CN115491364A - 亚胺还原酶突变体、亚胺还原酶和葡萄糖脱氢酶共表达酶及其应用 - Google Patents
亚胺还原酶突变体、亚胺还原酶和葡萄糖脱氢酶共表达酶及其应用 Download PDFInfo
- Publication number
- CN115491364A CN115491364A CN202110684461.5A CN202110684461A CN115491364A CN 115491364 A CN115491364 A CN 115491364A CN 202110684461 A CN202110684461 A CN 202110684461A CN 115491364 A CN115491364 A CN 115491364A
- Authority
- CN
- China
- Prior art keywords
- ala
- leu
- gly
- ser
- val
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 108090000854 Oxidoreductases Proteins 0.000 title claims abstract description 111
- 150000002466 imines Chemical class 0.000 title claims abstract description 109
- 102000004190 Enzymes Human genes 0.000 title claims abstract description 84
- 108090000790 Enzymes Proteins 0.000 title claims abstract description 84
- 108010050375 Glucose 1-Dehydrogenase Proteins 0.000 title claims abstract description 66
- SNICXCGAKADSCV-JTQLQIEISA-N (-)-Nicotine Chemical compound CN1CCC[C@H]1C1=CC=CN=C1 SNICXCGAKADSCV-JTQLQIEISA-N 0.000 claims abstract description 61
- 229960002715 nicotine Drugs 0.000 claims abstract description 61
- 229930182840 (S)-nicotine Natural products 0.000 claims abstract description 57
- 239000000758 substrate Substances 0.000 claims abstract description 54
- 102000004316 Oxidoreductases Human genes 0.000 claims description 98
- 239000000243 solution Substances 0.000 claims description 25
- MYKUKUCHPMASKF-UHFFFAOYSA-N Nornicotine Natural products C1CCNC1C1=CC=CN=C1 MYKUKUCHPMASKF-UHFFFAOYSA-N 0.000 claims description 17
- 239000000843 powder Substances 0.000 claims description 17
- 238000000034 method Methods 0.000 claims description 15
- 239000012634 fragment Substances 0.000 claims description 13
- 239000013598 vector Substances 0.000 claims description 12
- 241000894006 Bacteria Species 0.000 claims description 10
- 238000002156 mixing Methods 0.000 claims description 10
- PZDSRPCFNWOUFP-UHFFFAOYSA-N Compound III(S) Chemical compound CCC1COC(C=2C=CC(OCCCCCCCC=3ON=C(C)C=3)=CC=2)=N1 PZDSRPCFNWOUFP-UHFFFAOYSA-N 0.000 claims description 9
- 239000005515 coenzyme Substances 0.000 claims description 9
- 238000012258 culturing Methods 0.000 claims description 8
- -1 dihydrochloride Chemical compound 0.000 claims description 8
- 238000001976 enzyme digestion Methods 0.000 claims description 8
- 108090000623 proteins and genes Proteins 0.000 claims description 8
- 150000003839 salts Chemical class 0.000 claims description 8
- 238000004519 manufacturing process Methods 0.000 claims description 7
- 150000007523 nucleic acids Chemical class 0.000 claims description 7
- 108091008146 restriction endonucleases Proteins 0.000 claims description 7
- 108020004707 nucleic acids Proteins 0.000 claims description 6
- 102000039446 nucleic acids Human genes 0.000 claims description 6
- 230000004186 co-expression Effects 0.000 claims description 5
- 230000001939 inductive effect Effects 0.000 claims description 5
- 102000012410 DNA Ligases Human genes 0.000 claims description 4
- 108010061982 DNA Ligases Proteins 0.000 claims description 4
- QAOWNCQODCNURD-UHFFFAOYSA-L Sulfate Chemical compound [O-]S([O-])(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-L 0.000 claims description 4
- 239000007853 buffer solution Substances 0.000 claims description 4
- 238000004108 freeze drying Methods 0.000 claims description 4
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 claims description 3
- CPELXLSAUQHCOX-UHFFFAOYSA-N Hydrogen bromide Chemical compound Br CPELXLSAUQHCOX-UHFFFAOYSA-N 0.000 claims description 3
- 230000003197 catalytic effect Effects 0.000 claims description 3
- 238000011033 desalting Methods 0.000 claims description 3
- QAOWNCQODCNURD-UHFFFAOYSA-M hydrogensulfate Chemical compound OS([O-])(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-M 0.000 claims description 3
- 230000001131 transforming effect Effects 0.000 claims description 3
- 241000672609 Escherichia coli BL21 Species 0.000 claims description 2
- 239000013604 expression vector Substances 0.000 claims description 2
- 239000003292 glue Substances 0.000 claims description 2
- 125000003275 alpha amino acid group Chemical group 0.000 claims 2
- 230000001035 methylating effect Effects 0.000 claims 1
- 238000002360 preparation method Methods 0.000 abstract description 22
- 230000000694 effects Effects 0.000 abstract description 18
- YSYDPEFJZDXDGK-UHFFFAOYSA-N 4-(methylamino)-1-pyridin-3-ylbutan-2-one Chemical compound CNCCC(CC=1C=NC=CC=1)=O YSYDPEFJZDXDGK-UHFFFAOYSA-N 0.000 abstract description 10
- 238000006555 catalytic reaction Methods 0.000 abstract description 5
- SNICXCGAKADSCV-UHFFFAOYSA-N nicotine Natural products CN1CCCC1C1=CC=CN=C1 SNICXCGAKADSCV-UHFFFAOYSA-N 0.000 abstract description 5
- VPZXBVLAVMBEQI-UHFFFAOYSA-N glycyl-DL-alpha-alanine Natural products OC(=O)C(C)NC(=O)CN VPZXBVLAVMBEQI-UHFFFAOYSA-N 0.000 description 189
- BTBUEVAGZCKULD-XPUUQOCRSA-N Ala-Gly-His Chemical compound C[C@H](N)C(=O)NCC(=O)N[C@H](C(O)=O)CC1=CN=CN1 BTBUEVAGZCKULD-XPUUQOCRSA-N 0.000 description 108
- LKDIBBOKUAASNP-FXQIFTODSA-N Glu-Ala-Glu Chemical compound OC(=O)CC[C@H](N)C(=O)N[C@@H](C)C(=O)N[C@@H](CCC(O)=O)C(O)=O LKDIBBOKUAASNP-FXQIFTODSA-N 0.000 description 108
- 238000006243 chemical reaction Methods 0.000 description 84
- PYTZFYUXZZHOAD-WHFBIAKZSA-N Gly-Ala-Ala Chemical compound OC(=O)[C@H](C)NC(=O)[C@H](C)NC(=O)CN PYTZFYUXZZHOAD-WHFBIAKZSA-N 0.000 description 72
- 229940088598 enzyme Drugs 0.000 description 69
- ZUGXSSFMTXKHJS-ZLUOBGJFSA-N Ser-Ala-Ala Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](C)C(=O)N[C@@H](C)C(O)=O ZUGXSSFMTXKHJS-ZLUOBGJFSA-N 0.000 description 64
- YCRAFFCYWOUEOF-DLOVCJGASA-N Ala-Phe-Ser Chemical compound OC[C@@H](C(O)=O)NC(=O)[C@@H](NC(=O)[C@@H](N)C)CC1=CC=CC=C1 YCRAFFCYWOUEOF-DLOVCJGASA-N 0.000 description 61
- 108020004414 DNA Proteins 0.000 description 61
- OXRLYTYUXAQTHP-YUMQZZPRSA-N Leu-Gly-Ala Chemical compound [H]N[C@@H](CC(C)C)C(=O)NCC(=O)N[C@@H](C)C(O)=O OXRLYTYUXAQTHP-YUMQZZPRSA-N 0.000 description 60
- DKJPOZOEBONHFS-ZLUOBGJFSA-N Ala-Ala-Asp Chemical compound C[C@H](N)C(=O)N[C@@H](C)C(=O)N[C@H](C(O)=O)CC(O)=O DKJPOZOEBONHFS-ZLUOBGJFSA-N 0.000 description 54
- DVWVZSJAYIJZFI-FXQIFTODSA-N Ala-Arg-Asn Chemical compound [H]N[C@@H](C)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC(N)=O)C(O)=O DVWVZSJAYIJZFI-FXQIFTODSA-N 0.000 description 54
- IFTVANMRTIHKML-WDSKDSINSA-N Ala-Gln-Gly Chemical compound C[C@H](N)C(=O)N[C@@H](CCC(N)=O)C(=O)NCC(O)=O IFTVANMRTIHKML-WDSKDSINSA-N 0.000 description 54
- FVSOUJZKYWEFOB-KBIXCLLPSA-N Ala-Gln-Ile Chemical compound CC[C@H](C)[C@@H](C(O)=O)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@H](C)N FVSOUJZKYWEFOB-KBIXCLLPSA-N 0.000 description 54
- LMFXXZPPZDCPTA-ZKWXMUAHSA-N Ala-Gly-Ile Chemical compound CC[C@H](C)[C@@H](C(O)=O)NC(=O)CNC(=O)[C@H](C)N LMFXXZPPZDCPTA-ZKWXMUAHSA-N 0.000 description 54
- KMGOBAQSCKTBGD-DLOVCJGASA-N Ala-His-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@@H](NC(=O)[C@H](C)N)CC1=CN=CN1 KMGOBAQSCKTBGD-DLOVCJGASA-N 0.000 description 54
- MDNAVFBZPROEHO-UHFFFAOYSA-N Ala-Lys-Val Natural products CC(C)C(C(O)=O)NC(=O)C(NC(=O)C(C)N)CCCCN MDNAVFBZPROEHO-UHFFFAOYSA-N 0.000 description 54
- HOVPGJUNRLMIOZ-CIUDSAMLSA-N Ala-Ser-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@H](CO)NC(=O)[C@H](C)N HOVPGJUNRLMIOZ-CIUDSAMLSA-N 0.000 description 54
- YNOCMHZSWJMGBB-GCJQMDKQSA-N Ala-Thr-Asp Chemical compound [H]N[C@@H](C)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC(O)=O)C(O)=O YNOCMHZSWJMGBB-GCJQMDKQSA-N 0.000 description 54
- KUFVXLQLDHJVOG-SHGPDSBTSA-N Ala-Thr-Thr Chemical compound C[C@H]([C@@H](C(=O)N[C@@H]([C@@H](C)O)C(=O)O)NC(=O)[C@H](C)N)O KUFVXLQLDHJVOG-SHGPDSBTSA-N 0.000 description 54
- WOPJVEMFXYHZEE-SRVKXCTJSA-N Asp-Phe-Asp Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CC(O)=O)C(O)=O WOPJVEMFXYHZEE-SRVKXCTJSA-N 0.000 description 54
- ZBYLEBZCVKLPCY-FXQIFTODSA-N Asp-Ser-Arg Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](CO)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O ZBYLEBZCVKLPCY-FXQIFTODSA-N 0.000 description 54
- HILMIYALTUQTRC-XVKPBYJWSA-N Glu-Gly-Val Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)NCC(=O)N[C@@H](C(C)C)C(O)=O HILMIYALTUQTRC-XVKPBYJWSA-N 0.000 description 54
- LJPIRKICOISLKN-WHFBIAKZSA-N Gly-Ala-Ser Chemical compound NCC(=O)N[C@@H](C)C(=O)N[C@@H](CO)C(O)=O LJPIRKICOISLKN-WHFBIAKZSA-N 0.000 description 54
- TZOVVRJYUDETQG-RCOVLWMOSA-N Gly-Asp-Val Chemical compound CC(C)[C@@H](C(O)=O)NC(=O)[C@H](CC(O)=O)NC(=O)CN TZOVVRJYUDETQG-RCOVLWMOSA-N 0.000 description 54
- XLFHCWHXKSFVIB-BQBZGAKWSA-N Gly-Gln-Gln Chemical compound NCC(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCC(N)=O)C(O)=O XLFHCWHXKSFVIB-BQBZGAKWSA-N 0.000 description 54
- XPJBQTCXPJNIFE-ZETCQYMHSA-N Gly-Gly-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)CNC(=O)CN XPJBQTCXPJNIFE-ZETCQYMHSA-N 0.000 description 54
- UUYBFNKHOCJCHT-VHSXEESVSA-N Gly-Leu-Pro Chemical compound CC(C)C[C@@H](C(=O)N1CCC[C@@H]1C(=O)O)NC(=O)CN UUYBFNKHOCJCHT-VHSXEESVSA-N 0.000 description 54
- FXGRXIATVXUAHO-WEDXCCLWSA-N Gly-Lys-Thr Chemical compound C[C@@H](O)[C@@H](C(O)=O)NC(=O)[C@@H](NC(=O)CN)CCCCN FXGRXIATVXUAHO-WEDXCCLWSA-N 0.000 description 54
- FFALDIDGPLUDKV-ZDLURKLDSA-N Gly-Thr-Ser Chemical compound [H]NCC(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CO)C(O)=O FFALDIDGPLUDKV-ZDLURKLDSA-N 0.000 description 54
- JBCLFWXMTIKCCB-UHFFFAOYSA-N H-Gly-Phe-OH Natural products NCC(=O)NC(C(O)=O)CC1=CC=CC=C1 JBCLFWXMTIKCCB-UHFFFAOYSA-N 0.000 description 54
- NKVZTQVGUNLLQW-JBDRJPRFSA-N Ile-Ala-Ala Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](C)C(=O)N[C@@H](C)C(=O)O)N NKVZTQVGUNLLQW-JBDRJPRFSA-N 0.000 description 54
- DURWCDDDAWVPOP-JBDRJPRFSA-N Ile-Cys-Ser Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CS)C(=O)N[C@@H](CO)C(=O)O)N DURWCDDDAWVPOP-JBDRJPRFSA-N 0.000 description 54
- BQSLGJHIAGOZCD-CIUDSAMLSA-N Leu-Ala-Ser Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](C)C(=O)N[C@@H](CO)C(O)=O BQSLGJHIAGOZCD-CIUDSAMLSA-N 0.000 description 54
- ULXYQAJWJGLCNR-YUMQZZPRSA-N Leu-Asp-Gly Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](CC(O)=O)C(=O)NCC(O)=O ULXYQAJWJGLCNR-YUMQZZPRSA-N 0.000 description 54
- CFZZDVMBRYFFNU-QWRGUYRKSA-N Leu-His-Gly Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](CC1=CNC=N1)C(=O)NCC(O)=O CFZZDVMBRYFFNU-QWRGUYRKSA-N 0.000 description 54
- ORWTWZXGDBYVCP-BJDJZHNGSA-N Leu-Ile-Cys Chemical compound SC[C@@H](C(O)=O)NC(=O)[C@H]([C@@H](C)CC)NC(=O)[C@@H](N)CC(C)C ORWTWZXGDBYVCP-BJDJZHNGSA-N 0.000 description 54
- HNDWYLYAYNBWMP-AJNGGQMLSA-N Leu-Ile-Lys Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CCCCN)C(=O)O)NC(=O)[C@H](CC(C)C)N HNDWYLYAYNBWMP-AJNGGQMLSA-N 0.000 description 54
- LXKNSJLSGPNHSK-KKUMJFAQSA-N Leu-Leu-Lys Chemical compound CC(C)C[C@@H](C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCCCN)C(=O)O)N LXKNSJLSGPNHSK-KKUMJFAQSA-N 0.000 description 54
- AIRUUHAOKGVJAD-JYJNAYRXSA-N Leu-Phe-Glu Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CCC(O)=O)C(O)=O AIRUUHAOKGVJAD-JYJNAYRXSA-N 0.000 description 54
- AIRZWUMAHCDDHR-KKUMJFAQSA-N Lys-Leu-Leu Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC(C)C)C(O)=O AIRZWUMAHCDDHR-KKUMJFAQSA-N 0.000 description 54
- KDIIENQUNVNWHR-JYJNAYRXSA-N Pro-Arg-Phe Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC1=CC=CC=C1)C(O)=O KDIIENQUNVNWHR-JYJNAYRXSA-N 0.000 description 54
- PRKWBYCXBBSLSK-GUBZILKMSA-N Pro-Ser-Val Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](CO)C(=O)N[C@@H](C(C)C)C(O)=O PRKWBYCXBBSLSK-GUBZILKMSA-N 0.000 description 54
- YDTUEBLEAVANFH-RCWTZXSCSA-N Pro-Val-Thr Chemical compound C[C@@H](O)[C@@H](C(O)=O)NC(=O)[C@H](C(C)C)NC(=O)[C@@H]1CCCN1 YDTUEBLEAVANFH-RCWTZXSCSA-N 0.000 description 54
- WDXYVIIVDIDOSX-DCAQKATOSA-N Ser-Arg-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@@H](NC(=O)[C@@H](N)CO)CCCN=C(N)N WDXYVIIVDIDOSX-DCAQKATOSA-N 0.000 description 54
- QPFJSHSJFIYDJZ-GHCJXIJMSA-N Ser-Asp-Ile Chemical compound CC[C@H](C)[C@@H](C(O)=O)NC(=O)[C@H](CC(O)=O)NC(=O)[C@@H](N)CO QPFJSHSJFIYDJZ-GHCJXIJMSA-N 0.000 description 54
- WNDUPCKKKGSKIQ-CIUDSAMLSA-N Ser-Pro-Gln Chemical compound OC[C@H](N)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CCC(N)=O)C(O)=O WNDUPCKKKGSKIQ-CIUDSAMLSA-N 0.000 description 54
- PURRNJBBXDDWLX-ZDLURKLDSA-N Ser-Thr-Gly Chemical compound C[C@H]([C@@H](C(=O)NCC(=O)O)NC(=O)[C@H](CO)N)O PURRNJBBXDDWLX-ZDLURKLDSA-N 0.000 description 54
- KZSYAEWQMJEGRZ-RHYQMDGZSA-N Thr-Leu-Val Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](C(C)C)C(O)=O KZSYAEWQMJEGRZ-RHYQMDGZSA-N 0.000 description 54
- ASQFIHTXXMFENG-XPUUQOCRSA-N Val-Ala-Gly Chemical compound CC(C)[C@H](N)C(=O)N[C@@H](C)C(=O)NCC(O)=O ASQFIHTXXMFENG-XPUUQOCRSA-N 0.000 description 54
- RUCNAYOMFXRIKJ-DCAQKATOSA-N Val-Ala-Lys Chemical compound CC(C)[C@H](N)C(=O)N[C@@H](C)C(=O)N[C@H](C(O)=O)CCCCN RUCNAYOMFXRIKJ-DCAQKATOSA-N 0.000 description 54
- CWSIBTLMMQLPPZ-FXQIFTODSA-N Val-Cys-Ala Chemical compound C[C@@H](C(=O)O)NC(=O)[C@H](CS)NC(=O)[C@H](C(C)C)N CWSIBTLMMQLPPZ-FXQIFTODSA-N 0.000 description 54
- VFOHXOLPLACADK-GVXVVHGQSA-N Val-Gln-Leu Chemical compound CC(C)C[C@@H](C(=O)O)NC(=O)[C@H](CCC(=O)N)NC(=O)[C@H](C(C)C)N VFOHXOLPLACADK-GVXVVHGQSA-N 0.000 description 54
- OTJMMKPMLUNTQT-AVGNSLFASA-N Val-Leu-Arg Chemical compound CC(C)C[C@@H](C(=O)N[C@@H](CCCN=C(N)N)C(=O)O)NC(=O)[C@H](C(C)C)N OTJMMKPMLUNTQT-AVGNSLFASA-N 0.000 description 54
- LNWSJGJCLFUNTN-ZOBUZTSGSA-N Val-Trp-Asn Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CC1=CNC2=CC=CC=C21)C(=O)N[C@@H](CC(=O)N)C(=O)O)N LNWSJGJCLFUNTN-ZOBUZTSGSA-N 0.000 description 54
- 108010086434 alanyl-seryl-glycine Proteins 0.000 description 54
- KOSRFJWDECSPRO-UHFFFAOYSA-N alpha-L-glutamyl-L-glutamic acid Natural products OC(=O)CCC(N)C(=O)NC(CCC(O)=O)C(O)=O KOSRFJWDECSPRO-UHFFFAOYSA-N 0.000 description 54
- 108010026364 glycyl-glycyl-leucine Proteins 0.000 description 54
- 108010050848 glycylleucine Proteins 0.000 description 54
- 108010081551 glycylphenylalanine Proteins 0.000 description 54
- 108010040030 histidinoalanine Proteins 0.000 description 54
- 108010085325 histidylproline Proteins 0.000 description 54
- 108010044311 leucyl-glycyl-glycine Proteins 0.000 description 54
- 108010017391 lysylvaline Proteins 0.000 description 54
- 108010053725 prolylvaline Proteins 0.000 description 54
- QXUPRMQJDWJDFR-NRPADANISA-N Glu-Val-Ser Chemical compound CC(C)[C@H](NC(=O)[C@@H](N)CCC(O)=O)C(=O)N[C@@H](CO)C(O)=O QXUPRMQJDWJDFR-NRPADANISA-N 0.000 description 51
- 108010026333 seryl-proline Proteins 0.000 description 51
- FBEJIDRSQCGFJI-GUBZILKMSA-N Glu-Leu-Ser Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CO)C(O)=O FBEJIDRSQCGFJI-GUBZILKMSA-N 0.000 description 50
- MNZHHDPWDWQJCQ-YUMQZZPRSA-N Ala-Leu-Gly Chemical compound C[C@H](N)C(=O)N[C@@H](CC(C)C)C(=O)NCC(O)=O MNZHHDPWDWQJCQ-YUMQZZPRSA-N 0.000 description 48
- IOFVWPYSRSCWHI-JXUBOQSCSA-N Ala-Thr-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@H]([C@@H](C)O)NC(=O)[C@H](C)N IOFVWPYSRSCWHI-JXUBOQSCSA-N 0.000 description 48
- FZIJIFCXUCZHOL-CIUDSAMLSA-N Lys-Ala-Ala Chemical compound OC(=O)[C@H](C)NC(=O)[C@H](C)NC(=O)[C@@H](N)CCCCN FZIJIFCXUCZHOL-CIUDSAMLSA-N 0.000 description 48
- GURGCNUWVSDYTP-SRVKXCTJSA-N Pro-Leu-Gln Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(N)=O)C(O)=O GURGCNUWVSDYTP-SRVKXCTJSA-N 0.000 description 48
- 108010024654 phenylalanyl-prolyl-alanine Proteins 0.000 description 47
- FZWVCYCYWCLQDH-NHCYSSNCSA-N Ile-Leu-Gly Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CC(C)C)C(=O)NCC(=O)O)N FZWVCYCYWCLQDH-NHCYSSNCSA-N 0.000 description 45
- 108010034529 leucyl-lysine Proteins 0.000 description 43
- GFMJUESGWILPEN-MELADBBJSA-N Cys-Phe-Pro Chemical compound C1C[C@@H](N(C1)C(=O)[C@H](CC2=CC=CC=C2)NC(=O)[C@H](CS)N)C(=O)O GFMJUESGWILPEN-MELADBBJSA-N 0.000 description 41
- PHONXOACARQMPM-BQBZGAKWSA-N Gly-Ala-Met Chemical compound [H]NCC(=O)N[C@@H](C)C(=O)N[C@@H](CCSC)C(O)=O PHONXOACARQMPM-BQBZGAKWSA-N 0.000 description 41
- NYEYYMLUABXDMC-NHCYSSNCSA-N Ile-Gly-Leu Chemical compound CC[C@H](C)[C@@H](C(=O)NCC(=O)N[C@@H](CC(C)C)C(=O)O)N NYEYYMLUABXDMC-NHCYSSNCSA-N 0.000 description 41
- SVBJIZVVYJYGLA-DCAQKATOSA-N Leu-Ser-Val Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](CO)C(=O)N[C@@H](C(C)C)C(O)=O SVBJIZVVYJYGLA-DCAQKATOSA-N 0.000 description 41
- VIZLHGTVGKBBKO-AVGNSLFASA-N Met-Arg-His Chemical compound CSCC[C@@H](C(=O)N[C@@H](CCCN=C(N)N)C(=O)N[C@@H](CC1=CN=CN1)C(=O)O)N VIZLHGTVGKBBKO-AVGNSLFASA-N 0.000 description 41
- 108010025306 histidylleucine Proteins 0.000 description 41
- 108010005942 methionylglycine Proteins 0.000 description 41
- UESYBOXFJWJVSB-AVGNSLFASA-N Gln-Phe-Ser Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CO)C(O)=O UESYBOXFJWJVSB-AVGNSLFASA-N 0.000 description 40
- 108010044940 alanylglutamine Proteins 0.000 description 40
- TZDNWXDLYFIFPT-BJDJZHNGSA-N Ala-Ile-Leu Chemical compound [H]N[C@@H](C)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CC(C)C)C(O)=O TZDNWXDLYFIFPT-BJDJZHNGSA-N 0.000 description 39
- SEZADXQOJJTXPG-VFAJRCTISA-N Lys-Thr-Trp Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CC1=CNC2=CC=CC=C21)C(=O)O)NC(=O)[C@H](CCCCN)N)O SEZADXQOJJTXPG-VFAJRCTISA-N 0.000 description 37
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 description 36
- 108010028939 alanyl-alanyl-lysyl-alanine Proteins 0.000 description 36
- QCBYAHHNOHBXIH-UWVGGRQHSA-N His-Pro-Gly Chemical compound C([C@H](N)C(=O)N1[C@@H](CCC1)C(=O)NCC(O)=O)C1=CN=CN1 QCBYAHHNOHBXIH-UWVGGRQHSA-N 0.000 description 33
- 108010079364 N-glycylalanine Proteins 0.000 description 33
- 108010087924 alanylproline Proteins 0.000 description 30
- VLKZOEOYAKHREP-UHFFFAOYSA-N n-Hexane Chemical compound CCCCCC VLKZOEOYAKHREP-UHFFFAOYSA-N 0.000 description 28
- DYXOFPBJBAHWFY-JBDRJPRFSA-N Ala-Ser-Ile Chemical compound CC[C@H](C)[C@@H](C(O)=O)NC(=O)[C@H](CO)NC(=O)[C@H](C)N DYXOFPBJBAHWFY-JBDRJPRFSA-N 0.000 description 27
- XJLXINKUBYWONI-DQQFMEOOSA-N [[(2r,3r,4r,5r)-5-(6-aminopurin-9-yl)-3-hydroxy-4-phosphonooxyoxolan-2-yl]methoxy-hydroxyphosphoryl] [(2s,3r,4s,5s)-5-(3-carbamoylpyridin-1-ium-1-yl)-3,4-dihydroxyoxolan-2-yl]methyl phosphate Chemical compound NC(=O)C1=CC=C[N+]([C@@H]2[C@H]([C@@H](O)[C@H](COP([O-])(=O)OP(O)(=O)OC[C@@H]3[C@H]([C@@H](OP(O)(O)=O)[C@@H](O3)N3C4=NC=NC(N)=C4N=C3)O)O2)O)=C1 XJLXINKUBYWONI-DQQFMEOOSA-N 0.000 description 25
- IRJWAYCXIYUHQE-WHFBIAKZSA-N Gly-Ser-Ala Chemical compound OC(=O)[C@H](C)NC(=O)[C@H](CO)NC(=O)CN IRJWAYCXIYUHQE-WHFBIAKZSA-N 0.000 description 24
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 22
- 239000008103 glucose Substances 0.000 description 22
- 108010025153 lysyl-alanyl-alanine Proteins 0.000 description 18
- 238000003756 stirring Methods 0.000 description 18
- CQZDZKRHFWJXDF-WDSKDSINSA-N Gly-Gln-Ala Chemical compound OC(=O)[C@H](C)NC(=O)[C@H](CCC(N)=O)NC(=O)CN CQZDZKRHFWJXDF-WDSKDSINSA-N 0.000 description 17
- 239000012528 membrane Substances 0.000 description 16
- 238000004128 high performance liquid chromatography Methods 0.000 description 15
- 239000008363 phosphate buffer Substances 0.000 description 15
- LNNSWWRRYJLGNI-NAKRPEOUSA-N Ala-Ile-Val Chemical compound C[C@H](N)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](C(C)C)C(O)=O LNNSWWRRYJLGNI-NAKRPEOUSA-N 0.000 description 14
- CHIAUHSHDARFBD-ULQDDVLXSA-N His-Pro-Tyr Chemical compound C([C@H](N)C(=O)N1[C@@H](CCC1)C(=O)N[C@@H](CC=1C=CC(O)=CC=1)C(O)=O)C1=CN=CN1 CHIAUHSHDARFBD-ULQDDVLXSA-N 0.000 description 13
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 description 12
- VNFSAYFQLXPHPY-CIQUZCHMSA-N Ala-Thr-Ile Chemical compound [H]N[C@@H](C)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O VNFSAYFQLXPHPY-CIQUZCHMSA-N 0.000 description 11
- 239000011541 reaction mixture Substances 0.000 description 11
- 239000000047 product Substances 0.000 description 10
- NJIFPLAJSVUQOZ-JBDRJPRFSA-N Ala-Cys-Ile Chemical compound CC[C@H](C)[C@@H](C(=O)O)NC(=O)[C@H](CS)NC(=O)[C@H](C)N NJIFPLAJSVUQOZ-JBDRJPRFSA-N 0.000 description 9
- 239000007788 liquid Substances 0.000 description 9
- MYKUKUCHPMASKF-VIFPVBQESA-N (S)-nornicotine Chemical compound C1CCN[C@@H]1C1=CC=CN=C1 MYKUKUCHPMASKF-VIFPVBQESA-N 0.000 description 8
- HRNQLKCLPVKZNE-CIUDSAMLSA-N Ser-Ala-Leu Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](C)C(=O)N[C@@H](CC(C)C)C(O)=O HRNQLKCLPVKZNE-CIUDSAMLSA-N 0.000 description 8
- 239000012295 chemical reaction liquid Substances 0.000 description 8
- 238000010438 heat treatment Methods 0.000 description 8
- 229930027917 kanamycin Natural products 0.000 description 8
- SBUJHOSQTJFQJX-NOAMYHISSA-N kanamycin Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CN)O[C@@H]1O[C@H]1[C@H](O)[C@@H](O[C@@H]2[C@@H]([C@@H](N)[C@H](O)[C@@H](CO)O2)O)[C@H](N)C[C@@H]1N SBUJHOSQTJFQJX-NOAMYHISSA-N 0.000 description 8
- 229960000318 kanamycin Drugs 0.000 description 8
- 229930182823 kanamycin A Natural products 0.000 description 8
- WUHJHHGYVVJMQE-BJDJZHNGSA-N Ala-Leu-Ile Chemical compound [H]N[C@@H](C)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O WUHJHHGYVVJMQE-BJDJZHNGSA-N 0.000 description 7
- PWAIZUBWHRHYKS-MELADBBJSA-N Asp-Phe-Pro Chemical compound C1C[C@@H](N(C1)C(=O)[C@H](CC2=CC=CC=C2)NC(=O)[C@H](CC(=O)O)N)C(=O)O PWAIZUBWHRHYKS-MELADBBJSA-N 0.000 description 7
- 150000001413 amino acids Chemical group 0.000 description 7
- 230000035772 mutation Effects 0.000 description 7
- 239000013612 plasmid Substances 0.000 description 7
- CXRCVCURMBFFOL-FXQIFTODSA-N Ala-Ala-Pro Chemical compound C[C@H](N)C(=O)N[C@@H](C)C(=O)N1CCC[C@H]1C(O)=O CXRCVCURMBFFOL-FXQIFTODSA-N 0.000 description 6
- VQPPIMUZCZCOIL-GUBZILKMSA-N Leu-Gln-Ala Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](C)C(O)=O VQPPIMUZCZCOIL-GUBZILKMSA-N 0.000 description 6
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 6
- BVOVIGCHYNFJBZ-JXUBOQSCSA-N Thr-Leu-Ala Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](C)C(O)=O BVOVIGCHYNFJBZ-JXUBOQSCSA-N 0.000 description 6
- JMBRNXUOLJFURW-BEAPCOKYSA-N Thr-Phe-Pro Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)N2CCC[C@@H]2C(=O)O)N)O JMBRNXUOLJFURW-BEAPCOKYSA-N 0.000 description 6
- 238000004458 analytical method Methods 0.000 description 6
- 238000000605 extraction Methods 0.000 description 6
- 238000004305 normal phase HPLC Methods 0.000 description 6
- KRXIWXCXOARFNT-ZLUOBGJFSA-N Asp-Ala-Ala Chemical compound OC(=O)[C@H](C)NC(=O)[C@H](C)NC(=O)[C@@H](N)CC(O)=O KRXIWXCXOARFNT-ZLUOBGJFSA-N 0.000 description 5
- PTRKPHUGYULXPU-KKUMJFAQSA-N Leu-Phe-Ser Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CO)C(O)=O PTRKPHUGYULXPU-KKUMJFAQSA-N 0.000 description 5
- RNYLNYTYMXACRI-VFAJRCTISA-N Leu-Thr-Trp Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC1=CNC2=C1C=CC=C2)C(O)=O RNYLNYTYMXACRI-VFAJRCTISA-N 0.000 description 5
- PFZWARWVRNTPBR-IHPCNDPISA-N Lys-Leu-Trp Chemical compound CC(C)C[C@@H](C(=O)N[C@@H](CC1=CNC2=CC=CC=C21)C(=O)O)NC(=O)[C@H](CCCCN)N PFZWARWVRNTPBR-IHPCNDPISA-N 0.000 description 5
- 238000001514 detection method Methods 0.000 description 5
- 238000011068 loading method Methods 0.000 description 5
- 230000035484 reaction time Effects 0.000 description 5
- 238000012216 screening Methods 0.000 description 5
- 238000003786 synthesis reaction Methods 0.000 description 5
- WSFSSNUMVMOOMR-UHFFFAOYSA-N Formaldehyde Chemical compound O=C WSFSSNUMVMOOMR-UHFFFAOYSA-N 0.000 description 4
- VXEFAWJTFAUDJK-AVGNSLFASA-N Glu-Tyr-Ser Chemical compound C1=CC(=CC=C1C[C@@H](C(=O)N[C@@H](CO)C(=O)O)NC(=O)[C@H](CCC(=O)O)N)O VXEFAWJTFAUDJK-AVGNSLFASA-N 0.000 description 4
- PZZJMBYSYAKYPK-UWJYBYFXSA-N Ser-Ala-Tyr Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](C)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(O)=O PZZJMBYSYAKYPK-UWJYBYFXSA-N 0.000 description 4
- RFKVQLIXNVEOMB-WEDXCCLWSA-N Thr-Leu-Gly Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CC(C)C)C(=O)NCC(=O)O)N)O RFKVQLIXNVEOMB-WEDXCCLWSA-N 0.000 description 4
- HEDRZPFGACZZDS-MICDWDOJSA-N Trichloro(2H)methane Chemical compound [2H]C(Cl)(Cl)Cl HEDRZPFGACZZDS-MICDWDOJSA-N 0.000 description 4
- 238000004296 chiral HPLC Methods 0.000 description 4
- 230000002255 enzymatic effect Effects 0.000 description 4
- BPHPUYQFMNQIOC-NXRLNHOXSA-N isopropyl beta-D-thiogalactopyranoside Chemical compound CC(C)S[C@@H]1O[C@H](CO)[C@H](O)[C@H](O)[C@H]1O BPHPUYQFMNQIOC-NXRLNHOXSA-N 0.000 description 4
- 239000002609 medium Substances 0.000 description 4
- BDAGIHXWWSANSR-UHFFFAOYSA-N methanoic acid Natural products OC=O BDAGIHXWWSANSR-UHFFFAOYSA-N 0.000 description 4
- 239000006228 supernatant Substances 0.000 description 4
- 108010036211 5-HT-moduline Proteins 0.000 description 3
- 241000588724 Escherichia coli Species 0.000 description 3
- 241001198387 Escherichia coli BL21(DE3) Species 0.000 description 3
- FGSGPLRPQCZBSQ-AVGNSLFASA-N Glu-Phe-Ser Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CO)C(O)=O FGSGPLRPQCZBSQ-AVGNSLFASA-N 0.000 description 3
- ZVKDCQVQTGYBQT-LSJOCFKGSA-N His-Pro-Ala Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)N1CCC[C@H]1C(=O)N[C@@H](C)C(O)=O ZVKDCQVQTGYBQT-LSJOCFKGSA-N 0.000 description 3
- VCBWXASUBZIFLQ-IHRRRGAJSA-N His-Pro-Leu Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CC(C)C)C(O)=O VCBWXASUBZIFLQ-IHRRRGAJSA-N 0.000 description 3
- 239000012880 LB liquid culture medium Substances 0.000 description 3
- YOKVEHGYYQEQOP-QWRGUYRKSA-N Leu-Leu-Gly Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](CC(C)C)C(=O)NCC(O)=O YOKVEHGYYQEQOP-QWRGUYRKSA-N 0.000 description 3
- 230000001580 bacterial effect Effects 0.000 description 3
- 230000015572 biosynthetic process Effects 0.000 description 3
- 238000005119 centrifugation Methods 0.000 description 3
- 229930027945 nicotinamide-adenine dinucleotide Natural products 0.000 description 3
- 239000002773 nucleotide Substances 0.000 description 3
- 125000003729 nucleotide group Chemical group 0.000 description 3
- 238000001228 spectrum Methods 0.000 description 3
- SNICXCGAKADSCV-SNVBAGLBSA-N (+)-nicotine Chemical compound CN1CCC[C@@H]1C1=CC=CN=C1 SNICXCGAKADSCV-SNVBAGLBSA-N 0.000 description 2
- 229930182841 (R)-nicotine Natural products 0.000 description 2
- 238000005160 1H NMR spectroscopy Methods 0.000 description 2
- CTSZPNIMMLSKDV-UHFFFAOYSA-O 2-methyl-1-pyrrolinium Chemical compound CC1=[NH+]CCC1 CTSZPNIMMLSKDV-UHFFFAOYSA-O 0.000 description 2
- OSWFIVFLDKOXQC-UHFFFAOYSA-N 4-(3-methoxyphenyl)aniline Chemical compound COC1=CC=CC(C=2C=CC(N)=CC=2)=C1 OSWFIVFLDKOXQC-UHFFFAOYSA-N 0.000 description 2
- 241000607574 Aeromonas veronii Species 0.000 description 2
- YJIUYQKQBBQYHZ-ACZMJKKPSA-N Gln-Ala-Ala Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)N[C@@H](C)C(=O)N[C@@H](C)C(O)=O YJIUYQKQBBQYHZ-ACZMJKKPSA-N 0.000 description 2
- OWYIDJCNRWRSJY-QTKMDUPCSA-N His-Pro-Thr Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)N1CCC[C@H]1C(=O)N[C@@H]([C@@H](C)O)C(O)=O OWYIDJCNRWRSJY-QTKMDUPCSA-N 0.000 description 2
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 description 2
- BZLVMXJERCGZMT-UHFFFAOYSA-N Methyl tert-butyl ether Chemical compound COC(C)(C)C BZLVMXJERCGZMT-UHFFFAOYSA-N 0.000 description 2
- 244000061176 Nicotiana tabacum Species 0.000 description 2
- 235000002637 Nicotiana tabacum Nutrition 0.000 description 2
- 108091028043 Nucleic acid sequence Proteins 0.000 description 2
- SMFGCTXUBWEPKM-KBPBESRZSA-N Phe-Leu-Gly Chemical compound OC(=O)CNC(=O)[C@H](CC(C)C)NC(=O)[C@@H](N)CC1=CC=CC=C1 SMFGCTXUBWEPKM-KBPBESRZSA-N 0.000 description 2
- ABEFOXGAIIJDCL-SFJXLCSZSA-N Phe-Thr-Trp Chemical compound C([C@H](N)C(=O)N[C@@H]([C@H](O)C)C(=O)N[C@@H](CC=1C2=CC=CC=C2NC=1)C(O)=O)C1=CC=CC=C1 ABEFOXGAIIJDCL-SFJXLCSZSA-N 0.000 description 2
- 108091007187 Reductases Proteins 0.000 description 2
- 241001052560 Thallis Species 0.000 description 2
- UEXPMFIAZZHEAD-HSHDSVGOSA-N Val-Thr-Trp Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CC1=CNC2=CC=CC=C21)C(=O)O)NC(=O)[C@H](C(C)C)N)O UEXPMFIAZZHEAD-HSHDSVGOSA-N 0.000 description 2
- 238000002835 absorbance Methods 0.000 description 2
- 230000009286 beneficial effect Effects 0.000 description 2
- 239000000872 buffer Substances 0.000 description 2
- 230000008859 change Effects 0.000 description 2
- 238000012512 characterization method Methods 0.000 description 2
- 238000010276 construction Methods 0.000 description 2
- 235000019253 formic acid Nutrition 0.000 description 2
- 230000014509 gene expression Effects 0.000 description 2
- 229910052739 hydrogen Inorganic materials 0.000 description 2
- 239000001257 hydrogen Substances 0.000 description 2
- 230000006698 induction Effects 0.000 description 2
- 238000005374 membrane filtration Methods 0.000 description 2
- 230000011987 methylation Effects 0.000 description 2
- 238000007069 methylation reaction Methods 0.000 description 2
- 239000011259 mixed solution Substances 0.000 description 2
- DPNGWXJMIILTBS-UHFFFAOYSA-N myosmine Chemical compound C1CCN=C1C1=CC=CN=C1 DPNGWXJMIILTBS-UHFFFAOYSA-N 0.000 description 2
- 230000003287 optical effect Effects 0.000 description 2
- 239000008055 phosphate buffer solution Substances 0.000 description 2
- 238000012163 sequencing technique Methods 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 238000012795 verification Methods 0.000 description 2
- XCZXVTHYGSMQGH-NAKRPEOUSA-N Ala-Ile-Met Chemical compound C[C@H]([NH3+])C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CCSC)C([O-])=O XCZXVTHYGSMQGH-NAKRPEOUSA-N 0.000 description 1
- YNDLOUMBVDVALC-ZLUOBGJFSA-N Asn-Ala-Ala Chemical compound C[C@@H](C(=O)N[C@@H](C)C(=O)O)NC(=O)[C@H](CC(=O)N)N YNDLOUMBVDVALC-ZLUOBGJFSA-N 0.000 description 1
- 108020004705 Codon Proteins 0.000 description 1
- 238000002965 ELISA Methods 0.000 description 1
- 239000006142 Luria-Bertani Agar Substances 0.000 description 1
- UIJVKVHLCQSPOJ-XIRDDKMYSA-N Lys-Ser-Trp Chemical compound NCCCC[C@H](N)C(=O)N[C@@H](CO)C(=O)N[C@@H](Cc1c[nH]c2ccccc12)C(O)=O UIJVKVHLCQSPOJ-XIRDDKMYSA-N 0.000 description 1
- 102000016943 Muramidase Human genes 0.000 description 1
- 108010014251 Muramidase Proteins 0.000 description 1
- 108010062010 N-Acetylmuramoyl-L-alanine Amidase Proteins 0.000 description 1
- BQVUABVGYYSDCJ-UHFFFAOYSA-N Nalpha-L-Leucyl-L-tryptophan Natural products C1=CC=C2C(CC(NC(=O)C(N)CC(C)C)C(O)=O)=CNC2=C1 BQVUABVGYYSDCJ-UHFFFAOYSA-N 0.000 description 1
- 108010040201 Polymyxins Proteins 0.000 description 1
- 244000018795 Prunus mume Species 0.000 description 1
- 235000011158 Prunus mume Nutrition 0.000 description 1
- 240000005049 Prunus salicina Species 0.000 description 1
- MQUZANJDFOQOBX-SRVKXCTJSA-N Ser-Phe-Ser Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CO)C(O)=O MQUZANJDFOQOBX-SRVKXCTJSA-N 0.000 description 1
- PMZURENOXWZQFD-UHFFFAOYSA-L Sodium Sulfate Chemical compound [Na+].[Na+].[O-]S([O-])(=O)=O PMZURENOXWZQFD-UHFFFAOYSA-L 0.000 description 1
- 241000208292 Solanaceae Species 0.000 description 1
- 241000365977 Streptomyces sp. GF3587 Species 0.000 description 1
- 102000003929 Transaminases Human genes 0.000 description 1
- 108090000340 Transaminases Proteins 0.000 description 1
- 241000209140 Triticum Species 0.000 description 1
- 235000021307 Triticum Nutrition 0.000 description 1
- SUGLEXVWEJOCGN-ONUFPDRFSA-N Trp-Thr-Trp Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CC1=CNC2=CC=CC=C21)C(=O)O)NC(=O)[C@H](CC3=CNC4=CC=CC=C43)N)O SUGLEXVWEJOCGN-ONUFPDRFSA-N 0.000 description 1
- NVZVJIUDICCMHZ-BZSNNMDCSA-N Tyr-Phe-Ser Chemical compound [H]N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CO)C(O)=O NVZVJIUDICCMHZ-BZSNNMDCSA-N 0.000 description 1
- RFKJNTRMXGCKFE-FHWLQOOXSA-N Val-Leu-Trp Chemical compound C1=CC=C2C(C[C@H](NC(=O)[C@@H](NC(=O)[C@@H](N)C(C)C)CC(C)C)C(O)=O)=CNC2=C1 RFKJNTRMXGCKFE-FHWLQOOXSA-N 0.000 description 1
- 108010070783 alanyltyrosine Proteins 0.000 description 1
- 229930013930 alkaloid Natural products 0.000 description 1
- 150000003797 alkaloid derivatives Chemical class 0.000 description 1
- QFAADIRHLBXJJS-ZAZJUGBXSA-N amastatin Chemical compound CC(C)C[C@@H](N)[C@H](O)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](C(C)C)C(=O)N[C@H](C(O)=O)CC(O)=O QFAADIRHLBXJJS-ZAZJUGBXSA-N 0.000 description 1
- 108010052590 amastatin Proteins 0.000 description 1
- 150000001412 amines Chemical class 0.000 description 1
- 230000003321 amplification Effects 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 239000011248 coating agent Substances 0.000 description 1
- 238000000576 coating method Methods 0.000 description 1
- 238000001816 cooling Methods 0.000 description 1
- 125000004122 cyclic group Chemical group 0.000 description 1
- 230000009089 cytolysis Effects 0.000 description 1
- 230000001419 dependent effect Effects 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 238000004945 emulsification Methods 0.000 description 1
- RTZKZFJDLAIYFH-UHFFFAOYSA-N ether Substances CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 1
- 238000003810 ethyl acetate extraction Methods 0.000 description 1
- 239000000706 filtrate Substances 0.000 description 1
- 238000001914 filtration Methods 0.000 description 1
- 239000008098 formaldehyde solution Substances 0.000 description 1
- 235000009018 li Nutrition 0.000 description 1
- 229960000274 lysozyme Drugs 0.000 description 1
- 235000010335 lysozyme Nutrition 0.000 description 1
- 239000004325 lysozyme Substances 0.000 description 1
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 238000002703 mutagenesis Methods 0.000 description 1
- 231100000350 mutagenesis Toxicity 0.000 description 1
- 238000003199 nucleic acid amplification method Methods 0.000 description 1
- 238000005457 optimization Methods 0.000 description 1
- 238000004321 preservation Methods 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 238000011084 recovery Methods 0.000 description 1
- 230000009467 reduction Effects 0.000 description 1
- 230000008929 regeneration Effects 0.000 description 1
- 238000011069 regeneration method Methods 0.000 description 1
- 238000007363 ring formation reaction Methods 0.000 description 1
- 102220041265 rs587778687 Human genes 0.000 description 1
- 238000005070 sampling Methods 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 230000009466 transformation Effects 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/0004—Oxidoreductases (1.)
- C12N9/0012—Oxidoreductases (1.) acting on nitrogen containing compounds as donors (1.4, 1.5, 1.6, 1.7)
- C12N9/0026—Oxidoreductases (1.) acting on nitrogen containing compounds as donors (1.4, 1.5, 1.6, 1.7) acting on CH-NH groups of donors (1.5)
- C12N9/0028—Oxidoreductases (1.) acting on nitrogen containing compounds as donors (1.4, 1.5, 1.6, 1.7) acting on CH-NH groups of donors (1.5) with NAD or NADP as acceptor (1.5.1)
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N11/00—Carrier-bound or immobilised enzymes; Carrier-bound or immobilised microbial cells; Preparation thereof
- C12N11/02—Enzymes or microbial cells immobilised on or in an organic carrier
- C12N11/08—Enzymes or microbial cells immobilised on or in an organic carrier the carrier being a synthetic polymer
- C12N11/089—Enzymes or microbial cells immobilised on or in an organic carrier the carrier being a synthetic polymer obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
- C12N15/70—Vectors or expression systems specially adapted for E. coli
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/0004—Oxidoreductases (1.)
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/0004—Oxidoreductases (1.)
- C12N9/0006—Oxidoreductases (1.) acting on CH-OH groups as donors (1.1)
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P17/00—Preparation of heterocyclic carbon compounds with only O, N, S, Se or Te as ring hetero atoms
- C12P17/16—Preparation of heterocyclic carbon compounds with only O, N, S, Se or Te as ring hetero atoms containing two or more hetero rings
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P17/00—Preparation of heterocyclic carbon compounds with only O, N, S, Se or Te as ring hetero atoms
- C12P17/16—Preparation of heterocyclic carbon compounds with only O, N, S, Se or Te as ring hetero atoms containing two or more hetero rings
- C12P17/165—Heterorings having nitrogen atoms as the only ring heteroatoms
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P41/00—Processes using enzymes or microorganisms to separate optical isomers from a racemic mixture
- C12P41/002—Processes using enzymes or microorganisms to separate optical isomers from a racemic mixture by oxidation/reduction reactions
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Y—ENZYMES
- C12Y101/00—Oxidoreductases acting on the CH-OH group of donors (1.1)
- C12Y101/01—Oxidoreductases acting on the CH-OH group of donors (1.1) with NAD+ or NADP+ as acceptor (1.1.1)
- C12Y101/01047—Glucose 1-dehydrogenase (1.1.1.47)
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
- C12R2001/185—Escherichia
- C12R2001/19—Escherichia coli
Landscapes
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Organic Chemistry (AREA)
- Engineering & Computer Science (AREA)
- Genetics & Genomics (AREA)
- Wood Science & Technology (AREA)
- Zoology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- Biotechnology (AREA)
- General Health & Medical Sciences (AREA)
- Biochemistry (AREA)
- Microbiology (AREA)
- Biomedical Technology (AREA)
- Molecular Biology (AREA)
- Medicinal Chemistry (AREA)
- General Chemical & Material Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Physics & Mathematics (AREA)
- Biophysics (AREA)
- Plant Pathology (AREA)
- Analytical Chemistry (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Enzymes And Modification Thereof (AREA)
Abstract
本发明公开了一种亚胺还原酶突变体,以及这种亚胺还原酶突变体用于制备(S)‑尼古丁的制备方法。发明第一个目的是提供了一种亚胺还原酶突变体,解决现有技术使用酶得不到S构型的尼古丁,以及现有技术中酶活性低的问题,这种突变体可以将底物麦思明或4‑(甲胺基)‑1‑(吡啶‑3‑基)丁酮催化还原为(S)‑尼古丁。本发明第二个目的是提供这种亚胺还原酶与葡萄糖脱氢酶的共表达酶,具体是亚胺还原酶突变酶与葡萄糖脱氢酶的共表达酶,这种共表达酶应用与制备(S)‑尼古丁,降低了酶催化制备(S)‑尼古丁的成本。
Description
[技术领域]
本发明涉及医药化工领域,具体涉及一种亚胺还原酶突变体,以及这种突变体的应用,还涉及这种亚胺还原酶突变体与葡萄糖脱氢酶的共表达酶以及这种共表达酶的应用。
[背景技术]
(S)-尼古丁是一种存在茄科植物的生物碱,也是烟草的重要成分。虽然有化学合成法的相关报道,但是合成的是(R,S)-尼古丁,经拆分后才能获得(S)-尼古丁。所以化学合成成本比从烟叶中提取烟碱的成本要高。
使用酶催化的方式催化制备(S)-尼古丁,大大降低制备的成本,而且绿色环保。
Koichi Mitsukura等人在2010年发现Streptomyces sp.GF3587和3546的亚胺还原酶(Org.Biomol.Chem.,2010,8,4533-4535)。使用这两种酶将2-MPN还原为S-2MP或者R-2MP。
专利WO2014174505公开使用sp.GF3587和3546制备尼古丁的方法。反应底物为4-(甲胺基)-1-(吡啶-3-基)丁酮,转化率仅有23%,而且仅能制备(R)-尼古丁。
专利WO2020098978中公开了使用亚胺还原酶制备(S)-尼古丁方法。亚胺还原酶催化麦思明为去甲烟碱,去甲烟碱甲基化后得到(S)-尼古丁。使用的酶转化率在8小时后的可以达到77%,24小时后达到99%,专利中使用的酶来自Enzymicals。
现在公开使用酶催化制备(S)-尼古丁的技术,酶活性不够,或者仅能制备(R)-尼古丁。为此需要一种新的亚胺还原酶,能够催化底物制备(S)-尼古丁,而且酶的活性更高,在较高的底物起始浓度下,具有较高的转化率和光学纯度。
另外,亚胺还原酶是一种依赖辅酶NADPH的氧化酶,而专利WO2014174505、WO2020098978中的亚胺还原酶和葡萄糖脱氢酶都是单独制备后混合使用,这种方式增加了制备酶的步骤,提高了获得酶的成本。李骥璇等(生物技术通报,2019,35(1):105-111)公开了S-亚胺还原酶和葡萄糖脱氢酶共表达系统的构建及手性胺的合成,可以将亚胺还原酶和葡萄糖脱氢酶共同表达,降低了使用酶的成本。但是李骥璇等公开的共表达系统仅公开了催化2-MPN还原为S-2MP的反应,而且一般的葡萄糖脱氢酶的活性要大于亚胺还原酶,共表达系统的亚胺还原酶与葡萄糖脱氢酶是等量的,这使得葡萄糖脱氢酶过量,在实际生产中,过量的酶易引起乳化,给后面的提纯带来不便。为了解决这个问题,需要一种活性更高的亚胺还原酶与葡萄糖脱氢酶一起形成共表达酶,并且能够催化底物生成(S)-尼古丁。
[发明内容]
本发明第一个目的是提供了一种亚胺还原酶突变体,解决现有技术使用酶得不到S构型的尼古丁,以及现有技术中酶活性低的问题,这种突变体可以将底物麦思明或4-(甲胺基)-1-(吡啶-3-基)丁酮催化还原为(S)-尼古丁。
本发明第二个目的是提供这种亚胺还原酶与葡萄糖脱氢酶的共表达酶,具体是亚胺还原酶突变酶与葡萄糖脱氢酶的共表达酶,这种共表达酶应用与制备(S)-尼古丁,降低了酶催化制备(S)-尼古丁的成本。
为了达到发明目的,本发明的技术方案如下:
亚胺还原酶突变体,与氨基酸序列SEQ ID NO:1相比,存在残基差异包括V171、A172、Y230中的一个或多个。
SEQ ID NO:1是来源于气单胞菌Aeromonas veronii的野生型亚胺还原酶,经过突变后得到本发明的亚胺还原酶突变体,突变点包括V171、A172、Y230中的一个或多个。通过对比催化底物麦思明的活性,突变后的亚胺还原酶的活性是野生型的酶的活性31到50倍。
所述残基差异包括V171Y/N/A/S、A172V/F、Y230G/A/T中的一个或多个。
残基差异还包括:S14V/L/D、S33R/G/D/L/A/Q/N、L118V/M、C119/T/D、S130C/T、Q202L/A/S、L206Y/F、I209F/T、K234W/L/F/V、T235L/S、A239Y/L、V277L中的一个或多个。
亚胺还原酶突变体的氨基酸序列选自SEQ ID NO:3、5、7、9、11、13、15、17、19、21、23、25、27、29、31、33、35、37、39、41、43、45、47、49、51、53、55、57、59、61、63、65、67、69、71、73、75、77、79、81、83、85、87、89、91、93、95、97、99、101、105或107。本发明的亚胺还原酶突变体用于制备(S)-尼古丁,在合适的条件下,亚胺还原酶突变体催化底物I
或底物II
直接或经过化学反应得到(S)-尼古丁。
合适的条件是指当底物为麦斯时,将底物I、辅酶、葡萄糖、葡萄糖脱氢酶、缓冲液与亚胺还原酶混合后,反应得到化合物III(S)-去甲烟碱。
优选的,底物I的载量为10g/L至300g/L;酶的浓度为1g/L至10g/L;辅酶的用量为1g/L;葡萄糖脱氢酶的用量为2g/L;pH的范围为6至7;反应的温度为23℃至30℃;反应时间为15小时至24小时。
优选的,底物I的载量包括10g/L、50g/L、100g/L、150g/L、200g/L、250g/L、300g/L;
催化底物I使用的酶的浓度为1g/L、2g/L、3g/L、4g/L、5g/L、6g/L、7g/L、8g/L、9g/L、10g/L。
催化底物I时的反应温度为23℃、24℃、25℃、26℃、27℃、28℃、29℃、30℃。
催化底物I时的反应时间15小时、16小时、17小时、18小时、19小时、20小时、21小时、22小时、23小时、24小时。
化合物III(S)-去甲烟碱甲基化得到(S)-尼古丁。如(S)-去甲烟碱与甲酸/甲醛反应得到(S)-尼古丁。
催化麦思明得到(S)-去甲烟碱,再经甲基化得到(S)-尼古丁的反应如下:
底物II环化后经亚胺还原酶的催化得到(S)-尼古丁;
或底物II的盐经脱盐、环化经亚胺还原酶的催化得到(S)-尼古丁;
所述底物II的盐包括盐酸盐、二盐酸盐、氢溴酸盐、二氢溴酸盐、硫酸盐或硫酸氢盐。
底物II可环化为烯胺化合物或亚胺正离子化合物,结构如下:
亚胺还原酶催化时的合适条件为将底物II、辅酶、葡萄糖、葡萄糖脱氢酶、缓冲液与亚胺还原酶混合后,反应得到(S)-尼古丁。
优选的,底物II的载量为10g/L至300g/L;酶的浓度为1g/L至10g/L;辅酶的用量为1g/L;葡萄糖脱氢酶的用量为2g/L;pH的范围为6至7;反应的温度为23℃至30℃;反应时间为15小时至24小时。
优选的,底物II的载量包括10g/L、50g/L、100g/L、150g/L、200g/L、250g/L、300g/L;
催化底物II使用的酶的浓度为1g/L、2g/L、3g/L、4g/L、5g/L、6g/L、7g/L、8g/L、9g/L、10g/L。
催化底物II时的反应温度为23℃、24℃、25℃、26℃、27℃、28℃、29℃、30℃。
催化底物II时的反应时间15小时、16小时、17小时、18小时、19小时、20小时、21小时、22小时、23小时、24小时。
氨基酸序列SEQ ID NO:3、5、7、9、11、13、15、17、19、21、23、25、27、29、31、33、35、37、39、41、43、45、47、49、51、53、55、57、59、61、63、65、67、69、71、73、75、77、79、81、83、85、87、89、91、93、95、97、99、101、105或107;
对应核酸序列选自SEQ ID NO:4、6、8、10、12、14、16、18、20、22、24、26、28、30、32、34、36、38、40、42、44、46、48、50、52、54、56、58、60、62、64、66、68、70、72、74、76、78、80、82、84、86、88、90、92、94、96、98、100、102、104、106或108。
一种表达载体,包括以上的核酸。
一种载体细胞,以上的核苷酸或包括以上核酸的载体。
一种亚胺还原酶突变体的生产方法。将所述的亚胺还原酶突变体和葡萄糖脱氢酶共表达。用同时含有亚胺还原酶突变体基因与葡萄糖脱氢酶基因的细胞生产共表达酶。
具体来说,可以将这些核苷酸序列接入质粒中,如pET-30a(+),制备成载体。
将这些重组后的载体转入工程细菌,如大肠杆菌BL21(DE3),制备得到载体细胞。
亚胺还原酶突变体与葡萄糖脱氢酶的基因在同一个工程细菌中表达,制备成共表达酶。解决了单独制备葡萄糖脱氢酶的成本,亚胺还原酶与葡萄糖脱氢酶共同表达实现体系中辅酶的重复使用。
一种具体的制备亚胺还原酶突变体和葡萄糖脱氢酶共表达酶的方法:通过带有限制性内切酶位点的引物将葡萄糖脱氢酶基因从载体上扩增下来,分别用限制性内切酶BamHI和XhoI对带有亚胺还原酶基因片段的载体和扩增的葡萄糖脱氢酶基因片段进行双酶切,胶回收酶切片段,并用T4DNA连接酶连接后转化大肠杆菌BL21感受态细胞,获得同时含有亚胺还原酶基因和葡萄糖脱氢酶基因的重组菌,将上述重组菌培养,然后诱导、离心收集菌体、重悬,重悬后对细胞进行超声破碎,再将破碎后溶液进行冷冻干燥,获得亚胺还原酶和葡萄糖脱氢酶共表达酶粉。
将亚胺还原酶突变体和葡萄糖脱氢酶共表达酶应用于制备(S)-尼古丁:在合适条件下,如权利要求10所述的亚胺还原酶突变体和葡萄糖脱氢酶共表达酶催化底物I
或底物II
直接或经过化学反应得到(S)-尼古丁。
优选的,将底物I、辅酶、葡萄糖、缓冲液与亚胺还原酶和葡萄糖脱氢酶共表达酶混合后,反应得到化合物III(S)-去甲烟碱,然后将化合物III(S)-去甲烟碱甲基化得到(S)-尼古丁,
底物II环化后经亚胺还原酶和葡萄糖脱氢酶共表达酶催化得到(S)-尼古丁;
或底物II的盐经脱盐、环化经亚胺还原酶的催化得到(S)-尼古丁;
所述底物II的盐包括盐酸盐、二盐酸盐、氢溴酸盐、二氢溴酸盐、硫酸盐或硫酸氢盐。
相对于现有技术,本发明取得的有益效果:
1、经本发明亚胺还原酶突变体催化底物II或其盐为(S)-尼古丁,转化率为99.9%,化学纯度为99.9%,底物的载量为300g/L。
2、本发明的亚胺还原酶突变体可以催化底物I为(S)-去甲烟碱,酶的活性更高,且提高了底物的载量。
3、通过本发明的亚胺还原酶和葡萄糖脱氢酶共表达酶,在同一个工程菌中一起表达两种酶,在使用亚胺还原酶制备(S)-尼古丁的体系中实现辅酶循环再生的同时,减少了再次制备葡萄糖脱氢酶的步骤,降低了成本。
[附图说明]
图1为(S)-(3-吡咯烷-2-基)吡啶的合成路线图;
图2为(S)-(3-吡咯烷-2-基)吡啶标准品的反相非手性HPLC图谱;
图3为底物麦斯明标准品的反相非手性HPLC图谱;
图4为实施例5反应中控的反相非手性HPLC图谱(SEQ ID NO:1样品);
图5为(R,S)-(3-吡咯烷-2-基)吡啶的正相手性HPLC图谱;
图6为(S)-(3-吡咯烷-2-基)吡啶标准品的正相手性HPLC图谱;
图7为实施例5合成的(S)-(3-吡咯烷-2-基)吡啶的正相手性HPLC图谱(SEQ IDNO:1样品);
图8为实施例5合成的(S)-(3-吡咯烷-2-基)吡啶的正相手性HPLC图谱(SEQ IDNO:17样品);
图9为亚胺还原酶和葡萄糖脱氢酶共表达质粒图谱;
[具体实施例]
为进一步阐述本发明所采取的技术手段及其效果,以下结合实施例和附图对本发明作进一步地说明。可以理解的是,此处所描述的具体实施方式仅仅用于解释本发明,而非对本发明的限定。
实施例中未注明具体技术或条件者,按照本领域内的文献所描述的技术或条件,或者按照产品说明书进行。所用试剂或仪器未注明生产厂商者,均为可通过正规渠道商购获得的常规产品。
实施例1亚胺还原酶的表达及酶粉制备
(1)将来源于气单胞菌Aeromonas veronii的野生型亚胺还原酶的编码基因SEQID NO:2送至南京金斯瑞公司进行密码子优化和全基因合成,连接入质粒pET-30a(+)中;将重组质粒转入大肠杆菌BL21(DE3)中,将菌液涂布于含50μg/mL卡那霉素的LB琼脂平板上,置于37℃下培养过夜;挑取转化子进行测序验证,将验证正确的转化子命名为E.coliBL21/pET30a-No.2。
(2)将上述重组菌E.coli BL21/pET30a-No.2接种于5mL含有50μg/mL卡那霉素的LB液体培养基中,置于37℃下培养过夜;取1mL菌液接种于100mL含有50μg/mL卡那霉素的LB液体培养基中,置于37℃下培养3h,然后加入50μL 1M IPTG,25℃下诱导16小时;离心(4000rpm,4℃,10min)收集菌体,加入4倍体积的PBS缓冲液(pH=7.0)重悬,重悬后对细胞进行超声破碎,再将溶液进行冷冻干燥,获得亚胺还原酶酶粉,野生型亚胺还原酶的氨基酸序列如SEQ ID NO:1所示。
实施例2亚胺还原酶突变体文库的构建
本实施例以野生型亚胺还原酶的编码基因SEQ ID NO:1为模板,使用随机突变试剂盒(碧云天)进行易错PCR,引物见下表,使用DNA片段回收试剂盒(上海生工)并按照说明书回收上述易错PCR产物。
分别用限制性内切酶NdeI和BamHI对易错PCR产物和pET-30a(+)质粒进行双酶切,将回收的酶切产物用T4DNA连接酶连接在线性化pET-30a(+)质粒中,22℃下连接1小时。将连接产物转化入大肠杆菌BL21(DE3)感受态细胞(上海生工),37℃过夜培养,获得随机突变文库。
表1 易错PCR引物序列
序列编号 | 引物序列 |
SEQ ID NO:109 | TATA<u>CATATG</u>CGCCATCTGAGCGTGATTGG |
SEQ ID NO:110 | TTC<u>GGATCC</u>TTACTGCGCCGCGCCGTTGC |
突变文库经过筛选后,将获得的有益突变酶基因SEQ ID NO:37作为模板进行定点饱和突变及再次进行易错PCR,进行多轮次的建库与筛选。
实施例3亚胺还原酶突变体的初筛
(1)挑取文库中的菌落至96孔微量培养板上,培养板的每孔含有150μL LB液体培养基和50μg/mL卡那霉素,将菌体置于37℃、220rpm摇床中培养过夜;将20μL菌液转接至含有380μL LB液体培养基和50μg/mL卡那霉素的96孔板中,继续在37℃、220rpm的摇床中培养2~3h,添加IPTG至终浓度为0.4mM进行诱导,然后降温至25℃培养过夜(至少16小时)。
(2)将96孔板离心(10min,4000rpm,25℃)收集菌体,用200μL裂解液(含1g/L溶菌酶和0.5g/L硫酸多粘菌素B)重悬菌体,于25℃、600rpm下振荡2h进行裂解,裂解后的细胞碎片通过离心(4000rpm,4℃,10min)进行沉淀,获得上清液即粗酶液,用于酶标仪进行活性初筛。
(3)反应混合液的配制:取1mL麦斯明溶液(pH6.0 50mM),0.25mL NADPH溶液(20mM),13.75mL磷酸盐缓冲液(pH6.0 0.1M)置于加样槽中,混匀即得反应混合液。用液体工作站进行液体分装,取酶标板每孔分装150μL反应混合液,然后加入上述粗酶液50μL,并立即于25℃检测340nm处吸光光度值的变化。
酶活性定义:单位时间内生成1μmol产物所需要的酶的用量为一个酶活单位。
其中:
ΔA340:吸光值变化值;
V0:反应总体积;
N:酶液稀释倍数;
T:反应时间;
V1:加酶量;
6.22为NADPH斜率常数
实施例4亚胺还原酶突变体的复筛
(1)将实施例3中酶活性高于母本的突变体接种于100mL含50μg/mL卡那霉素的LB液体培养基中,37℃振荡培养至OD600=0.6时,加入IPTG至终浓度为0.5mM,在25℃下进行诱导表达。诱导16h后,离心(10min,4000rpm,25℃)收集菌体。菌体用超声破碎仪(JY92-2D,宁波新芝生物科技股份有限公司)破碎细胞,离心(4℃,10000rpm,20min)获得上清液,部分用于活性检测,剩余上清液进行冷冻干燥,制备酶粉。
(2)取5mL离心管,加入50mg底物麦斯明,加入2mL磷酸盐缓冲液(pH6.0 0.1M)。再向反应瓶中加入0.5mg NADP和90mg葡萄糖,搅拌至完全溶解,再向其中分别加入葡萄糖脱氢酶5mg和待筛选粗酶液0.2mL,混匀并升温至25℃,以300r/min搅拌反应24h。反应结束后,分别取反应液100μL加入到900μL乙腈中充分振荡,使用0.22μm过滤膜过滤,进行HPLC分析。
选取催化活性优于母本的突变体进行测序,分析突变位点,获得亚胺还原酶突变体的编码基因SEQ ID NO:4、6、8、10、12、14、16、18、20、22、24、26、28、30、32、34、36、38、40、42、44、46、48、50、52、54、56、58、60、62、64、66、68、70、72、74、76、78、80、82、84、86、88、90、92、94、96、98、100、102、104、106或108,相应的转氨酶突变体的氨基酸序列分别如SEQ IDNO:3、5、7、9、11、13、15、17、19、21、23、25、27、29、31、33、35、37、39、41、43、45、47、49、51、53、55、57、59、61、63、65、67、69、71、73、75、77、79、81、83、85、87、89、91、93、95、97、99、101、105或107所示。
不同亚胺还原酶突变体的活性检测结果见表2。
表2 亚胺还原酶突变体的突变位点与活性检测
注:++表示酶活性是SEQ ID NO:1的1.2至3倍;+++表示酶活性是SEQ ID NO:1的5至10倍;++++表示酶活性是SEQ ID NO:1的11至20倍;+++++表示酶活性是SEQ ID NO:1的21至30倍;++++++表示酶活性是SEQ ID NO:1的31至50倍;
实施例5表2中标为“++”的亚胺还原酶突变体在制备S-(3-吡咯烷-2-基)吡啶中的应用
取10mL反应瓶,加入100mg底物麦斯明,加入4mL磷酸盐缓冲液(pH6.0 0.1M),调节pH至6.0。再向反应瓶中加入2.5mg NADP和180mg葡萄糖,搅拌至完全溶解,再向其中分别加入10mg葡萄糖脱氢酶和50mg亚胺还原酶突变体酶粉,混匀并升温至25℃,以300r/min搅拌反应24h。反应结束后,分别取反应液100μL加入到900μL乙腈中充分振荡,使用0.22μm过滤膜过滤,进行HPLC分析。取反应液500μL,调pH11以上,加1000μL正己烷萃取,取正己烷层过膜后供正相HPLC分析手性。SEQ ID NO:1转化率图谱见图4,SEQ ID NO:1手性检测图谱见图7,SEQ ID NO:17手性检测图谱见图8.
SEQ ID NO:1及活性最高的三个突变体转化率及手性结果如表3所示。
表3
实施例6表2中标为“+++”的亚胺还原酶突变体在制备S-(3-吡咯烷-2-基)吡啶中的应用
取10mL反应瓶,加入250mg底物麦斯明,加入4mL磷酸盐缓冲液(pH6.0 0.1M),调节pH至6.0。再向反应瓶中加入2.5mg NADP和450mg葡萄糖,搅拌至完全溶解,再向其中分别加入10mg葡萄糖脱氢酶和50mg亚胺还原酶突变体酶粉,混匀并升温至25℃,以300r/min搅拌反应24h。反应结束后,分别取反应液100μL加入到900μL乙腈中充分振荡,使用0.22μm过滤膜过滤,进行HPLC分析。取反应液500μL,调pH11以上,加1000μL正己烷萃取,取正己烷层过膜后供正相HPLC分析手性。
SEQ ID NO:1及活性最高的三个突变体转化率及手性结果如表4所示。
表4
酶 | 转化率% | ee% |
SEQ ID NO:1 | 18.7 | 95.1 |
SEQ ID NO:25 | 98.1 | 99.3 |
SEQ ID NO:31 | 98.1 | 99.7 |
SEQ ID NO:33 | 90.3 | 99.6 |
实施例7表2中标为“++++”的亚胺还原酶突变体在制备S-(3-吡咯烷-2-基)吡啶中的应用
取10mL反应瓶,加入500mg底物麦斯明,加入4mL磷酸盐缓冲液(pH6.0 0.1M),调节pH至6.0。再向反应瓶中加入2.5mg NADP和900mg葡萄糖,搅拌至完全溶解,再向其中分别加入10mg葡萄糖脱氢酶和50mg亚胺还原酶突变体酶粉,混匀并升温至25℃,以300r/min搅拌反应24h。反应结束后,分别取反应液100μL加入到900μL乙腈中充分振荡,使用0.22μm过滤膜过滤,进行HPLC分析。取反应液500μL,调pH11以上,加1000μL正己烷萃取,取正己烷层过膜后供正相HPLC分析手性。
SEQ ID NO:1及活性最高的三个突变体转化率及手性结果如表5所示。
表5
酶 | 转化率% | ee% |
SEQ ID NO:1 | 8.9 | 95.6 |
SEQ ID NO:37 | 99.9 | 99.6 |
SEQ ID NO:47 | 99.8 | 99.9 |
SEQ ID NO:53 | 95.5 | 99.7 |
实施例8表2中标为“+++++”的亚胺还原酶突变体在制备S-(3-吡咯烷-2-基)吡啶中的应用
取10mL反应瓶,加入1000mg底物麦斯明,加入4mL磷酸盐缓冲液(pH6.0 0.1M),调节pH至6.0。再向反应瓶中加入5mg NADP和1800mg葡萄糖,搅拌至完全溶解,再向其中分别加入10mg葡萄糖脱氢酶和50mg亚胺还原酶突变体酶粉,混匀并升温至25℃,以300r/min搅拌反应16h。反应结束后,分别取反应液100μL加入到900μL乙腈中充分振荡,使用0.22μm过滤膜过滤,进行HPLC分析。取反应液500μL,调pH11以上,加1000μL正己烷萃取,取正己烷层过膜后供正相HPLC分析手性。
SEQ ID NO:37及活性最高的三个突变体转化率及手性结果如表6所示。
表6
酶 | 转化率% | ee% |
SEQ ID NO:37 | 46.8 | 99.7 |
SEQ ID NO:61 | 99.3 | 99.8 |
SEQ ID NO:65 | 99.5 | 99.7 |
SEQ ID NO:71 | 99.9 | 99.6 |
实施例9表2中标为“++++++”的亚胺还原酶突变体在制备S-(3-吡咯烷-2-基)吡啶中的应用
取20mL反应瓶,加入3g底物麦斯明,加入7mL磷酸盐缓冲液(pH6.00.1M),调节pH至6.0。再向反应瓶中加入10mg NADP和5.4g葡萄糖,搅拌至完全溶解,再向其中分别加入20mg葡萄糖脱氢酶和100mg亚胺还原酶突变体酶粉,混匀并升温至25℃,以300r/min搅拌反应16h。反应结束后,分别取反应液100μL加入到900μL乙腈中充分振荡,使用0.22μm过滤膜过滤,进行HPLC分析。取反应液500μL,调pH11以上,加1000μL正己烷萃取,取正己烷层过膜后供正相HPLC分析手性。
SEQ ID NO:37及活性最高的三个突变体转化率及手性结果如表7所示。
表7
酶 | 转化率% | ee% |
SEQ ID NO:37 | 33.3 | 99.8 |
SEQ ID NO:81 | 99.1 | 99.9 |
SEQ ID NO:85 | 99.6 | 99.6 |
SEQ ID NO:95 | 99.9 | 99.9 |
实施例10以4-(甲胺基)-1-(吡啶-3-基)丁酮为底物筛选表2中亚胺还原酶突变体
取5mL离心管,加入30mg底物4-(甲胺基)-1-(吡啶-3-基)丁酮,加入2mL磷酸盐缓冲液(pH6.0 0.1M)。再向反应瓶中加入0.3mg NADP和50mg葡萄糖,搅拌至完全溶解,再向其中分别加入3mg葡萄糖脱氢酶和30mg亚胺还原酶突变体酶粉,混匀并升温至25℃,以300r/min搅拌反应24h。反应结束后,分别取反应液100μL加入到900μL乙腈中充分振荡,使用0.22μm过滤膜过滤,进行HPLC分析,以(S)-尼古丁的面积比为转化率。对转化率高的样品分别取反应液1mL进行乙酸乙酯萃取,用于ee值检测。
SEQ ID NO:1及活性较高的部分突变体转化率及手性结果如表8所示。
表8
实施例11SEQ ID NO:37所示亚胺还原酶突变体在(S)-尼古丁制备中的应用
向50mL三口圆底烧瓶中加入4.5g 4-(甲胺基)-1-(吡啶-3-基)丁酮,加入20mL0.1M的磷酸盐缓冲液,调节pH至7.0。再向反应瓶中加入4.8g葡萄糖,搅拌至完全溶解。,0.04g葡萄糖脱氢酶和0.008g NADP盐,搅拌至完全溶解。然后将第二个烧瓶中的溶液缓慢加入第一个烧瓶中,升温至30℃,以300r/min搅拌反应16h。取样检测转化率99%,终止反应,过滤,滤液用氢氧化钠溶液调节至pH=10后用甲基叔丁基醚萃取,无水硫酸钠干燥,浓缩后得(S)-尼古丁2.6g。产品纯度99%,光学纯度100%。
对制备得到的(S)-尼古丁进行核磁氢谱表征,数据为:1H-NMR(400MHz,CDCl3):δppm 8.54(1H,d),8.50(1H,dd),7.70(1H,dt),7.24-7.27(1H,m),3.22-3.27(1H,m),3.08(1H,t),2.27-2.34(1H,m),2.17-2.24(1H,m),2.16(3H,m),1.91-2.02(1H,m),1.79-1.87(1H,m),1.68-1.76(1H,m)。表明(S)-尼古丁被成功合成。
实施例12SEQ ID NO:47所示亚胺还原酶突变体在(S)-尼古丁制备中的应用
向50mL三口圆底烧瓶中加入4.5g 4-(甲胺基)-1-(吡啶-3-基)丁酮,加入20mL0.1M的磷酸盐缓冲液,调节pH至7.0。再向反应瓶中加入4.8g葡萄糖,搅拌至完全溶解。在另一个50mL烧瓶中加入10mL 0.1M的磷酸盐缓冲液,0.3g SEQ ID NO:53所示亚胺还原酶,0.04g葡萄糖脱氢酶和0.008g NADP盐,搅拌至完全溶解。然后将第二个烧瓶中的溶液缓慢加入第一个烧瓶中,升温至30℃,以300r/min搅拌反应16h。取反应液100μL加入到900μL乙腈中充分振荡,使用0.22μm过滤膜过滤,进行HPLC分析,以(S)-尼古丁的面积比为转化率。取反应液1mL进行乙酸乙酯萃取,用于ee值检测。转化率99.5%,ee值99.6%。
实施例13SEQ ID NO:61所示亚胺还原酶突变体在(S)-尼古丁制备中的应用
向50mL三口圆底烧瓶中加入4.5g 4-(甲胺基)-1-(吡啶-3-基)丁酮,加入20mL0.1M的磷酸盐缓冲液,调节pH至7.0。再向反应瓶中加入4.8g葡萄糖,搅拌至完全溶解。在另一个50mL烧瓶中加入10mL 0.1M的磷酸盐缓冲液,0.3g SEQ ID NO:61所示亚胺还原酶,0.04g葡萄糖脱氢酶和0.008g NADP盐,搅拌至完全溶解。然后将第二个烧瓶中的溶液缓慢加入第一个烧瓶中,升温至30℃,以300r/min搅拌反应16h。取反应液100μL加入到900μL乙腈中充分振荡,使用0.22μm过滤膜过滤,进行HPLC分析,以(S)-尼古丁的面积比为转化率。取反应液1mL进行乙酸乙酯萃取,用于ee值检测。转化率93.2%,ee值99.7%。
实施例14SEQ ID NO:85所示亚胺还原酶突变体在(S)-尼古丁制备中的应用
向50mL三口圆底烧瓶中加入4.5g 4-(甲胺基)-1-(吡啶-3-基)丁酮,加入20mL0.1M的磷酸盐缓冲液,调节pH至7.0。再向反应瓶中加入4.8g葡萄糖,搅拌至完全溶解。在另一个50mL烧瓶中加入10mL 0.1M的磷酸盐缓冲液,0.3g SEQ ID NO:85所示亚胺还原酶,0.04g葡萄糖脱氢酶和0.008g NADP盐,搅拌至完全溶解。然后将第二个烧瓶中的溶液缓慢加入第一个烧瓶中,升温至30℃,以300r/min搅拌反应16h。取反应液100μL加入到900μL乙腈中充分振荡,使用0.22μm过滤膜过滤,进行HPLC分析,以(S)-尼古丁的面积比为转化率。取反应液1mL进行乙酸乙酯萃取,用于ee值检测。转化率100%,ee值99.8%。
实施例15亚胺还原酶与葡萄糖脱氢酶的共表达酶粉制备
通过带有限制性内切酶位点的引物(见下表)将葡萄糖脱氢酶基因从
pET30a-GDH(序列见SEQ ID NO:111)载体上扩增下来。分别用限制性内切酶BamHI和XhoI对带有亚胺还原酶基因片段的pET30a载体和扩增的葡萄糖脱氢酶基因片段进行双酶切。胶回收酶切片段,并用T4DNA连接酶连接后转化大肠杆菌BL21(DE3)感受态细胞,获得同时含有亚胺还原酶基因和葡萄糖脱氢酶基因的重组菌(质粒示意图见图9)。
表9 葡萄糖脱氢酶扩增引物
序列编号 | 引物序列 |
SEQ ID NO:112 | GCC<u>GGATCC</u>AATAATTTTGTTTAACTTTAAGAAGG |
SEQ ID NO:113 | GG<u>CTCGAG</u>TTAGCCACGACCCGCTTGAAAGC |
将上述重组菌接种于5mL含有50μg/mL卡那霉素的LB液体培养基中,置于37℃下培养过夜;取1mL菌液接种于100mL含有50μg/mL卡那霉素的LB液体培养基中,置于37℃下培养3h,然后加入50μL 1M IPTG,25℃下诱导16小时;离心(4000rpm,4℃,10min)收集菌体,加入4倍体积的PBS缓冲液(pH=7.0)重悬,重悬后对细胞进行超声破碎,再将溶液进行冷冻干燥,获得亚胺还原酶和葡萄糖脱氢酶共表达酶粉。
实施例16亚胺还原酶与葡萄糖脱氢酶的共表达酶粉在S-(3-吡咯烷-2-基)吡啶制备中的应用
取10mL反应瓶,加入750mg底物麦斯明,加入4mL磷酸盐缓冲液(pH6.0 0.1M),调节pH至6.0。再向反应瓶中加入5mg NADP和1200mg葡萄糖,搅拌至完全溶解,再向其中加入共表达酶粉(此处亚胺还原酶为SEQ ID NO:37所示突变酶),混匀并升温至25℃,以300r/min搅拌反应16h。反应结束后,分别取反应液100μL加入到900μL乙腈中充分振荡,使用0.22μm过滤膜过滤,进行HPLC分析。取反应液500μL,调pH11以上,加1000μL正己烷萃取,取正己烷层过膜后供正相HPLC分析手性。转化率99.6%,ee值99.8%。
实施例17亚胺还原酶与葡萄糖脱氢酶的共表达酶粉在(S)-尼古丁制备中的应用
向50mL三口圆底烧瓶中加入4.5g 4-(甲胺基)-1-(吡啶-3-基)丁酮,加入30mL0.1M的磷酸盐缓冲液,调节pH至7.0。再向反应瓶中加入4.8g葡萄糖和0.008g NADP盐,搅拌至完全溶解。直接加入0.3g共表达酶粉(此处亚胺还原酶为SEQ ID NO:37所示突变酶),搅拌至完全溶解,升温至30℃,以300r/min搅拌反应16h。取反应液100μL加入到900μL乙腈中充分振荡,使用0.22μm过滤膜过滤,进行HPLC分析,以(S)-尼古丁的面积比为转化率。取反应液1mL进行乙酸乙酯萃取,用于ee值检测。转化率99.6%,ee值99.8%。
实施例18用(S)-去甲烟碱制备(S)-尼古丁
将实施例5-9或16制备的(S)-去甲烟碱制备(S)-尼古丁。
向三个500mL三口瓶中均加入使用实施例16方法制备的107g(S)-去甲烟碱,80g37%甲醛溶液,升温到75℃。滴加60g 85%甲酸溶液,滴加完毕后,保温反应24h。反应完成后,加入氢氧化钠,调节pH=12。水相用甲叔醚萃取,合并萃取液,浓缩,减压蒸馏,得无色液体80g,即(S)-尼古丁。
对所得产物进行核磁氢谱表征,核磁数据结果为:1H-NMR(400MHz,CDCl3):δppm8.54(1H,d),8.50(1H,dd),7.70(1H,dt),7.24-7.27(1H,m),3.22-3.27(1H,m),3.08(1H,t),2.27-2.34(1H,m),2.17-2.24(1H,m),2.16(3H,m),1.91-2.02(1H,m),1.79-1.87(1H,m),1.68-1.76(1H,m),表明(S)-尼古丁被成功合成。
申请人声明,本发明通过上述实施例来说明本发明的详细方法,但本发明并不局限于上述详细方法,即不意味着本发明必须依赖上述详细方法才能实施。所属技术领域的技术人员应该明了,对本发明的任何改进,对本发明产品各原料的等效替换及辅助成分的添加、具体方式的选择等,均落在本发明的保护范围和公开范围之内。
序列表
<120> 亚胺还原酶突变体、亚胺还原酶和葡萄糖脱氢酶共表达酶及其应用
<160> 113
<170> SIPOSequenceListing 1.0
<210> 1
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 1
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Ser Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Ser Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Ser Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Val Ala Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Tyr Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 2
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 2
atgcgccatc tgagcgtgat tggcctgggc gcgatgggca gcgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtagtg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gtggcggcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgtat gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 3
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 3
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Ser Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Ser Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Ser Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Val Ala Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Ala Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 4
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 4
atgcgccatc tgagcgtgat tggcctgggc gcgatgggca gcgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtagtg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gtggcggcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccggcg gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 5
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 5
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Ser Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Ser Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Ser Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Tyr Ala Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Tyr Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 6
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 6
atgcgccatc tgagcgtgat tggcctgggc gcgatgggca gcgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtagtg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg tacgcggcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgtat gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 7
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 7
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Ser Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Ser Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Ser Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Val Ala Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Thr Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 8
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 8
atgcgccatc tgagcgtgat tggcctgggc gcgatgggca gcgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtagtg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gtggcggcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgacg gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 9
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 9
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Ser Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Ser Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Ser Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Val Ala Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 10
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 10
atgcgccatc tgagcgtgat tggcctgggc gcgatgggca gcgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtagtg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gtggcggcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggc gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 11
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 11
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Ser Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Ser Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Ser Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Asn Ala Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Tyr Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 12
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 12
atgcgccatc tgagcgtgat tggcctgggc gcgatgggca gcgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtagtg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg aacgcggcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgtat gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 13
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 13
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Ser Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Ser Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Ser Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Val Phe Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Tyr Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 14
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 14
atgcgccatc tgagcgtgat tggcctgggc gcgatgggca gcgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtagtg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gtgtttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgtat gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 15
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 15
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Ser Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Ser Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Val Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Cys Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Val Ala Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Phe Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Tyr Ala Ser Leu Trp Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 16
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 16
atgcgccatc tgagcgtgat tggcctgggc gcgatgggca gcgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtagtg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tgtttgcttt 360
ccggcgcaga ttggcaccag cgatgcgtgt attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gtggcggcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgtttctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgtat gcgagcctgt ggacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 17
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 17
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Val Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Ser Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Ser Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Asn Ala Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Tyr Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 18
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 18
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcg tcgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtagtg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg aatgcggcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgtat gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 19
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 19
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Val Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Ser Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Ser Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Val Ala Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Thr Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Tyr Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 20
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 20
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcg tcgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtagtg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gtggcggcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgactctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgtat gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 21
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 21
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Ser Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Arg Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Ser Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Asn Ala Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Tyr Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Leu Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 22
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 22
atgcgccatc tgagcgtgat tggcctgggc gcgatgggca gcgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtagtg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg aacgcggcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgtat gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 23
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 23
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Ser Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Ser Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Ser Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ser Ala Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Tyr Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 24
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 24
atgcgccatc tgagcgtgat tggcctgggc gcgatgggca gcgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtagtg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg agcgcggcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgtat gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 25
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 25
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Ser Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Ser Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Ser Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Ala Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Tyr Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 26
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 26
atgcgccatc tgagcgtgat tggcctgggc gcgatgggca gcgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtagtg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggcggcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgtat gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 27
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 27
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Ser Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Ser Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Ser Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Val Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Tyr Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 28
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 28
atgcgccatc tgagcgtgat tggcctgggc gcgatgggca gcgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtagtg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gtggtggcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgtat gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 29
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 29
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Ser Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Ser Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Ser Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Ala Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Tyr Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Lys Leu Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 30
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 30
atgcgccatc tgagcgtgat tggcctgggc gcgatgggca gcgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtagtg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggcggcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaatatag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctga agctttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 31
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 31
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Ser Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Ser Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Ser Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Tyr Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 32
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 32
atgcgccatc tgagcgtgat tggcctgggc gcgatgggca gcgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtagtg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggtggcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgtat gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 33
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 33
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Ser Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Ser Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Ser Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Ala Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 34
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 34
atgcgccatc tgagcgtgat tggcctgggc gcgatgggca gcgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtagtg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggcggcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggc gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 35
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 35
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Ser Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Ser Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Ser Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Val Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 36
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 36
atgcgccatc tgagcgtgat tggcctgggc gcgatgggca gcgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtagtg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gtggtggcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggc gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 37
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 37
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Ser Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Ser Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Ser Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 38
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 38
atgcgccatc tgagcgtgat tggcctgggc gcgatgggca gcgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtagtg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 39
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 39
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Leu Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Gly Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Ser Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Val Phe Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Leu Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Tyr Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 40
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 40
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcc tggcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtgggg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gtgtttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcttttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgtat gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 41
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 41
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Gln Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Ser Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Ser Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 42
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 42
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcc aggcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtagtg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 43
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 43
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Ser Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Asp Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Ser Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Leu Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 44
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 44
atgcgccatc tgagcgtgat tggcctgggc gcgatgggca gcgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtgatg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctgc tgacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 45
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 45
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Val Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Asp Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Thr Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Ser Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Phe Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Ala Ala Ser Leu Lys Thr Trp Ser Ala Tyr Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 46
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 46
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcg ttgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtgatg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgactttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcgtttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccggct gcgagcctga agacttggag cgcgtatatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 47
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 47
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Val Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Ser Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Thr Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Ser Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Phe Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Lys Leu Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 48
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 48
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcg ttgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtagtg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgactttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcgtttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctga agctttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 49
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 49
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Ser Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Gly Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Thr Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 50
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 50
atgcgccatc tgagcgtgat tggcctgggc gcgatgggca gcgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtgggg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgact attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 51
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 51
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Gln Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Ser Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Val Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Ser Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Leu Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Ala Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 52
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 52
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcc aggcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtagtg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tgtttgcttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcttttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccggct gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 53
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 53
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Gln Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Leu Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Val Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Ser Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Leu Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Leu Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 54
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 54
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcc aggcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtcttg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tgtttgcttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcttttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgctg gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 55
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 55
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Gln Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Gly Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Asp Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Thr Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Phe Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Ala Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 56
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 56
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcc aggcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtgggg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctggatttt 360
ccggcgcaga ttggcaccag cgatgcgact attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcgtttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcggctttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 57
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 57
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Gln Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Ala Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Ser Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Phe Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Lys Thr Trp Ser Ala Leu Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 58
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 58
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcc aggcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtgcgg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaatttag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctga agacttggag cgcgctgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 59
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 59
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Gln Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Ala Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Val Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Ser Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 60
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 60
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcc aggcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtgcgg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tgtttgcttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 61
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 61
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Val Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Gly Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Thr Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Leu Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 62
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 62
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcg ttgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtgggg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgact attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgctgctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 63
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 63
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Ser Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Gly Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Val Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Thr Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Leu Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 64
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 64
atgcgccatc tgagcgtgat tggcctgggc gcgatgggca gcgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtgggg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tgtttgcttt 360
ccggcgcaga ttggcaccag cgatgcgact attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctgc tgacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 65
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 65
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Ser Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Gly Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Asp Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Thr Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Ala Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Lys Thr Trp Ser Ala Tyr Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 66
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 66
atgcgccatc tgagcgtgat tggcctgggc gcgatgggca gcgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtgggg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctggatttt 360
ccggcgcaga ttggcaccag cgatgcgact attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcggcgttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctga agacttggag cgcgtatatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 67
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 67
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Gln Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Asp Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Thr Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Leu Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 68
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 68
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcc aggcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtgatg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgact attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctgc tgacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 69
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 69
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Val Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Gly Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Thr Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Thr Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Phe Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Lys Leu Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 70
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 70
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcg ttgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtgggg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgactttt 360
ccggcgcaga ttggcaccag cgatgcgact attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcgtttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctga agctgtggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 71
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 71
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Val Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Asp Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Thr Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Cys Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Phe Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Lys Thr Trp Ser Ala Tyr Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 72
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 72
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcg ttgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtgatg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgactttt 360
ccggcgcaga ttggcaccag cgatgcgtgt attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcgtttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctga agacttggag cgcgtatatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 73
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 73
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Gln Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Gln Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Val Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Thr Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Tyr Ser Pro
195 200 205
Leu Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 74
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 74
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcc aggcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtcagg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tgtttgcttt 360
ccggcgcaga ttggcaccag cgatgcgact attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaatatag cccgctgctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 75
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 75
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Gln Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Gly Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Val Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Cys Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Leu Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Thr Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 76
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 76
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcc aggcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtgggg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tgtttgcttt 360
ccggcgcaga ttggcaccag cgatgcgtgt attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgctgttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgact gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 77
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 77
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Ser Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Gly Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Asp Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Thr Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Ala Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Lys Thr Trp Ser Ala Leu Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 78
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 78
atgcgccatc tgagcgtgat tggcctgggc gcgatgggca gcgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtgggg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctggatttt 360
ccggcgcaga ttggcaccag cgatgcgact attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcggctttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctga agacttggag cgcgctgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 79
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 79
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Gln Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Ala Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Cys Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Ser Phe Ser Glu Phe Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Leu Ala Ser Leu Lys Thr Trp Ser Ala Leu Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 80
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 80
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcc aggcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtgcgg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgtgt attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgagcttta gcgaatttag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgctg gcgagcctga agacttggag cgcgctgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 81
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 81
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Gln Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Gly Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Asp Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Leu Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Phe Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 82
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 82
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcc aggcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtgggg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctggatttt 360
ccggcgcaga ttggcaccag cgatgcgctg attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctgt ttacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 83
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 83
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Val Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Ala Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Cys Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Leu Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Leu Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 84
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 84
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcg ttgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtgcgg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgtgt attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgctgctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaact gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 85
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 85
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Ser Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Gly Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Val Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Leu Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Ala Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 86
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 86
atgcgccatc tgagcgtgat tggcctgggc gcgatgggca gcgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtggcg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tgtttgcttt 360
ccggcgcaga ttggcaccag cgatgcgctg attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcggcgttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 87
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 87
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Ser Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Gly Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Val Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Cys Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Ala Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Lys Ser Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 88
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 88
atgcgccatc tgagcgtgat tggcctgggc gcgatgggca gcgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtggcg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tgtatgcttt 360
ccggcgcaga ttggcaccag cgatgcgtgt attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcggcgttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctga agagctggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 89
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 89
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Gln Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Asp Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Val Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Leu Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Leu Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 90
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 90
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcc aggcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtgatg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tgtttgcttt 360
ccggcgcaga ttggcaccag cgatgcgctg attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctgc tgacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 91
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 91
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Val Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Gly Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Asp Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Leu Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Lys Leu Trp Ser Ala Leu Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 92
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 92
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcg ttgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtggcg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctggatttt 360
ccggcgcaga ttggcaccag cgatgcgctg attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctga agctgtggag cgcgctgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 93
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 93
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Val Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Ser Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Met Thr Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Leu Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Lys Thr Trp Ser Ala Tyr Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 94
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 94
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcg ttgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtagtg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tatgaccttt 360
ccggcgcaga ttggcaccag cgatgcgctg attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctga agacttggag cgcgtatatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 95
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 95
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Gln Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Asn Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Val Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Leu Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Phe Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Tyr Ser Pro
195 200 205
Thr Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Leu Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 96
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 96
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcc aggcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtaatg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tgtgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgctg attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcgtttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaatatag cccgaccctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctgc tgacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 97
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 97
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Gln Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Gly Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Val Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Leu Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Leu Phe Ser Glu Leu Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Val Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 98
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 98
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcc aggcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtgggg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tgtgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgctg attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgttgttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctgg tgacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 99
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 99
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Leu Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Gly Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Asp Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Cys Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Ala Phe Ser Glu Leu Ser Pro
195 200 205
Thr Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Lys Thr Trp Ser Ala Ala Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Leu Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 100
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 100
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcc tggcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtgggg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctggatttt 360
ccggcgcaga ttggcaccag cgatgcgtgt attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcggcgttta gcgaactgag cccgacactg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaact gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 101
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 101
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Gln Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Gly Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Cys Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Phe Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Leu Ala Ser Leu Val Thr Trp Ser Ala Leu Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 102
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 102
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcc aggcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtgggg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgtgc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaatttag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgctg gcgagcctgg ttacttggag cgcgctgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 103
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 103
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Gln Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Gln Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Leu Asp Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Thr Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Tyr Phe Ser Glu Tyr Ser Pro
195 200 205
Ile Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Phe Thr Trp Ser Ala Leu Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 104
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 104
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcc aggcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtcagg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctggatttt 360
ccggcgcaga ttggcaccag cgatgcgact attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgtatttta gcgaatatag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctgt ttacttggag cgcgctgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 105
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 105
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Val Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Gly Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Val Cys Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Thr Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Gln Phe Ser Glu Leu Ser Pro
195 200 205
Thr Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Val Leu Trp Ser Ala Leu Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 106
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 106
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcg ttgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtgggg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tgtttgcttt 360
ccggcgcaga ttggcaccag cgatgcgact attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgactctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctgg ttctttggag cgcgctgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 107
<211> 292
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 107
Met Arg His Leu Ser Val Ile Gly Leu Gly Ala Met Gly Gln Ala Leu
1 5 10 15
Ala Thr Thr Leu Leu Lys Ala Gly His Pro Val Thr Val Trp Asn Arg
20 25 30
Gly Ala Ala Lys Ala Ala Pro Leu Gln Ala Leu Gly Ala Thr Leu Ala
35 40 45
Pro Ser Val Gly Ala Ala Ile Ala Ala Ser Asp Ile Thr Leu Val Cys
50 55 60
Val Asp Asn Tyr Ala Val Ser Gln Leu Leu Leu Asp Glu Ala Ser Asp
65 70 75 80
Ala Val Ala Gly Lys Leu Leu Val Gln Leu Ser Thr Gly Ser Pro Gln
85 90 95
Gly Ala Arg Ala Leu Glu Ser Trp Ser His Ala Arg Gly Ala Arg Tyr
100 105 110
Leu Asp Gly Ala Ile Val Thr Phe Pro Ala Gln Ile Gly Thr Ser Asp
115 120 125
Ala Cys Ile Ile Cys Ser Gly Ala Ser Ala Ala Phe Ser Glu Ala Glu
130 135 140
Pro Val Leu Ser Leu Leu Ala Pro Thr Leu Asp His Val Ala Glu Ala
145 150 155 160
Val Gly Ala Ala Ala Ala Gln Asp Cys Ala Ala Val Ala Tyr Phe Ala
165 170 175
Gly Gly Leu Leu Gly Ala Leu His Gly Ala Leu Ile Cys Glu Ala Glu
180 185 190
Gly Leu Pro Val Ala Lys Val Cys Ala Ala Phe Ser Glu Leu Ser Pro
195 200 205
Phe Leu Gly Gly Asp Val Ala His Leu Gly Lys Thr Leu Ala Ser Gly
210 215 220
Asp Phe Asp His Pro Gly Ala Ser Leu Lys Leu Trp Ser Ala Leu Ile
225 230 235 240
Ser Arg Leu Ala Gly His Ala Thr Asp Ala Gly Ile Asp Ser Arg Phe
245 250 255
Pro Arg Phe Ala Ala Asp Leu Phe Glu Glu Gly Val Ala Gln Gly Phe
260 265 270
Gly Gln Gln Glu Val Ser Ala Leu Ile Lys Val Leu Arg Ala Arg Asn
275 280 285
Gly Ala Ala Gln
290
<210> 108
<211> 879
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 108
atgcgccatc tgagcgtgat tggcctgggc gcgatgggcc aggcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtgggg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tgttactttt 360
ccggcgcaga ttggcaccag cgatgcgtgc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gcggttgcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcggcgttta gcgaactgag cccgtttctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgggg gcgagcctga agctttggag cgcgctgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaa 879
<210> 109
<211> 30
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 109
tatacatatg cgccatctga gcgtgattgg 30
<210> 110
<211> 29
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 110
ttcggatcct tactgcgccg cgccgttgc 29
<210> 111
<211> 35
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 111
gccggatcca ataattttgt ttaactttaa gaagg 35
<210> 112
<211> 31
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 112
ggctcgagtt agccacgacc cgcttgaaag c 31
<210> 113
<211> 1707
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 113
atgcgccatc tgagcgtgat tggcctgggc gcgatgggca gcgcgctggc gaccaccctg 60
ctgaaggcag gacacccagt aacagtttgg aatcgtagtg ctgcaaaggc tgctccactc 120
caagctctag gtgctacttt ggctcctagt gttggagccg ctatcgccgc gagcgatatt 180
accctggtgt gcgtggataa ctatgcggtg agccagctgc tgctggatga agcgagcgat 240
gcagtagccg ggaagttgct tgtacaactt tcgacgggaa gtcctcaagg tgcacgtgcc 300
cttgagagct ggagccatgc gcgcggcgcg cgctatctgg atggcgcgat tctgtgcttt 360
ccggcgcaga ttggcaccag cgatgcgagc attatttgca gcggcgcgag cgcggcgttt 420
agcgaagcgg aaccggtgct gtctctcctc gctcccacac tagaccacgt tgccgaggct 480
gtgggagcgg ccgcggctca agactgcgcg gtggcggcgt atttcgctgg aggcctgctg 540
ggcgcgctgc atggcgcgct gatttgcgaa gcggaaggcc tgccggtggc gaaagtgtgc 600
gcgcagttta gcgaactgag cccgattctg ggcggcgatg tggcgcatct gggcaagaca 660
ttggcgagcg gcgatttcga ccacccgtat gcgagcctga agacttggag cgcggcgatt 720
agccgcctgg cgggccatgc gaccgatgcg ggcattgata gccgctttcc gcgctttgcg 780
gcggatctgt ttgaagaagg cgtggcgcag ggctttggcc agcaggaagt gagcgcgctg 840
attaaagtgc tgcgcgcgcg caacggcgcg gcgcagtaag gatccaataa ttttgtttaa 900
ctttaagaag gagatataca tatgtacaag gacctggaag gcaaagtggt tgtgatcacc 960
ggcagcagca ccggtctggg caagagcatg gcgattcgtt tcgcgaccga gaaggcgaaa 1020
gttgtggtta actatcgtag caaagaggat gaagcgaaca gcgtgctgga ggaaatcaag 1080
aaagttggtg gcgaagcgat tgcggtgaag ggtgacgtga ccgttgagag cgatgtgatc 1140
aacctggttc agagcgcgat taaggaattt ggtaaactgg acgttatgat caacaacgcg 1200
ggcctggaaa acccggtgag cagccacgag atgagcctga gcgactggaa caaggttatc 1260
gataccaacc tgaccggtgc gttcctgggc agccgtgaag cgattaagta ctttgtggag 1320
aacgatatca aaggtaccgt gattaacatg agcagcgttc acgagaaaat cccgtggccg 1380
ctgttcgttc actacgcggc gagcaagggt ggcatgaaac tgatgaccga gaccctggcg 1440
ctggaatatg cgccgaaggg catccgtgtt aacaacattg gtccgggcgc gatcaacacc 1500
ccgattaacg cggagaaatt tgcggacccg gaacaacgtg cggatgtgga gagcatgatc 1560
ccgatgggtt atattggcga accggaagaa attgcggcgg ttgcggcgtg gctggcgagc 1620
agcgaggcga gctacgtgac cggtattacc ctgttcgcgg acggtggcat gacccagtat 1680
ccgagctttc aagcgggtcg tggctaa 1707
Claims (17)
1.亚胺还原酶突变体,其特征在于:与氨基酸序列SEQ ID NO:1相比,存在残基差异包括V171、A172、Y230中的一个或多个。
2.根据权利要求1所述的亚胺还原酶突变体,其特征在于:所述残基差异包括V171Y/N/A/S、A172V/F、Y230G/A/T中的一个或多个。
3.根据权利要求1或2所述的亚胺还原酶突变体,其特征在于:亚胺还原酶突变体的氨基酸序列选自SEQ ID NO:3、5、7、9、11、13、15、17、19、21、23、25、27、29、31、33、35、37、39、41、43、45、47、49、51、53、55、57、59、61、63、65、67、69、71、73、75、77、79、81、83、85、87、89、91、93、95、97、99、101、105或107。
5.根据权利要求4所述的制备(S)-尼古丁的方法,其特征在于:将底物I、辅酶、葡萄糖、葡萄糖脱氢酶、缓冲液与亚胺还原酶混合后,反应得到化合物III(S)-去甲烟碱,然后将化合物III(S)-去甲烟碱甲基化得到(S)-尼古丁。
7.根据权利要求6所述的制备(S)-尼古丁的方法,其特征在于:底物II环化后经亚胺还原酶的催化得到(S)-尼古丁;
或底物II的盐经脱盐、环化经亚胺还原酶的催化得到(S)-尼古丁;
所述底物II的盐包括盐酸盐、二盐酸盐、氢溴酸盐、二氢溴酸盐、硫酸盐或硫酸氢盐。
8.一种核酸,其特征在于:所述核酸可以编码权利要求1至3中任一项所述的亚胺还原酶突变体。
9.一种表达载体,其特征在于:包括权利要求8的核酸。
10.一种细胞,其特征在于:包括权利要求8的核酸或权利要求9的载体。
11.一种如权利要求1至3中任意一项的亚胺还原酶突变体的生产方法,其特征在于培养权利要求1至3中所述的细胞得到所述的亚胺还原酶突变体。
12.如权利要求11所述的生产方法,其特征在于所述的亚胺还原酶突变体和葡萄糖脱氢酶共表达。
13.如权利要求12所述的生产方法,其特征在于:用同时含有亚胺还原酶突变体基因与葡萄糖脱氢酶基因的细胞生产共表达酶。
14.如权利要求13所述的生产方法,其特征在于:通过带有限制性内切酶位点的引物将葡萄糖脱氢酶基因从载体上扩增下来,分别用限制性内切酶BamHI和XhoI对带有亚胺还原酶基因片段的载体和扩增的葡萄糖脱氢酶基因片段进行双酶切,胶回收酶切片段,并用T4DNA连接酶连接后转化大肠杆菌BL21感受态细胞,获得同时含有亚胺还原酶基因和葡萄糖脱氢酶基因的重组菌,将上述重组菌培养,然后诱导、离心收集菌体、重悬,重悬后对细胞进行超声破碎,再将破碎后溶液进行冷冻干燥,获得亚胺还原酶和葡萄糖脱氢酶共表达酶粉。
15.通过权利要求11-14任意一项所述的生产方法制备得到的亚胺还原酶突变体或者亚胺还原酶突变体与葡萄糖脱氢酶的共表达酶。
Priority Applications (4)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
EP22827500.4A EP4361260A1 (en) | 2021-06-20 | 2022-06-20 | Imine reductase mutant, co-expressed enzyme of imine reductase and glucose dehydrogenase, and application thereof |
KR1020237044009A KR20240011764A (ko) | 2021-06-20 | 2022-06-20 | 이민 환원 효소 돌연변이체, 이민 환원 효소와 포도당 탈수소효소의 공동발현 효소 및 이의 응용 |
PCT/CN2022/099711 WO2022268006A1 (zh) | 2021-06-20 | 2022-06-20 | 亚胺还原酶突变体、亚胺还原酶和葡萄糖脱氢酶共表达酶及其应用 |
US18/389,868 US20240141306A1 (en) | 2021-06-20 | 2023-12-20 | Imine reductase mutant, co-expressed recombinant enzyme of imine reductase and glucose dehydrogenase, and applications thereof |
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN202110682123 | 2021-06-20 | ||
CN2021106821238 | 2021-06-20 |
Publications (1)
Publication Number | Publication Date |
---|---|
CN115491364A true CN115491364A (zh) | 2022-12-20 |
Family
ID=84464625
Family Applications (2)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CN202110684461.5A Pending CN115491364A (zh) | 2021-06-20 | 2021-06-21 | 亚胺还原酶突变体、亚胺还原酶和葡萄糖脱氢酶共表达酶及其应用 |
CN202280035192.2A Pending CN117751184A (zh) | 2021-06-20 | 2022-06-20 | 亚胺还原酶突变体、亚胺还原酶和葡萄糖脱氢酶共表达酶及其应用 |
Family Applications After (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CN202280035192.2A Pending CN117751184A (zh) | 2021-06-20 | 2022-06-20 | 亚胺还原酶突变体、亚胺还原酶和葡萄糖脱氢酶共表达酶及其应用 |
Country Status (5)
Country | Link |
---|---|
US (1) | US20240141306A1 (zh) |
EP (1) | EP4361260A1 (zh) |
KR (1) | KR20240011764A (zh) |
CN (2) | CN115491364A (zh) |
WO (1) | WO2022268006A1 (zh) |
Families Citing this family (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN117363667B (zh) * | 2023-12-07 | 2024-03-22 | 欣雅利华生物技术(上海)有限公司 | 亚胺还原酶在制备达泊西汀中间体和/或达泊西汀中的用途 |
Family Cites Families (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
IL225900A0 (en) | 2013-04-22 | 2014-03-31 | Perrigo Api Ltd | A process for the preparation of nicotine that includes the enzymatic reduction of 4-(methylamino)-1-(3-pyridinyl)-1-butanone |
HUE051799T2 (hu) | 2018-11-16 | 2021-03-29 | Zanoprima Lifesciences Ltd | Eljárás (S)-nikotin elõállítására myosmine vegyületbõl |
CN110564788B (zh) * | 2019-09-30 | 2021-05-04 | 江南大学 | 一种利用亚胺还原酶生产麻黄碱的方法 |
-
2021
- 2021-06-21 CN CN202110684461.5A patent/CN115491364A/zh active Pending
-
2022
- 2022-06-20 WO PCT/CN2022/099711 patent/WO2022268006A1/zh active Application Filing
- 2022-06-20 EP EP22827500.4A patent/EP4361260A1/en active Pending
- 2022-06-20 CN CN202280035192.2A patent/CN117751184A/zh active Pending
- 2022-06-20 KR KR1020237044009A patent/KR20240011764A/ko unknown
-
2023
- 2023-12-20 US US18/389,868 patent/US20240141306A1/en active Pending
Also Published As
Publication number | Publication date |
---|---|
CN117751184A (zh) | 2024-03-22 |
KR20240011764A (ko) | 2024-01-26 |
US20240141306A1 (en) | 2024-05-02 |
WO2022268006A1 (zh) | 2022-12-29 |
EP4361260A1 (en) | 2024-05-01 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
US10837036B2 (en) | Method for preparing L-aspartic acid with maleic acid by whole-cell biocatalysis | |
WO2018023209A1 (zh) | 一种制备烟酰胺单核苷酸的方法 | |
WO2018023207A1 (zh) | 一种制备烟酰胺单核苷酸的方法 | |
CN109852644B (zh) | 一种制备布瓦西坦中间体的方法 | |
JP7491588B2 (ja) | 操作されたパントテン酸キナーゼ改変体酵素 | |
CN113801859B (zh) | 一种用于制备手性醇化合物的羰基还原酶突变体及其用途 | |
US20240141306A1 (en) | Imine reductase mutant, co-expressed recombinant enzyme of imine reductase and glucose dehydrogenase, and applications thereof | |
CN115927224A (zh) | 一种羰基还原酶突变体及其应用 | |
CN115992189A (zh) | 一种细菌色氨酸-5-羟化酶及其应用 | |
JP2005198655A (ja) | セファロスポリンcアシラーゼ | |
CN110396507B (zh) | 源自Cnuibacter physcomitrellae的L-泛解酸内酯脱氢酶 | |
WO2023015712A1 (zh) | 一种s-尼古丁的制备方法 | |
CN117535256A (zh) | 一种羰基还原酶及其在玻色因合成中应用 | |
EP3567105A1 (en) | Highly active s-cyanohydrin lyase and application thereof | |
CN110396506B (zh) | 源自Nocardia asteroides的L-泛解酸内酯脱氢酶及其应用 | |
CN115704014A (zh) | 亚胺还原酶突变体及其在合成(r)-2-(2,5-二氟苯基)吡咯烷中的应用 | |
CN113106078B (zh) | 一种亮氨酸脱氢酶突变体及其编码基因、基因工程菌以及在制备l-叔亮氨酸中的应用 | |
CN114774383B (zh) | 一类亚胺还原酶突变体及其在催化合成手性2-芳基吡咯烷中的应用 | |
CN114075557B (zh) | 重组转氨酶及其在合成(r)-2-(2,5-二氟苯基)吡咯烷中的应用 | |
JP6844073B1 (ja) | (1r,3r)−3−(トリフルオロメチル)シクロヘキサン−1−オール及びその中間体の製造法 | |
CN110527671B (zh) | 源自Nocardia farcinica的L-泛解酸内酯脱氢酶及其应用 | |
CN118006574A (zh) | 一种酶突变体、级联催化合成手性s-4-芳基噁唑烷酮的方法和应用 | |
CN115537405A (zh) | 一种酮还原酶及其在制备(s)-1-(3-氯苯基)-1,3-丙二醇中的应用 | |
CN117402920A (zh) | 酮还原酶BsSDR10突变体及其在不对称还原α-唑烷基取代的苯乙酮衍生物中的应用 | |
CN116710558A (zh) | 工程化磷酸戊糖变位酶变体酶 |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
PB01 | Publication | ||
PB01 | Publication | ||
TA01 | Transfer of patent application right | ||
TA01 | Transfer of patent application right |
Effective date of registration: 20231008 Address after: 401221 Chongqing Changshou District Chongqing (Changshou) Chemical Industry Park Fine Chemical Industry Zone 1 Applicant after: PORTON FINE CHEMICALS Ltd. Address before: 401120 15-3, Unit 2, Building 184, No. 55, Cuiyu Road, Yubei District, Chongqing Applicant before: Chen Zecong |
|
SE01 | Entry into force of request for substantive examination | ||
SE01 | Entry into force of request for substantive examination |