] Therefore, in one aspect of the present invention, a varicella-zoster virus surface protein antigen variant is provided, comprising a variant consisting of a truncation of the carboxyl terminus of any amino acid residue from amino acids 525 to 543 of the varicella-zoster virus surface protein (gE) antigen represented by the amino acid sequence shown in SEQ ID NO: 1.

[0036] Specifically, in the present invention, a varicella-zoster virus surface protein antigen variant is provided, characterized in that the antigen variant includes a variant selected from the following group:

[0037] a) truncation of the carboxyl terminus at amino acid residue 525;

[0038] b) truncation of the carboxyl terminus at amino acid residue 526;

[0039] c) carboxyl-terminal truncation of amino acid residue 527;

[0040] d) carboxyl-terminal truncation at amino acid residue 528;

[0041] e) truncation of the carboxyl terminus at amino acid residue 529;

[0042] f) carboxyl-terminal truncation at amino acid residue 530;

[0043] g) truncation of the carboxyl terminus of amino acid residue 531;

[0044] h) truncation of the carboxyl terminus at amino acid residue 532;

[0045] i) truncation of the carboxyl terminus at amino acid residue 533;

[0046] j) truncation of the carboxyl terminus at amino acid residue 534;

[0047] k) carboxyl-terminal truncation of amino acid residue 535;

[0048] 1) truncation of the carboxyl terminus of amino acid residue 536;

[0049] m) carboxyl-terminal truncation of amino acid residue 537;

[0050] n) truncation of the carboxyl terminus at amino acid residue 538;

[0051] o) carboxyl-terminal truncation at amino acid residue 539;

[0052] p) carboxyl-terminal truncation of amino acid residue 540;

[0053] q) truncation of the carboxyl terminus at amino acid residue 541;

[0054] r) carboxyl-terminal truncation of amino acid residue 542; and

[0055] s) Carboxyl-terminal truncation at amino acid residue 543.

[0056] The antigen variant is characterized in that it preferably includes a variant selected from the following groups: j) truncation of the carboxyl terminal of the amino acid residue 534; k) truncation of the carboxyl terminal of the amino acid residue 535; l) truncation of the carboxyl terminal of the amino acid residue 536; m) truncation of the carboxyl terminal of the amino acid residue 537; n) truncation of the carboxyl terminal of the amino acid residue 538; o) truncation of the carboxyl terminal of the amino acid residue 539; p) truncation of the carboxyl terminal of the amino acid residue 540; however, the changes are not limited to this.

[0057] SEQ ID NO: 1:

[0058] mgtvnkpvvg vlmgfgiitg tlritnpvra svlryddfhX1 dedkldtnsv yepyyhsdhaesswvnrges srkaydhnsp yiwprndydg flenahehhg vynqgrgids gerlmqptqm saqedlgddtgihviptlng ddrhkivnvd qrqygdvfkg dlnpkpqgqr lievsveenh pftlrapiqr iygvrytetwsflpsltctg daapaiqhic lkhttcfqdv vvdvdcaent kedqlaeisy rfqgkkeadq pwivvntstlfdeleldppe iepgvlkvlr tekqylgvyi wnmrgsdgts tyatflvtwk gdektrnptp avtpqprgaefhmwnyhshv fsvgdtfsla mhlqykihea pfdlllewly vpidptcqpm rlystclyhp napqclshmnsgctftsphl aqrvastvyq ncehadnyta yclgishmep sfglilhdgg ttlkfvdtpe slsglyvfvvyfnghveava ytvvstvdhf vnaieergfp ptagqppatt kpkeitpvnp gtsplX2ryaa wtgglaavvllclviflict akrmrvkayr vdkspynqsm yyaglpvddf edsestdtee efgnaiggsh ggssytvyidktr (where X1 is T or I, and X2 is L or I).

[0059] In the present invention, the varicella zoster virus surface protein antigen variant is characterized in that it is a varicella zoster virus surface protein antigen variant consisting of 534 to 540 amino acids, which is derived from the varicella zoster virus surface protein antigen of the amino acid sequence shown in SEQ ID NO: 1, which consists of 623 amino acids, or a variant in which the carboxyl terminus is truncated in some amino acid residues. For example, as used herein, the term "a variant in which the carboxyl terminus of the 534th amino acid residue is truncated" means that in the direction from the amino terminus (N terminus) to the carboxyl terminus (C terminus), the 1st to 534th amino acid residues are retained, while the continuous amino acid residues from the 535th amino acid residue to the carboxyl terminus are truncated.

[0060] According to a specific embodiment of the present invention, the 40th amino acid residue in the amino acid sequence shown by SEQ ID NO: 1 is threonine.

[0061] According to a specific embodiment of the present invention, the amino acid residue at position 536 in the amino acid sequence shown by SEQ ID NO: 1 is leucine.

[0062] In the present invention, the varicella-zoster virus surface protein antigen variant is characterized by being represented by any one of the amino acid sequences of SEQ ID NOs: 2 to 8 and SEQ ID NOs: 21 to 23, as shown below:

[0063] a) an antigen variant represented by the amino acid sequence shown in SEQ ID NO: 2, which is obtained by truncating the carboxyl terminus of the 534th amino acid residue;

[0064] b) an antigen variant represented by the amino acid sequence shown in SEQ ID NO: 3, which is obtained by truncating the carboxyl terminus of the 535th amino acid residue;

[0065] c) an antigen variant represented by the amino acid sequence shown in SEQ ID NO: 4, which is obtained by truncating the carboxyl terminus of the 536th amino acid residue;

[0066] d) an antigen variant represented by the amino acid sequence shown in SEQ ID NO: 5, which is obtained by truncating the carboxyl terminus of the 537th amino acid residue;

[0067] e) an antigen variant represented by the amino acid sequence shown in SEQ ID NO: 6, which is obtained by truncating the carboxyl terminus of the 538th amino acid residue;

[0068] f) an antigen variant represented by the amino acid sequence shown in SEQ ID NO: 7, which is obtained by truncating the carboxyl terminus of the 539th amino acid residue;

[0069] g) an antigen variant represented by the amino acid sequence shown in SEQ ID NO: 8, which is obtained by truncating the carboxyl terminus of the 540th amino acid residue;

[0070] h) an antigen variant represented by the amino acid sequence shown in SEQ ID NO: 21, which is obtained by truncating the carboxyl terminus of the 525th amino acid residue;

[0071] i) an antigen variant represented by the amino acid sequence shown in SEQ ID NO: 22, which is obtained by truncating the carboxyl terminus of the 530th amino acid residue; or

[0072] j) An antigen variant represented by the amino acid sequence shown in SEQ ID NO: 23, which is obtained by truncating the carboxyl terminus of the 543rd amino acid residue.

[0073] The glycoprotein from which the surface protein of the present invention is derived is a component of the envelope of varicella-zoster virus of evolutionary branch 1, and is a peptide fragment (truncated protein) consisting of 534 to 540 amino acids, which is obtained by partial truncation of the carboxyl terminus.

[0074] Taking into account biologically equivalent amino acid changes, the amino acid sequences used in the present invention are understood to include sequences having substantial identity with the sequences of SEQ ID NOs: 2 to 8 and SEQ ID NOs: 21 to 23. The above substantial identity means that when the sequence of the present invention as described above is aligned with any other sequence to obtain maximum correspondence and the sequences aligned using algorithms commonly used in the art are analyzed, the other sequences have at least 70% homology, more particularly 80% homology, even more particularly 90% homology, and most particularly 95% homology with the sequence of the present invention while having the same function.

[0075] In one embodiment of the present invention, the following antigen variants are identified as having high immunogenicity: a) an antigen variant represented by the amino acid sequence shown in SEQ ID NO: 2, which is obtained by truncating the carboxyl terminus of the 534th amino acid residue; b) an antigen variant represented by the amino acid sequence shown in SEQ ID NO: 3, which is obtained by truncating the carboxyl terminus of the 535th amino acid residue; c) an antigen variant represented by the amino acid sequence shown in SEQ ID NO: 4, which is obtained by truncating the carboxyl terminus of the 536th amino acid residue; d) an antigen variant represented by the amino acid sequence shown in SEQ ID NO: 5, which is obtained by truncating the carboxyl terminus of the 537th amino acid residue; e) an antigen variant represented by the amino acid sequence shown in SEQ ID NO: 6, which is obtained by truncating the carboxyl terminus of the 538th amino acid residue; f) an antigen variant represented by the amino acid sequence shown in SEQ ID NO: 7, which is obtained by truncating the carboxyl terminus of the 539th amino acid residue; or g) an antigen variant represented by the amino acid sequence shown in SEQ ID NO: 8 The antigen variant represented by the amino acid sequence shown in NO:8 is obtained by truncating the carboxyl terminus of the 540th amino acid residue.

[0076] In the early stage of vaccine development, attenuated live cells or dead cells were mainly used as antigens. However, due to safety issues, the development of such antigens is turning to the development of protein antigens with clear structure and composition. However, protein antigens are generally problematic because they have low immunogenicity compared to conventional vaccines. From the perspective of excellent stability and immunogenicity and high-level expression in host cells, the antigen variants of the present invention can be effectively used as vaccines for preventing or treating chickenpox or herpes zoster induced by varicella-zoster virus.

[0077] In another aspect of the present invention, provided are genes encoding antigen variants, recombinant vectors comprising the antigen variants, and host cells transformed with the recombinant vectors.

[0078] In the present invention, the gene is characterized in that it can be represented by any one of the nucleotide sequences of SEQ ID NOs: 9 to 15.

[0079] As used herein, the term "vector" refers to a DNA construct comprising a DNA sequence that is operably linked to a suitable control sequence that enables expression of the DNA sequence in a suitable host. A vector may be a plasmid, a phage particle, or simply a potential genomic insert. Once transformed into a suitable host, the vector may replicate and function independently of the host genome, or may be integrated into its genome in some cases. In this specification, "plasmid" and "vector" are sometimes used interchangeably because plasmids are currently the most commonly used vector form. For the purposes of the present invention, plasmid vectors are preferably used. Typical plasmid vectors that can be used for this purpose have the following structure, including: (a) an origin of replication that allows efficient replication so that hundreds of plasmid vectors can be produced per host cell; (b) an antibiotic resistance gene that allows selection of host cells transformed with the plasmid vector, and (c) a restriction enzyme site that allows insertion of exogenous DNA fragments. Even if there is no suitable restriction enzyme cleavage site in the vector, the vector can be easily connected to the exogenous DNA using a synthetic oligonucleotide adapter or joint according to conventional methods.

[0080] As used herein, the term "recombinant vector" generally refers to a recombinant vector into which a heterologous DNA fragment is inserted, which is generally in the form of a double-stranded DNA fragment. Here, heterologous DNA refers to foreign DNA that is not naturally found in the host cell. Once the recombinant vector enters the host cell, it can replicate independently of the host chromosomal DNA, thereby producing several copies of the vector and the (heterologous) DNA inserted therein.

[0081] After ligation, the gene or recombinant vector is transformed or transfected into the host cell. For "transformation" or "transfection", several types of techniques commonly used to introduce exogenous nucleic acid (DNA or RNA) into prokaryotic or eukaryotic host cells can be used, examples of which include electroporation, calcium phosphate precipitation, DEAE-dextran transfection and lipofection.

[0082] As is well known in the art, to increase the expression level of a transfected gene in a host cell, the gene in question should be operably linked to transcriptional and translational expression control sequences that function in the selected expression host.

[0083] As used herein, the term "transformation" refers to the introduction of DNA into a host so that the DNA can be replicated as an extrachromosomal factor or by chromosomal integration. Of course, it should be understood that not all vectors have the same function when expressing the gene sequence of the present invention. Similarly, not all host cells have equivalent functions relative to the same expression system. However, those skilled in the art can make appropriate choices between various vectors, expression control sequences and hosts without excessive experimental burden within the scope of the present invention. For example, the host must be considered and the vector must be selected because the vector must be replicated therein. In this regard, the copy number of the vector, its ability to regulate the copy number and the expression of other proteins encoded by the vector must also be considered.

[0084] In the present invention, the host cell to be transformed is preferably selected from, but not limited to, the following group: animal cells, plant cells, yeast, Escherichia coli and insect cells.

[0085] Specifically, in the present invention, when it is applied to a living body, as long as it is a non-toxic or attenuated microorganism, any microorganism can be used as the microorganism to be transformed into a host cell. Examples thereof may include gram-negative bacteria, such as Escherichia coli (E.coli), Salmonella typhi, Salmonella typhimurium, Vibrio cholerae, Mycobacterium bovis and Shigella; Gram-positive bacteria, such as Bacillus, Lactobacillus, Lactococcus, Staphylococcus, Listeria monocytogenes and Streptococcus. Its preferred example may include lactic acid bacteria as edible microorganisms. However, microorganisms are not limited thereto.

[0086] The lactic acid bacteria may include Lactobacillus, Streptococcus and Bifidobacterium. Representative examples of the genus Lactobacillus may include Lactobacillus acidophilus, Lactobacillus plantarum, Lactobacillus fermentum, Lactobacillus delbrueckii, Lactobacillus johnsonii, Lactobacillus reuteri, Lactobacillus bulgaricus, and Lactobacillus casei; representative species of Streptococcus may include Streptococcus thermophiles; and representative examples of Bifidobacterium may include Bifidobacterium infantis, Bifidobacterium bifidum, Bifidobacterium longum, Bifidobacterium breve, Bifidobacterium lactis. lactis and Bifidobacterium adolescentis, more preferably Lactobacillus casei. However, the lactic acid bacteria are not limited thereto.

[0087] The microorganism can also be a eukaryotic cell, including fungi, such as Aspergillus sp.; yeasts, such as Pichia pastoris, Saccharomyces cerevisiae, Schizosaccharomyces sp., and Neurospora crassa, other lower eukaryotic cells, and higher eukaryotic cells, such as cells from insects.

[0088] Microorganism can be derived from plant or mammal.Preferred examples thereof can include monkey kidney cell (COS-7 cell), NS0 cell, SP2 / 0, Chinese hamster ovary (CHO) cell, W138, hamster kidney (BHK) cell, MDCK, myeloma cell line, HuT78 cell and HEK293 cell, wherein CHO cell is preferred.However, microorganism is not limited thereto.

[0089] Another aspect of the present invention provides a method for producing varicella-zoster virus antigens, the method comprising the step of culturing host cells.

[0090] Where a recombinant vector capable of expressing a varicella zoster virus antigen is introduced into a host cell, the antigen can be produced by culturing the host cells for a time sufficient to allow the antigen to be expressed therein, or more preferably, for a time sufficient to allow the antigen to be secreted into the medium in which the host cells are cultured.

[0091] In some cases, the antigen expressed can be separated from the host cell and purified to homogeneity. The separation or purification of the antigen can be carried out by conventional separation and purification methods for proteins, such as chromatography. The example of chromatography can include affinity chromatography, including protein A column or protein G column, ion exchange chromatography and hydrophobic chromatography. In addition to the above-mentioned chromatography, the antigen can also be separated and purified by further combination filtration, ultrafiltration, salting out, dialysis, etc.

[0092] In yet another aspect of the present invention, a vaccine composition for preventing or treating varicella or herpes zoster is provided, comprising a varicella zoster virus antigen variant as an active ingredient.

[0093] In another aspect of the present invention, a method for preventing or treating varicella or herpes zoster is provided, wherein the vaccine composition comprises a varicella zoster virus antigen variant as an active ingredient.

[0094] In another aspect of the present invention, a use of a vaccine composition is provided, wherein the vaccine composition comprises a varicella-zoster virus antigen variant as an active ingredient, for preventing or treating varicella or herpes zoster.

[0095] In yet another aspect of the present invention, there is provided use of a vaccine composition comprising an antigen variant as an active ingredient in the preparation of a medicament for preventing or treating varicella or herpes zoster.

[0096] As used herein, the term "preventing" refers to inhibiting the occurrence of a condition or disease in a subject who has not yet been diagnosed as having the condition or disease and who is likely to have the condition or disease.

[0097] As used herein, the term "treatment" refers to (a) inhibiting the progression of a condition or disease or its symptoms; (b) alleviating a condition or disease or its symptoms; (c) eliminating a condition or disease or its symptoms. The composition of the present invention activates an immune response against the varicella zoster virus in an individual suffering from chickenpox or herpes zoster caused by varicella zoster virus infection, thereby inhibiting the progression of the disease, eliminating or alleviating its symptoms. Therefore, the composition of the present invention itself can be a therapeutic composition for chickenpox or herpes zoster, or can be used as a therapeutic adjuvant for a disease administered in combination with other pharmacological ingredients.

[0098] Therefore, in the present specification, the term "treatment" or "therapeutic agent" also includes the meaning of "adjuvant therapy" or "therapy adjuvant".

[0099] As used herein, the term "active ingredient" refers to a vaccine composition sufficient to produce a desired effect, including but not limited to inducing or increasing an immune response to varicella zoster virus in a patient, preventing, alleviating or eliminating reactivation of varicella zoster virus in a patient infected with the same virus or vaccinated with a live varicella zoster virus vaccine, preventing herpes zoster (HZ) and / or postherpetic neuralgia (PHN), and reducing the severity or duration of HZ and / or PHN. It will be appreciated by those skilled in the art that the level of such desired effect may vary.

[0100] As used herein, the term "immune response" refers to a cell-mediated (T cell) immune response and / or an antibody (B cell) response.

[0101] The vaccine compositions of the present invention can be used to prevent varicella and / or HZ and / or PHN, or reduce the severity or duration of varicella and / or HZ and / or PHN in immunocompetent and immunocompromised patient populations, including but not limited to healthy individuals and immunocompromised patients undergoing hematopoietic cell transplantation (HCT) or solid organ transplantation (SOT), patients infected with HIV, patients with autoimmune diseases, and patients with blood cancers; individuals undergoing chemotherapy for various solid malignancies; and patients undergoing chronic immunosuppressive therapy for various diseases, including rheumatoid arthritis (RA), systemic lupus erythematosus (SLE), Crohn's disease, psoriasis, and multiple sclerosis.

[0102] In the present invention, the vaccine composition is characterized in that it further comprises a pharmaceutically acceptable carrier, excipient or diluent.

[0103] The vaccine composition of the present invention can be prepared in unit dosage form or in the form of a multi-dose container by using a pharmaceutically acceptable carrier and / or excipient for formulation according to methods that can be easily implemented by those skilled in the art. Here, the dosage form can be formulated in the form of oral preparations, such as powders, granules, tablets, capsules, suspensions, emulsions, syrups and aerosols, external preparations, suppositories and sterile injection solutions prepared and used according to conventional methods. Suitable formulations known in the art can be those disclosed in Remington's Pharmaceutical Sciences, published by Mark Publishing Company, Easton, Pennsylvania.

[0104] Solid preparations for oral administration include tablets, pills, powders, granules, capsules, etc., and these solid preparations are prepared by mixing with at least one excipient such as starch, calcium carbonate, sucrose, lactose and gelatin. In addition to simple excipients, solid preparations also use lubricants, such as magnesium stearate and talc.

[0105] Liquid preparations for oral administration include suspensions, oral liquids, emulsions, syrups, etc., and these liquid preparations may contain various excipients such as wetting agents, sweeteners, flavors and preservatives in addition to commonly used simple diluents such as water and liquid paraffin.

[0106] Preparations for parenteral administration include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilized preparations and suppositories. As the base of the suppository, Vitsol, polyethylene glycol, Tween 61, cocoa butter, lauric acid, glycerin gelatin and the like can be used.

[0107] In the present invention, the vaccine composition is characterized in that it further comprises an adjuvant. Generally, a protein antigen alone cannot strongly induce an immune response, so the effect of the vaccine composition can be improved by mixing with an adjuvant.

[0108] As used herein, the term "adjuvant" refers to a substance that non-specifically promotes the immune response to an antigen during the initial activation of immune cells, including agents, molecules, etc., each of which is not an immunogen to the host but enhances immunity by increasing the activity of cells in the immune system (Warren et al., Annu. Rev. Immunol, 4: 369, 1986). The adjuvant used in the present invention that can enhance the immune response can be administered simultaneously with the vaccine composition, or can be administered sequentially at a certain time interval.

[0109] The adjuvant of the present invention is characterized in that it is selected from but not limited to the following group: calcium hydroxide calcium phosphate, mineral oil, squalene, toll-like receptor (TLR) antagonist, detergent, liposome, saponin, cytokine, and a combination thereof.

[0110] In yet another aspect of the present invention, there is provided a method for treating or preventing a disease or disorder in a patient, the method comprising the step of administering to the patient a therapeutically effective amount of the vaccine composition.

[0111] The optimal dose of the vaccine composition of the present invention can be determined by standard studies, which include observing an appropriate immune response in a subject. After the initial vaccination, the subject may be given one or more booster immunizations at appropriate intervals.

[0112] The appropriate dose of the vaccine composition of the present invention depends on factors such as the formulation method, administration method, patient's age, weight, gender, pathological condition, diet, administration time, administration route, excretion rate and reaction sensitivity, and can be appropriately determined by a person skilled in the art taking the above factors into consideration.

[0113] The vaccine composition of the present invention can be administered by the commonly used routes in the medical field. Parenteral administration is preferred and can be administered, for example, by intravenous, intraperitoneal, intramuscular, intraarterial, oral, intracardiac, intramedullary, intradural, transdermal, enteral, subcutaneous, sublingual or topical routes. Generally, the vaccine composition of the present invention is characterized in that it comprises a therapeutically effective amount of a varicella zoster virus surface protein antigen variant according to the present invention as an active ingredient.

[0114] Invention method

[0115] Hereinafter, the present invention will be described in more detail by way of examples. These examples are for illustrative purposes only, and it will be apparent to those skilled in the art that the scope of the present invention is not to be construed as being limited by these examples.

[0116] Example 1: Generation of surface protein (gE) constructs

[0117] In order to produce the surface protein (gE) construct, PCR was used to obtain the required gE fragment. Then, each gE fragment was cut with a restriction enzyme and inserted into the pcDNA3.1 vector. The sequence of the gE fragment inserted into the pcDNA3.1 vector was identified by sequencing. The DNA of the pcDNA3.1 vector containing the gE fragment identified by sequence was obtained by midiprep. The amino acid sequence of the surface protein (gE) fragment is shown in Table 1 below.

[0118] [Table 1]

[0119]

[0120] Example 2: Transient transfection

[0121] To identify the expression level of gE fragment, it was transiently transfected into 293 cells using Lipofectamine 3000. 5 × 10 510 cells were added to each well of a 6-well plate and cultured. Then, the next day, samples for transfection were prepared. 0.2 μg of DNA and 0.4 μg of P3000 were added to a test tube and then diluted with 125 μL of optiMEM; then 3.75 μL of Lipofectamine 3000 was added to another test tube and then diluted with 125 μL of optiMEM. The diluted DNA and an equal amount of diluted Lipofectamine 3000 were transferred to the test tube. Then, the test tubes were mixed and incubated at room temperature for 10 minutes to prepare a DNA-liposome mixture. After the incubation was completed, the DNA-liposome mixture was added to a 6-well plate containing 293 cells, and then cultured in a CO2 incubator for 2 days. After the culture was completed, the supernatant was obtained, 4× sample buffer containing β-mercaptoethanol was added thereto, and then heated at 100°C for 5 minutes. After heating, the resultant was kept frozen until Western blotting was performed.

[0122] Example 3: Immunoblotting

[0123] Western blotting was performed to compare the expression levels of gE fragments. Each sample was electrophoresed on a NuPAGE 4-12% Bis-Tris gel and then transferred to a PVDF membrane. The membrane was blocked in 5% skim milk for 1 hour, incubated with a monoclonal gE antibody (1 μg / mL) for 2 hours, washed with TBST (Tween 0.05%), and then incubated with goat anti-mouse IgG-HRP diluted 5000× for 1 hour. The incubated membrane was washed with TBST and then developed with an ECL substrate. Detection was performed using a Chemidoc machine. As a result of performing Western blotting, as shown in Figure 3 As shown, it was found that the antigens, gE 534 amino acid, gE 537 amino acid, and gE 540 amino acid showed higher expression levels.

[0124] Example 4: Comparison of expression levels with GlaxoSmithKline Biologicals' varicella zoster virus surface protein antigen

[0125] In terms of expression level, the surface protein antigen contained in Shingrix (GSK gE 546 amino acids), which is a currently commercialized varicella zoster vaccine of GlaxoSmithKline Biologicals SA, and the antigen produced by the present inventors (Mogam gE 537 amino acids) were compared. Western blotting was performed in the same manner as described in Example 3. The difference between the surface protein antigen contained in Shingrix of GlaxoSmithKline Biologicals SA (GSK gE 546 amino acids) and the antigen produced by the present inventors (Mogam gE 537 amino acids) is shown in Table 2 below. As a result of performing Western blotting, as shown in Figure 4 As shown, the antigen produced by the present inventors was found to have a higher expression level than the surface protein antigen of GlaxoSmithKline Biologicals.

[0126] [Table 2]

[0127] Mogam gE 537 amino acids GSK gE 546 amino acids source Clade 1 (wild type, Dumas strain) Clade 3 (wild type) Amino acid 40 T I Amino acid 536 L I C-end w / o YAAWTGGLA YAAWTGGLA

[0128] Example 5: Comparison of immunogenicity with GlaxoSmithKline Biologicals' Varicella-Zoster Virus Surface Protein Antigen

[0129] In terms of immunogenicity, the surface protein antigen (GSK gE 546 amino acids) contained in Shingrix, a currently commercialized GlaxoSmithKline Biologicals varicella zoster vaccine, and the antigen (Mogam gE 537 amino acids) produced by the inventors were compared and animal experiments were conducted. From the perspective of humans having a history of varicella infection, in order to mimic varicella infection in mice, female C57BL / 6 mice were given a primary immunization (LAV primary immunization) by subcutaneous injection of a live attenuated vaccine (LAV.3,000pfu) once. 28 days after the LAV primary immunization (day 0), mice were given a secondary immunization by intramuscular injection of a Mogam gE or GSK gE antigen composition containing or not containing an adjuvant. Blood samples were collected 42 days after the LAV primary immunization (day 42) to measure the humoral immune response to gE; leukocytes were collected from spleen samples 42 days after the LAV primary immunization (day 42) to measure the cell-mediated immune response (CMI) to gE or VZV. The immunization day, blood collection day and spleen sample day were calculated from the LAV initial immunization day (i.e., day 0). The overall animal experimental method is described in Table 3 below.

[0130] [Table 3]

[0131]

[0132]

[0133] Example 5-1: Comparison of humoral immune responses

[0134] After primary and secondary immunization, enzyme-linked immunosorbent assay (ELISA) was performed to measure gE antigen-specific IgG potency. Recombinant gE protein (1 μg / mL) was distributed on the ELISA plate and incubated overnight at 4°C to coat the protein antigen thereon. Each antigen-coated ELISA plate was washed 3 times and then blocked for 1 hour with a phosphate-buffered saline (PBS) solution containing 2% bovine serum albumin (BSA). After completing the blocking reaction with BSA, the ELISA plate was washed. Then, diluted serum samples were added thereto and incubated for 2 hours. Horseradish peroxidase (HRP)-conjugated goat anti-mouse IgG, IgG1 or IgG2c antibodies were added thereto and incubated for 1 hour. After the final incubation, the ELISA plate was washed and the HRP reaction was induced by adding 3,3', 5,5'-tetramethylbenzidine (TMB, manufactured by KPL) as an HRP substrate. Then, TMB stop solution was added to stop the HRP reaction, and the optical density (OD) was measured at 450 nm using an ELISA microplate reader (Spectramax 250, microplate reader) to check the amount of antibody produced. Figure 5 As shown, the luminescence intensity of G6 containing the antigen (Mogam gE 537 amino acids) produced by the present inventors and an adjuvant was the highest. From these results, it was found that the highest humoral immune response was induced in G6.

[0135] Example 5-2: Comparison of cell-mediated immune responses

[0136] After primary and secondary immunization, IFN-γ ELISA was performed to check the amount of secreted IFN-γ, which is a representative cytokine secreted by T cells after antigen stimulation. Leukocytes collected from mice were stimulated with VZV lysate for 3 days. Then, centrifugation was performed to obtain supernatant, and the supernatant was analyzed with a mouse IFN-γ ELISA kit. IFN-γ capture antibody (4 μg / mL) was distributed on the ELISA plate, and overnight incubation was performed at room temperature to allow IFN-γ capture antibody to be coated thereon. Each antibody-coated ELISA plate was washed 3 times, and then blocked with PBS containing 1% bovine serum albumin (BSA) for 1 hour. After completing the blocking reaction with BSA, the ELISA plate was washed. Then, the supernatant obtained after stimulating leukocytes was added thereto, and incubated at room temperature for 2 hours. After the culture was completed, the ELISA plate was washed and cultured at room temperature with biotinylated mouse IFN-γ detection antibody (400 ng / mL) for 2 hours. Washed, then incubated with streptavidin-HRP for another 20 minutes. The final incubated ELISA plate was washed and then reacted with the substrate solution at room temperature for 20 minutes. Stop solution was added thereto to terminate the reaction, and then the optical density (OD) was measured at 450 nm using an ELISA microplate reader (Spectramax 250, microplate reader) to examine the amount of IFN-γ cytokine produced. As a result, Figure 6 As shown, G6 containing the antigen produced by the present inventors (Mogam gE 537 amino acids) and adjuvant showed the largest amount of IFN-γ cytokine. From these results, it was found that the highest cell-mediated immune response was induced in G6.

[0137] Example 6: Identification of antigen-specific immunogenicity of gE antigen fragments

[0138] Example 6-1: Transient transfection for antigen production

[0139] To express the gE fragment, use Expifectamine TM 293 transfection kit was used to transiently transfect the cells into 293 cells. 6 ×10 cells / mL and cultured in a 125 mL flask. Then, transfection was performed the next day. 293 cells were cultured at a concentration of 2.9 × 10 6 cells / mL was diluted to a total of 25.5 mL and complexes were prepared for transfection. 30 g of DNA was placed in a 15 mL test tube and adjusted to 1.5 mL using Opti-MEM. In this way, complex 1 was prepared. 81 μL Express Fectamine TMThe 293 reagent was placed in another 15 mL test tube and adjusted to 1.5 mL using Opti-MEM. In this way, complex 2 was prepared. Incubate at room temperature for 5 minutes. After 5 minutes, complex 1 was transferred to the tube containing complex 2 and mixed. Then, the test tube was incubated at room temperature for 20 minutes to prepare the DNA-lipid complex. After the incubation was completed, all the DNA-lipid complexes were placed in a 125 mL flask containing 293 cells and cultured in an incubator. After 20 hours, the enhancer was treated. 150 μL of ExpiFectamine TM 293 Transfection Enhancer 1 was placed in a 1.5 mL test tube and ExpiFectamine was added to it. TM 293 transfection enhancer 2 to 1.5 mL. Then, the resultant was added to 293 cells and cultured in an incubator for 5 days. After 5 days, the culture supernatant was obtained, filtered through a 0.45 μm filter, and frozen until purification.

[0140] Example 6-2: Purification to obtain antigen

[0141] The culture solution that had been frozen was thawed and an equal amount of PBS was added to the culture solution. It was filtered using a 0.22 μm filter and then subjected to anion exchange chromatography. 5 M NaCl was added to the eluate and then subjected to hydrophobic interaction chromatography. The eluate subjected to chromatographic separation was filtered through a 0.22 μm filter and frozen until animal experiments were performed.

[0142] Example 6-3: Immunization

[0143] Animal experiments were performed to identify the immunogenicity of the gE antigen fragment. Female C57BL / 6 mice were injected intramuscularly with the gE antigen fragment at 2-week intervals, and blood samples were collected from the mice 2 weeks after the secondary immunization. Serum was separated from the collected blood samples and then frozen and stored until the antibody titer was determined.

[0144] Example 6-4: Measurement of antigen-specific IgG potency and responders

[0145] Enzyme-linked immunosorbent assay (ELISA) was performed to measure antigen-specific IgG potency. VZV surface protein (1 μg / mL) was coated on ELISA plates, incubated overnight at 4°C, and each ELISA plate was washed 3 times. Then, the ELISA plate was blocked with phosphate-buffered saline (PBS) solution containing 2% bovine serum albumin (BSA) for 1 hour. The ELISA plate was washed. Then, the diluted serum sample was added thereto and incubated for 2 hours. Horseradish peroxidase (HRP)-coupled goat anti-mouse IgG antibody was added thereto and incubated for 1 hour. After the final incubation, the ELISA plate was washed and the HRP reaction was induced by adding 3,3',5,5'-tetramethylbenzidine (TMB) as a substrate. Then, the ELISA stop solution was added thereto to stop the HRP reaction, and the optical density (OD) was measured at 450nm using a spectrometer.

[0146] In order to identify whether the sequence differences between the gE antigen fragments cause differences in antigen-specific immunogenicity, animal immunization was performed by the method described in Example 6-3. The serum obtained in the animal experiment according to the above-mentioned antibody titer measurement method was used to measure the antigen-specific antibody titer. The results are shown in FIG. Figure 7 As shown, the OD value after immunization with gE 534 amino acids or gE 543 amino acids was higher than the OD value after immunization with gE 500 amino acids, gE 510 amino acids, gE 525 amino acids or gE 546 amino acids.

[0147] After measuring antibody titer, the individual with OD value of 0.6 or higher was used as responder, and the result was summarized. As a result, there was no antigen-specific response for gE 500 amino acid, and the number of responders after gE 510 amino acid immunization was the same as the number of responders after gE 546 amino acid immunization. The number of responders after gE 525 amino acid, gE 534 amino acid, gE 537 amino acid or gE 540 amino acid immunization was higher than the number of responders after gE 510 amino acid or gE 546 amino acid immunization. That is, it has been determined that gE 525 amino acid to gE 540 amino acid show a higher number of responders ( Figure 8 ).

[0148] As a result, in the antibody titer assay, gE 534 amino acid to gE 543 amino acid showed higher values, while in antigen-specific responders, gE 525 amino acid to gE 540 amino acid showed higher values. Therefore, when the above two results are put together, it can be determined that gE 534 amino acid to 540 amino acid has higher immunogenicity.

[0149] Industrial Applicability

[0150] The varicella zoster virus surface protein (gE) antigen variant of the present invention is a protein antigen. When used as a vaccine composition, the antigen variant shows excellent safety and high expression level in host cells compared to live virus vaccines. Therefore, this antigen variant can be used as a vaccine for preventing or treating chickenpox or herpes zoster induced by varicella zoster virus.

[0151] As mentioned above, specific parts of the present invention have been described in detail. However, it will be apparent to those skilled in the art that such specific description is only used to illustrate preferred embodiments, and the scope of the present invention is not limited thereto. Therefore, the essential scope of the present invention will be limited by the appended claims and their equivalents.

[0152] Sequence Listing Free Text

[0153] Attached electronic file.

Claims

1. A varicella-zoster virus surface protein antigen variant, comprising a variant, which is a truncation of the carboxyl terminus of any one amino acid residue selected from amino acid residues 534 to 536 and 538 to 540 in a varicella-zoster virus surface protein (gE) antigen, wherein the amino acid sequence of the varicella-zoster virus surface protein (gE) antigen is as shown in SEQ ID NO: 1; wherein the 40th amino acid residue in the amino acid sequence shown in SEQ ID NO: 1 is threonine, and The 536th amino acid residue in the amino acid sequence shown in SEQ ID NO: 1 is leucine.

2. The varicella-zoster virus surface protein antigen variant according to claim 1, wherein the antigen variant is represented by any one of the amino acid sequences shown in SEQ ID NOs: 2 to 4 and 6 to 8.

3. A gene encoding the varicella-zoster virus surface protein antigen variant according to claim 1 or 2.

4. The gene according to claim 3, wherein The gene is represented by any one of the nucleotide sequences shown in SEQ ID NOs: 9 to 11 and 13 to 15.

5. A recombinant vector comprising: the gene according to claim 3. A host cell transformed with the recombinant vector according to claim 5.

7. A vaccine composition for preventing or treating varicella or herpes zoster, comprising an active ingredient: a varicella zoster virus surface protein antigen variant, which comprises a truncation of the carboxyl terminus of any one of the amino acid residues selected from the amino acid residues at positions 534 to 540 in the varicella zoster virus surface protein (gE) antigen, the amino acid sequence of the varicella zoster virus surface protein (gE) antigen being as shown in SEQ ID NO: 1; wherein the 40th amino acid residue in the amino acid sequence shown in SEQ ID NO: 1 is threonine, and The 536th amino acid residue in the amino acid sequence shown in SEQ ID NO: 1 is leucine.

8. Use of a vaccine composition, comprising a varicella-zoster virus surface protein antigen variant as an active ingredient for preventing or treating varicella or herpes zoster, wherein the varicella-zoster virus surface protein antigen variant comprises a truncation of the carboxyl terminal of any one of the amino acid residues selected from the amino acid residues at positions 534 to 540 in the varicella-zoster virus surface protein (gE) antigen, and the amino acid sequence of the varicella-zoster virus surface protein (gE) antigen is as shown in SEQ ID NO: 1; wherein the 40th amino acid residue in the amino acid sequence shown in SEQ ID NO: 1 is threonine, and The 536th amino acid residue in the amino acid sequence shown in SEQ ID NO: 1 is leucine.

9. Use of a vaccine composition in the preparation of a medicament for preventing or treating varicella or herpes zoster, the vaccine composition comprising a varicella zoster virus surface protein antigen variant as an active ingredient; the varicella zoster virus surface protein antigen variant comprises a truncation of the carboxyl terminal of any one of the amino acid residues selected from the amino acid residues at positions 534 to 540 in the varicella zoster virus surface protein (gE) antigen, the amino acid sequence of the varicella zoster virus surface protein (gE) antigen being as shown in SEQ ID NO: 1; wherein the 40th amino acid residue in the amino acid sequence shown in SEQ ID NO: 1 is threonine, and The 536th amino acid residue in the amino acid sequence shown in SEQ ID NO: 1 is leucine.

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