CN106434586A - Trehalose synthase mutant and gene thereof - Google Patents
Trehalose synthase mutant and gene thereof Download PDFInfo
- Publication number
- CN106434586A CN106434586A CN201610876031.2A CN201610876031A CN106434586A CN 106434586 A CN106434586 A CN 106434586A CN 201610876031 A CN201610876031 A CN 201610876031A CN 106434586 A CN106434586 A CN 106434586A
- Authority
- CN
- China
- Prior art keywords
- leu
- asp
- arg
- pro
- ala
- 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.)
- Granted
Links
- 108090000623 proteins and genes Proteins 0.000 title claims abstract description 21
- 108010045348 trehalose synthase Proteins 0.000 title abstract description 4
- 102000004190 Enzymes Human genes 0.000 claims abstract description 52
- 108090000790 Enzymes Proteins 0.000 claims abstract description 52
- HDTRYLNUVZCQOY-UHFFFAOYSA-N α-D-glucopyranosyl-α-D-glucopyranoside Natural products OC1C(O)C(O)C(CO)OC1OC1C(O)C(O)C(O)C(CO)O1 HDTRYLNUVZCQOY-UHFFFAOYSA-N 0.000 claims description 50
- HDTRYLNUVZCQOY-WSWWMNSNSA-N Trehalose Natural products O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@@H](O)[C@@H](CO)O1 HDTRYLNUVZCQOY-WSWWMNSNSA-N 0.000 claims description 50
- 108090000364 Ligases Proteins 0.000 claims description 33
- 102000003960 Ligases Human genes 0.000 claims description 32
- HDTRYLNUVZCQOY-LIZSDCNHSA-N alpha,alpha-trehalose Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 HDTRYLNUVZCQOY-LIZSDCNHSA-N 0.000 claims description 21
- 239000013598 vector Substances 0.000 claims description 7
- 230000009261 transgenic effect Effects 0.000 claims description 6
- 241000186359 Mycobacterium Species 0.000 claims description 4
- 239000000203 mixture Substances 0.000 claims description 4
- 230000002194 synthesizing effect Effects 0.000 claims description 4
- 239000002773 nucleotide Substances 0.000 claims description 2
- 125000003729 nucleotide group Chemical group 0.000 claims description 2
- 125000003275 alpha amino acid group Chemical group 0.000 claims 1
- 230000000694 effects Effects 0.000 abstract description 23
- 230000035772 mutation Effects 0.000 abstract description 18
- 150000001413 amino acids Chemical class 0.000 abstract description 10
- 235000001014 amino acid Nutrition 0.000 abstract description 9
- QNAYBMKLOCPYGJ-REOHCLBHSA-N L-alanine Chemical compound C[C@H](N)C(O)=O QNAYBMKLOCPYGJ-REOHCLBHSA-N 0.000 abstract description 8
- 235000004279 alanine Nutrition 0.000 abstract description 8
- 241000187480 Mycobacterium smegmatis Species 0.000 abstract 1
- 229940088598 enzyme Drugs 0.000 description 45
- 108010057821 leucylproline Proteins 0.000 description 40
- 108010044940 alanylglutamine Proteins 0.000 description 30
- FSXRLASFHBWESK-UHFFFAOYSA-N dipeptide phenylalanyl-tyrosine Natural products C=1C=C(O)C=CC=1CC(C(O)=O)NC(=O)C(N)CC1=CC=CC=C1 FSXRLASFHBWESK-UHFFFAOYSA-N 0.000 description 30
- XBGGUPMXALFZOT-UHFFFAOYSA-N glycyl-L-tyrosine hemihydrate Natural products NCC(=O)NC(C(O)=O)CC1=CC=C(O)C=C1 XBGGUPMXALFZOT-UHFFFAOYSA-N 0.000 description 30
- NONWUQAWAANERO-BZSNNMDCSA-N Asp-Phe-Tyr Chemical compound C([C@H](NC(=O)[C@H](CC(O)=O)N)C(=O)N[C@@H](CC=1C=CC(O)=CC=1)C(O)=O)C1=CC=CC=C1 NONWUQAWAANERO-BZSNNMDCSA-N 0.000 description 20
- 108010077245 asparaginyl-proline Proteins 0.000 description 20
- 108010079547 glutamylmethionine Proteins 0.000 description 20
- 108010087823 glycyltyrosine Proteins 0.000 description 20
- 238000006243 chemical reaction Methods 0.000 description 17
- 241000282326 Felis catus Species 0.000 description 15
- 241000894006 Bacteria Species 0.000 description 11
- 241000588724 Escherichia coli Species 0.000 description 11
- INOZZBHURUDQQR-AJNGGQMLSA-N (2s)-2-[[(2s)-2-[[(2s)-2-[[(2s)-2-amino-3-(1h-imidazol-5-yl)propanoyl]amino]-3-carboxypropanoyl]amino]-4-carboxybutanoyl]amino]-4-methylpentanoic acid Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](CC(O)=O)NC(=O)[C@@H](N)CC1=CN=CN1 INOZZBHURUDQQR-AJNGGQMLSA-N 0.000 description 10
- JAMAWBXXKFGFGX-KZVJFYERSA-N Ala-Arg-Thr Chemical compound [H]N[C@@H](C)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H]([C@@H](C)O)C(O)=O JAMAWBXXKFGFGX-KZVJFYERSA-N 0.000 description 10
- ZEXDYVGDZJBRMO-ACZMJKKPSA-N Ala-Asn-Gln Chemical compound C[C@@H](C(=O)N[C@@H](CC(=O)N)C(=O)N[C@@H](CCC(=O)N)C(=O)O)N ZEXDYVGDZJBRMO-ACZMJKKPSA-N 0.000 description 10
- BLGHHPHXVJWCNK-GUBZILKMSA-N Ala-Gln-Leu Chemical compound [H]N[C@@H](C)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CC(C)C)C(O)=O BLGHHPHXVJWCNK-GUBZILKMSA-N 0.000 description 10
- KXEVYGKATAMXJJ-ACZMJKKPSA-N Ala-Glu-Asp Chemical compound C[C@H](N)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(O)=O)C(O)=O KXEVYGKATAMXJJ-ACZMJKKPSA-N 0.000 description 10
- ROLXPVQSRCPVGK-XDTLVQLUSA-N Ala-Glu-Tyr Chemical compound N[C@@H](C)C(=O)N[C@@H](CCC(=O)O)C(=O)N[C@@H](CC1=CC=C(C=C1)O)C(=O)O ROLXPVQSRCPVGK-XDTLVQLUSA-N 0.000 description 10
- QHASENCZLDHBGX-ONGXEEELSA-N Ala-Gly-Phe Chemical compound C[C@H](N)C(=O)NCC(=O)N[C@H](C(O)=O)CC1=CC=CC=C1 QHASENCZLDHBGX-ONGXEEELSA-N 0.000 description 10
- SDZRIBWEVVRDQI-CIUDSAMLSA-N Ala-Lys-Asp Chemical compound [H]N[C@@H](C)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC(O)=O)C(O)=O SDZRIBWEVVRDQI-CIUDSAMLSA-N 0.000 description 10
- DHONNEYAZPNGSG-UBHSHLNASA-N Ala-Val-Phe Chemical compound C[C@H](N)C(=O)N[C@@H](C(C)C)C(=O)N[C@H](C(O)=O)CC1=CC=CC=C1 DHONNEYAZPNGSG-UBHSHLNASA-N 0.000 description 10
- NLYYHIKRBRMAJV-AEJSXWLSSA-N Ala-Val-Pro Chemical compound C[C@@H](C(=O)N[C@@H](C(C)C)C(=O)N1CCC[C@@H]1C(=O)O)N NLYYHIKRBRMAJV-AEJSXWLSSA-N 0.000 description 10
- UXJCMQFPDWCHKX-DCAQKATOSA-N Arg-Arg-Glu Chemical compound NC(N)=NCCC[C@H](N)C(=O)N[C@@H](CCCN=C(N)N)C(=O)N[C@@H](CCC(O)=O)C(O)=O UXJCMQFPDWCHKX-DCAQKATOSA-N 0.000 description 10
- CPSHGRGUPZBMOK-CIUDSAMLSA-N Arg-Asn-Gln Chemical compound [H]N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CCC(N)=O)C(O)=O CPSHGRGUPZBMOK-CIUDSAMLSA-N 0.000 description 10
- HPSVTWMFWCHKFN-GARJFASQSA-N Arg-Glu-Pro Chemical compound C1C[C@@H](N(C1)C(=O)[C@H](CCC(=O)O)NC(=O)[C@H](CCCN=C(N)N)N)C(=O)O HPSVTWMFWCHKFN-GARJFASQSA-N 0.000 description 10
- OQCWXQJLCDPRHV-UWVGGRQHSA-N Arg-Gly-Leu Chemical compound [H]N[C@@H](CCCNC(N)=N)C(=O)NCC(=O)N[C@@H](CC(C)C)C(O)=O OQCWXQJLCDPRHV-UWVGGRQHSA-N 0.000 description 10
- QEHMMRSQJMOYNO-DCAQKATOSA-N Arg-His-Asn Chemical compound C1=C(NC=N1)C[C@@H](C(=O)N[C@@H](CC(=O)N)C(=O)O)NC(=O)[C@H](CCCN=C(N)N)N QEHMMRSQJMOYNO-DCAQKATOSA-N 0.000 description 10
- OFIYLHVAAJYRBC-HJWJTTGWSA-N Arg-Ile-Phe Chemical compound CC[C@H](C)[C@H](NC(=O)[C@@H](N)CCCNC(N)=N)C(=O)N[C@@H](Cc1ccccc1)C(O)=O OFIYLHVAAJYRBC-HJWJTTGWSA-N 0.000 description 10
- YKZJPIPFKGYHKY-DCAQKATOSA-N Arg-Leu-Asp Chemical compound [H]N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC(O)=O)C(O)=O YKZJPIPFKGYHKY-DCAQKATOSA-N 0.000 description 10
- PZBSKYJGKNNYNK-ULQDDVLXSA-N Arg-Leu-Tyr Chemical compound CC(C)C[C@H](NC(=O)[C@@H](N)CCCN=C(N)N)C(=O)N[C@@H](Cc1ccc(O)cc1)C(O)=O PZBSKYJGKNNYNK-ULQDDVLXSA-N 0.000 description 10
- YVTHEZNOKSAWRW-DCAQKATOSA-N Arg-Lys-Ala Chemical compound [H]N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](C)C(O)=O YVTHEZNOKSAWRW-DCAQKATOSA-N 0.000 description 10
- MNBHKGYCLBUIBC-UFYCRDLUSA-N Arg-Phe-Phe Chemical compound C([C@H](NC(=O)[C@H](CCCNC(N)=N)N)C(=O)N[C@@H](CC=1C=CC=CC=1)C(O)=O)C1=CC=CC=C1 MNBHKGYCLBUIBC-UFYCRDLUSA-N 0.000 description 10
- PLTGTJAZQRGMPP-FXQIFTODSA-N Asn-Pro-Ala Chemical compound OC(=O)[C@H](C)NC(=O)[C@@H]1CCCN1C(=O)[C@@H](N)CC(N)=O PLTGTJAZQRGMPP-FXQIFTODSA-N 0.000 description 10
- SKQTXVZTCGSRJS-SRVKXCTJSA-N Asn-Tyr-Asp Chemical compound C1=CC(=CC=C1C[C@@H](C(=O)N[C@@H](CC(=O)O)C(=O)O)NC(=O)[C@H](CC(=O)N)N)O SKQTXVZTCGSRJS-SRVKXCTJSA-N 0.000 description 10
- 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 10
- QHAJMRDEWNAIBQ-FXQIFTODSA-N Asp-Arg-Asn Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC(N)=O)C(O)=O QHAJMRDEWNAIBQ-FXQIFTODSA-N 0.000 description 10
- ZLGKHJHFYSRUBH-FXQIFTODSA-N Asp-Arg-Asp Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC(O)=O)C(O)=O ZLGKHJHFYSRUBH-FXQIFTODSA-N 0.000 description 10
- SVFOIXMRMLROHO-SRVKXCTJSA-N Asp-Asp-Phe Chemical compound OC(=O)C[C@H](N)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@H](C(O)=O)CC1=CC=CC=C1 SVFOIXMRMLROHO-SRVKXCTJSA-N 0.000 description 10
- YBMUFUWSMIKJQA-GUBZILKMSA-N Asp-Gln-His Chemical compound C1=C(NC=N1)C[C@@H](C(=O)O)NC(=O)[C@H](CCC(=O)N)NC(=O)[C@H](CC(=O)O)N YBMUFUWSMIKJQA-GUBZILKMSA-N 0.000 description 10
- RATOMFTUDRYMKX-ACZMJKKPSA-N Asp-Glu-Cys Chemical compound C(CC(=O)O)[C@@H](C(=O)N[C@@H](CS)C(=O)O)NC(=O)[C@H](CC(=O)O)N RATOMFTUDRYMKX-ACZMJKKPSA-N 0.000 description 10
- OMMIEVATLAGRCK-BYPYZUCNSA-N Asp-Gly-Gly Chemical compound OC(=O)C[C@H](N)C(=O)NCC(=O)NCC(O)=O OMMIEVATLAGRCK-BYPYZUCNSA-N 0.000 description 10
- PSLSTUMPZILTAH-BYULHYEWSA-N Asp-Gly-Ile Chemical compound CC[C@H](C)[C@@H](C(O)=O)NC(=O)CNC(=O)[C@@H](N)CC(O)=O PSLSTUMPZILTAH-BYULHYEWSA-N 0.000 description 10
- KHGPWGKPYHPOIK-QWRGUYRKSA-N Asp-Gly-Phe Chemical compound [H]N[C@@H](CC(O)=O)C(=O)NCC(=O)N[C@@H](CC1=CC=CC=C1)C(O)=O KHGPWGKPYHPOIK-QWRGUYRKSA-N 0.000 description 10
- AYFVRYXNDHBECD-YUMQZZPRSA-N Asp-Leu-Gly Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](CC(C)C)C(=O)NCC(O)=O AYFVRYXNDHBECD-YUMQZZPRSA-N 0.000 description 10
- AHWRSSLYSGLBGD-CIUDSAMLSA-N Asp-Pro-Glu Chemical compound OC(=O)C[C@H](N)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CCC(O)=O)C(O)=O AHWRSSLYSGLBGD-CIUDSAMLSA-N 0.000 description 10
- DINOVZWPTMGSRF-QXEWZRGKSA-N Asp-Pro-Val Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N1CCC[C@H]1C(=O)N[C@@H](C(C)C)C(O)=O DINOVZWPTMGSRF-QXEWZRGKSA-N 0.000 description 10
- MGSVBZIBCCKGCY-ZLUOBGJFSA-N Asp-Ser-Ser Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](CO)C(=O)N[C@@H](CO)C(O)=O MGSVBZIBCCKGCY-ZLUOBGJFSA-N 0.000 description 10
- MNQMTYSEKZHIDF-GCJQMDKQSA-N Asp-Thr-Ala Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](C)C(O)=O MNQMTYSEKZHIDF-GCJQMDKQSA-N 0.000 description 10
- OTKUAVXGMREHRX-CFMVVWHZSA-N Asp-Tyr-Ile Chemical compound CC[C@H](C)[C@@H](C(O)=O)NC(=O)[C@@H](NC(=O)[C@@H](N)CC(O)=O)CC1=CC=C(O)C=C1 OTKUAVXGMREHRX-CFMVVWHZSA-N 0.000 description 10
- 108020004705 Codon Proteins 0.000 description 10
- XXDATQFUGMAJRV-XIRDDKMYSA-N Cys-Leu-Trp Chemical compound [H]N[C@@H](CS)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC1=CNC2=C1C=CC=C2)C(O)=O XXDATQFUGMAJRV-XIRDDKMYSA-N 0.000 description 10
- YTMBNLHIDIKJIU-HCXYKTFWSA-N D-Arginyl-L-arginyl-D-glutaminyl-L-phenylalanine Chemical compound NC(=N)NCCC[C@@H](N)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@H](CCC(O)=N)C(=O)N[C@H](C(O)=O)CC1=CC=CC=C1 YTMBNLHIDIKJIU-HCXYKTFWSA-N 0.000 description 10
- CYTSBCIIEHUPDU-ACZMJKKPSA-N Gln-Asp-Ala Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](C)C(O)=O CYTSBCIIEHUPDU-ACZMJKKPSA-N 0.000 description 10
- MCAVASRGVBVPMX-FXQIFTODSA-N Gln-Glu-Ala Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](C)C(O)=O MCAVASRGVBVPMX-FXQIFTODSA-N 0.000 description 10
- ZNZPKVQURDQFFS-FXQIFTODSA-N Gln-Glu-Ser Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CO)C(O)=O ZNZPKVQURDQFFS-FXQIFTODSA-N 0.000 description 10
- NSNUZSPSADIMJQ-WDSKDSINSA-N Gln-Gly-Asp Chemical compound NC(=O)CC[C@H](N)C(=O)NCC(=O)N[C@@H](CC(O)=O)C(O)=O NSNUZSPSADIMJQ-WDSKDSINSA-N 0.000 description 10
- VLOLPWWCNKWRNB-LOKLDPHHSA-N Gln-Thr-Pro Chemical compound C[C@H]([C@@H](C(=O)N1CCC[C@@H]1C(=O)O)NC(=O)[C@H](CCC(=O)N)N)O VLOLPWWCNKWRNB-LOKLDPHHSA-N 0.000 description 10
- RBSKVTZUFMIWFU-XEGUGMAKSA-N Gln-Trp-Ala Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)N[C@@H](CC1=CNC2=C1C=CC=C2)C(=O)N[C@@H](C)C(O)=O RBSKVTZUFMIWFU-XEGUGMAKSA-N 0.000 description 10
- RGNMNWULPAYDAH-JSGCOSHPSA-N Gln-Trp-Gly Chemical compound C1=CC=C2C(=C1)C(=CN2)C[C@@H](C(=O)NCC(=O)O)NC(=O)[C@H](CCC(=O)N)N RGNMNWULPAYDAH-JSGCOSHPSA-N 0.000 description 10
- CTJRFALAOYAJBX-NWLDYVSISA-N Gln-Trp-Thr Chemical compound C[C@H]([C@@H](C(=O)O)NC(=O)[C@H](CC1=CNC2=CC=CC=C21)NC(=O)[C@H](CCC(=O)N)N)O CTJRFALAOYAJBX-NWLDYVSISA-N 0.000 description 10
- DIXKFOPPGWKZLY-CIUDSAMLSA-N Glu-Arg-Asp Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC(O)=O)C(O)=O DIXKFOPPGWKZLY-CIUDSAMLSA-N 0.000 description 10
- CGYDXNKRIMJMLV-GUBZILKMSA-N Glu-Arg-Glu Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCC(O)=O)C(O)=O CGYDXNKRIMJMLV-GUBZILKMSA-N 0.000 description 10
- ZOXBSICWUDAOHX-GUBZILKMSA-N Glu-Asn-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@H](CC(N)=O)NC(=O)[C@@H](N)CCC(O)=O ZOXBSICWUDAOHX-GUBZILKMSA-N 0.000 description 10
- ATVYZJGOZLVXDK-IUCAKERBSA-N Glu-Leu-Gly Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(C)C)C(=O)NCC(O)=O ATVYZJGOZLVXDK-IUCAKERBSA-N 0.000 description 10
- ILWHFUZZCFYSKT-AVGNSLFASA-N Glu-Lys-Leu Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC(C)C)C(O)=O ILWHFUZZCFYSKT-AVGNSLFASA-N 0.000 description 10
- ALMBZBOCGSVSAI-ACZMJKKPSA-N Glu-Ser-Asn Chemical compound C(CC(=O)O)[C@@H](C(=O)N[C@@H](CO)C(=O)N[C@@H](CC(=O)N)C(=O)O)N ALMBZBOCGSVSAI-ACZMJKKPSA-N 0.000 description 10
- JLCYOCDGIUZMKQ-JBACZVJFSA-N Glu-Trp-Phe Chemical compound C1=CC=C(C=C1)C[C@@H](C(=O)O)NC(=O)[C@H](CC2=CNC3=CC=CC=C32)NC(=O)[C@H](CCC(=O)O)N JLCYOCDGIUZMKQ-JBACZVJFSA-N 0.000 description 10
- KXRORHJIRAOQPG-SOUVJXGZSA-N Glu-Tyr-Pro Chemical compound C1C[C@@H](N(C1)C(=O)[C@H](CC2=CC=C(C=C2)O)NC(=O)[C@H](CCC(=O)O)N)C(=O)O KXRORHJIRAOQPG-SOUVJXGZSA-N 0.000 description 10
- WKJKBELXHCTHIJ-WPRPVWTQSA-N Gly-Arg-Val Chemical compound CC(C)[C@@H](C(O)=O)NC(=O)[C@@H](NC(=O)CN)CCCN=C(N)N WKJKBELXHCTHIJ-WPRPVWTQSA-N 0.000 description 10
- QSTLUOIOYLYLLF-WDSKDSINSA-N Gly-Asp-Glu Chemical compound [H]NCC(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CCC(O)=O)C(O)=O QSTLUOIOYLYLLF-WDSKDSINSA-N 0.000 description 10
- XBWMTPAIUQIWKA-BYULHYEWSA-N Gly-Asp-Ile Chemical compound CC[C@H](C)[C@@H](C(O)=O)NC(=O)[C@H](CC(O)=O)NC(=O)CN XBWMTPAIUQIWKA-BYULHYEWSA-N 0.000 description 10
- FZQLXNIMCPJVJE-YUMQZZPRSA-N Gly-Asp-Leu Chemical compound [H]NCC(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CC(C)C)C(O)=O FZQLXNIMCPJVJE-YUMQZZPRSA-N 0.000 description 10
- LXXANCRPFBSSKS-IUCAKERBSA-N Gly-Gln-Leu Chemical compound [H]NCC(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CC(C)C)C(O)=O LXXANCRPFBSSKS-IUCAKERBSA-N 0.000 description 10
- CCQOOWAONKGYKQ-BYPYZUCNSA-N Gly-Gly-Ala Chemical compound OC(=O)[C@H](C)NC(=O)CNC(=O)CN CCQOOWAONKGYKQ-BYPYZUCNSA-N 0.000 description 10
- SXJHOPPTOJACOA-QXEWZRGKSA-N Gly-Ile-Arg Chemical compound NCC(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@H](C(O)=O)CCCN=C(N)N SXJHOPPTOJACOA-QXEWZRGKSA-N 0.000 description 10
- COVXELOAORHTND-LSJOCFKGSA-N Gly-Ile-Val Chemical compound NCC(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](C(C)C)C(O)=O COVXELOAORHTND-LSJOCFKGSA-N 0.000 description 10
- IALQAMYQJBZNSK-WHFBIAKZSA-N Gly-Ser-Asn Chemical compound [H]NCC(=O)N[C@@H](CO)C(=O)N[C@@H](CC(N)=O)C(O)=O IALQAMYQJBZNSK-WHFBIAKZSA-N 0.000 description 10
- MKIAPEZXQDILRR-YUMQZZPRSA-N Gly-Ser-His Chemical compound C1=C(NC=N1)C[C@@H](C(=O)O)NC(=O)[C@H](CO)NC(=O)CN MKIAPEZXQDILRR-YUMQZZPRSA-N 0.000 description 10
- CUVBTVWFVIIDOC-YEPSODPASA-N Gly-Thr-Val Chemical compound CC(C)[C@@H](C(O)=O)NC(=O)[C@H]([C@@H](C)O)NC(=O)CN CUVBTVWFVIIDOC-YEPSODPASA-N 0.000 description 10
- KOYUSMBPJOVSOO-XEGUGMAKSA-N Gly-Tyr-Ile Chemical compound [H]NCC(=O)N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O KOYUSMBPJOVSOO-XEGUGMAKSA-N 0.000 description 10
- DNVDEMWIYLVIQU-RCOVLWMOSA-N Gly-Val-Asp Chemical compound NCC(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CC(O)=O)C(O)=O DNVDEMWIYLVIQU-RCOVLWMOSA-N 0.000 description 10
- FLUVGKKRRMLNPU-CQDKDKBSSA-N His-Ala-Phe Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)N[C@@H](C)C(=O)N[C@@H](CC1=CC=CC=C1)C(O)=O FLUVGKKRRMLNPU-CQDKDKBSSA-N 0.000 description 10
- PDSUIXMZYNURGI-AVGNSLFASA-N His-Arg-Arg Chemical compound NC(N)=NCCC[C@@H](C(O)=O)NC(=O)[C@H](CCCN=C(N)N)NC(=O)[C@@H](N)CC1=CN=CN1 PDSUIXMZYNURGI-AVGNSLFASA-N 0.000 description 10
- YVCGJPIKRMGNPA-LSJOCFKGSA-N His-Met-Ala Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)N[C@@H](CCSC)C(=O)N[C@@H](C)C(O)=O YVCGJPIKRMGNPA-LSJOCFKGSA-N 0.000 description 10
- GNBHSMFBUNEWCJ-DCAQKATOSA-N His-Pro-Asn Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CC(N)=O)C(O)=O GNBHSMFBUNEWCJ-DCAQKATOSA-N 0.000 description 10
- PFOUFRJYHWZJKW-NKIYYHGXSA-N His-Thr-Gln Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CCC(=O)N)C(=O)O)NC(=O)[C@H](CC1=CN=CN1)N)O PFOUFRJYHWZJKW-NKIYYHGXSA-N 0.000 description 10
- UWSMZKRTOZEGDD-CUJWVEQBSA-N His-Thr-Ser Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CO)C(O)=O UWSMZKRTOZEGDD-CUJWVEQBSA-N 0.000 description 10
- FVEWRQXNISSYFO-ZPFDUUQYSA-N Ile-Arg-Glu Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CCCN=C(N)N)C(=O)N[C@@H](CCC(=O)O)C(=O)O)N FVEWRQXNISSYFO-ZPFDUUQYSA-N 0.000 description 10
- HVWXAQVMRBKKFE-UGYAYLCHSA-N Ile-Asp-Asp Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CC(=O)O)C(=O)N[C@@H](CC(=O)O)C(=O)O)N HVWXAQVMRBKKFE-UGYAYLCHSA-N 0.000 description 10
- LPXHYGGZJOCAFR-MNXVOIDGSA-N Ile-Glu-Leu Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CCC(=O)O)C(=O)N[C@@H](CC(C)C)C(=O)O)N LPXHYGGZJOCAFR-MNXVOIDGSA-N 0.000 description 10
- LWWILHPVAKKLQS-QXEWZRGKSA-N Ile-Gly-Met Chemical compound CC[C@H](C)[C@@H](C(=O)NCC(=O)N[C@@H](CCSC)C(=O)O)N LWWILHPVAKKLQS-QXEWZRGKSA-N 0.000 description 10
- AXNGDPAKKCEKGY-QPHKQPEJSA-N Ile-Ile-Thr Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H]([C@@H](C)O)C(=O)O)N AXNGDPAKKCEKGY-QPHKQPEJSA-N 0.000 description 10
- KLBVGHCGHUNHEA-BJDJZHNGSA-N Ile-Leu-Ala Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](C)C(=O)O)N KLBVGHCGHUNHEA-BJDJZHNGSA-N 0.000 description 10
- 241000880493 Leptailurus serval Species 0.000 description 10
- CQQGCWPXDHTTNF-GUBZILKMSA-N Leu-Ala-Glu Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](C)C(=O)N[C@H](C(O)=O)CCC(O)=O CQQGCWPXDHTTNF-GUBZILKMSA-N 0.000 description 10
- SUPVSFFZWVOEOI-CQDKDKBSSA-N Leu-Ala-Tyr Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](C)C(=O)N[C@H](C(O)=O)CC1=CC=C(O)C=C1 SUPVSFFZWVOEOI-CQDKDKBSSA-N 0.000 description 10
- SUPVSFFZWVOEOI-UHFFFAOYSA-N Leu-Ala-Tyr Natural products CC(C)CC(N)C(=O)NC(C)C(=O)NC(C(O)=O)CC1=CC=C(O)C=C1 SUPVSFFZWVOEOI-UHFFFAOYSA-N 0.000 description 10
- HBJZFCIVFIBNSV-DCAQKATOSA-N Leu-Arg-Asn Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](CCCN=C(N)N)C(=O)N[C@@H](CC(N)=O)C(O)=O HBJZFCIVFIBNSV-DCAQKATOSA-N 0.000 description 10
- JKGHDYGZRDWHGA-SRVKXCTJSA-N Leu-Asn-Leu Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CC(C)C)C(O)=O JKGHDYGZRDWHGA-SRVKXCTJSA-N 0.000 description 10
- USTCFDAQCLDPBD-XIRDDKMYSA-N Leu-Asn-Trp Chemical compound CC(C)C[C@@H](C(=O)N[C@@H](CC(=O)N)C(=O)N[C@@H](CC1=CNC2=CC=CC=C21)C(=O)O)N USTCFDAQCLDPBD-XIRDDKMYSA-N 0.000 description 10
- RXGLHDWAZQECBI-SRVKXCTJSA-N Leu-Leu-Ser Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CO)C(O)=O RXGLHDWAZQECBI-SRVKXCTJSA-N 0.000 description 10
- JLWZLIQRYCTYBD-IHRRRGAJSA-N Leu-Lys-Arg Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O JLWZLIQRYCTYBD-IHRRRGAJSA-N 0.000 description 10
- UCBPDSYUVAAHCD-UWVGGRQHSA-N Leu-Pro-Gly Chemical compound CC(C)C[C@H](N)C(=O)N1CCC[C@H]1C(=O)NCC(O)=O UCBPDSYUVAAHCD-UWVGGRQHSA-N 0.000 description 10
- PWPBLZXWFXJFHE-RHYQMDGZSA-N Leu-Pro-Thr Chemical compound CC(C)C[C@H](N)C(=O)N1CCC[C@H]1C(=O)N[C@@H]([C@@H](C)O)C(O)=O PWPBLZXWFXJFHE-RHYQMDGZSA-N 0.000 description 10
- IDGZVZJLYFTXSL-DCAQKATOSA-N Leu-Ser-Arg Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](CO)C(=O)N[C@H](C(O)=O)CCCN=C(N)N IDGZVZJLYFTXSL-DCAQKATOSA-N 0.000 description 10
- QWWPYKKLXWOITQ-VOAKCMCISA-N Leu-Thr-Leu Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@H](C(O)=O)CC(C)C QWWPYKKLXWOITQ-VOAKCMCISA-N 0.000 description 10
- YIRIDPUGZKHMHT-ACRUOGEOSA-N Leu-Tyr-Tyr Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(O)=O YIRIDPUGZKHMHT-ACRUOGEOSA-N 0.000 description 10
- FBNPMTNBFFAMMH-AVGNSLFASA-N Leu-Val-Arg Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](C(C)C)C(=O)N[C@H](C(O)=O)CCCN=C(N)N FBNPMTNBFFAMMH-AVGNSLFASA-N 0.000 description 10
- FBNPMTNBFFAMMH-UHFFFAOYSA-N Leu-Val-Arg Natural products CC(C)CC(N)C(=O)NC(C(C)C)C(=O)NC(C(O)=O)CCCN=C(N)N FBNPMTNBFFAMMH-UHFFFAOYSA-N 0.000 description 10
- GIKFNMZSGYAPEJ-HJGDQZAQSA-N Lys-Thr-Asp Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC(O)=O)C(O)=O GIKFNMZSGYAPEJ-HJGDQZAQSA-N 0.000 description 10
- KXYLFJIQDIMURW-IHPCNDPISA-N Lys-Trp-Leu Chemical compound C1=CC=C2C(C[C@@H](C(=O)N[C@@H](CC(C)C)C(O)=O)NC(=O)[C@@H](N)CCCCN)=CNC2=C1 KXYLFJIQDIMURW-IHPCNDPISA-N 0.000 description 10
- DRRXXZBXDMLGFC-IHRRRGAJSA-N Lys-Val-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@H](C(C)C)NC(=O)[C@@H](N)CCCCN DRRXXZBXDMLGFC-IHRRRGAJSA-N 0.000 description 10
- DTICLBJHRYSJLH-GUBZILKMSA-N Met-Ala-Val Chemical compound CSCC[C@H](N)C(=O)N[C@@H](C)C(=O)N[C@@H](C(C)C)C(O)=O DTICLBJHRYSJLH-GUBZILKMSA-N 0.000 description 10
- GPAHWYRSHCKICP-GUBZILKMSA-N Met-Glu-Glu Chemical compound CSCC[C@H](N)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCC(O)=O)C(O)=O GPAHWYRSHCKICP-GUBZILKMSA-N 0.000 description 10
- UROWNMBTQGGTHB-DCAQKATOSA-N Met-Leu-Asp Chemical compound CSCC[C@H](N)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC(O)=O)C(O)=O UROWNMBTQGGTHB-DCAQKATOSA-N 0.000 description 10
- CIIJWIAORKTXAH-FJXKBIBVSA-N Met-Thr-Gly Chemical compound CSCC[C@H](N)C(=O)N[C@@H]([C@@H](C)O)C(=O)NCC(O)=O CIIJWIAORKTXAH-FJXKBIBVSA-N 0.000 description 10
- IIHMNTBFPMRJCN-RCWTZXSCSA-N Met-Val-Thr Chemical compound CSCC[C@H](N)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H]([C@@H](C)O)C(O)=O IIHMNTBFPMRJCN-RCWTZXSCSA-N 0.000 description 10
- YBAFDPFAUTYYRW-UHFFFAOYSA-N N-L-alpha-glutamyl-L-leucine Natural products CC(C)CC(C(O)=O)NC(=O)C(N)CCC(O)=O YBAFDPFAUTYYRW-UHFFFAOYSA-N 0.000 description 10
- SITLTJHOQZFJGG-UHFFFAOYSA-N N-L-alpha-glutamyl-L-valine Natural products CC(C)C(C(O)=O)NC(=O)C(N)CCC(O)=O SITLTJHOQZFJGG-UHFFFAOYSA-N 0.000 description 10
- XMBSYZWANAQXEV-UHFFFAOYSA-N N-alpha-L-glutamyl-L-phenylalanine Natural products OC(=O)CCC(N)C(=O)NC(C(O)=O)CC1=CC=CC=C1 XMBSYZWANAQXEV-UHFFFAOYSA-N 0.000 description 10
- KZNQNBZMBZJQJO-UHFFFAOYSA-N N-glycyl-L-proline Natural products NCC(=O)N1CCCC1C(O)=O KZNQNBZMBZJQJO-UHFFFAOYSA-N 0.000 description 10
- KAGCQPSEVAETCA-JYJNAYRXSA-N Phe-Gln-Leu Chemical compound CC(C)C[C@@H](C(=O)O)NC(=O)[C@H](CCC(=O)N)NC(=O)[C@H](CC1=CC=CC=C1)N KAGCQPSEVAETCA-JYJNAYRXSA-N 0.000 description 10
- JEBWZLWTRPZQRX-QWRGUYRKSA-N Phe-Gly-Asp Chemical compound [H]N[C@@H](CC1=CC=CC=C1)C(=O)NCC(=O)N[C@@H](CC(O)=O)C(O)=O JEBWZLWTRPZQRX-QWRGUYRKSA-N 0.000 description 10
- XEXSSIBQYNKFBX-KBPBESRZSA-N Phe-Gly-His Chemical compound C([C@H](N)C(=O)NCC(=O)N[C@@H](CC=1N=CNC=1)C(O)=O)C1=CC=CC=C1 XEXSSIBQYNKFBX-KBPBESRZSA-N 0.000 description 10
- FINLZXKJWTYYLC-ACRUOGEOSA-N Phe-His-Phe Chemical compound C([C@H](N)C(=O)N[C@@H](CC=1N=CNC=1)C(=O)N[C@@H](CC=1C=CC=CC=1)C(O)=O)C1=CC=CC=C1 FINLZXKJWTYYLC-ACRUOGEOSA-N 0.000 description 10
- ZIQQNOXKEFDPBE-BZSNNMDCSA-N Phe-Lys-His Chemical compound C1=CC=C(C=C1)C[C@@H](C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC2=CN=CN2)C(=O)O)N ZIQQNOXKEFDPBE-BZSNNMDCSA-N 0.000 description 10
- CZQZSMJXFGGBHM-KKUMJFAQSA-N Phe-Pro-Gln Chemical compound C1C[C@H](N(C1)C(=O)[C@H](CC2=CC=CC=C2)N)C(=O)N[C@@H](CCC(=O)N)C(=O)O CZQZSMJXFGGBHM-KKUMJFAQSA-N 0.000 description 10
- GMWNQSGWWGKTSF-LFSVMHDDSA-N Phe-Thr-Ala Chemical compound [H]N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](C)C(O)=O GMWNQSGWWGKTSF-LFSVMHDDSA-N 0.000 description 10
- AOKZOUGUMLBPSS-PMVMPFDFSA-N Phe-Trp-Leu Chemical compound [H]N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CC1=CNC2=C1C=CC=C2)C(=O)N[C@@H](CC(C)C)C(O)=O AOKZOUGUMLBPSS-PMVMPFDFSA-N 0.000 description 10
- BAONJAHBAUDJKA-BZSNNMDCSA-N Phe-Tyr-Asp Chemical compound C([C@H](N)C(=O)N[C@@H](CC=1C=CC(O)=CC=1)C(=O)N[C@@H](CC(O)=O)C(O)=O)C1=CC=CC=C1 BAONJAHBAUDJKA-BZSNNMDCSA-N 0.000 description 10
- IPVPGAADZXRZSH-RNXOBYDBSA-N Phe-Tyr-Trp Chemical compound [H]N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CC1=CNC2=C1C=CC=C2)C(O)=O IPVPGAADZXRZSH-RNXOBYDBSA-N 0.000 description 10
- XZGWNSIRZIUHHP-SRVKXCTJSA-N Pro-Arg-Met Chemical compound CSCC[C@@H](C(=O)O)NC(=O)[C@H](CCCN=C(N)N)NC(=O)[C@@H]1CCCN1 XZGWNSIRZIUHHP-SRVKXCTJSA-N 0.000 description 10
- SGCZFWSQERRKBD-BQBZGAKWSA-N Pro-Asp-Gly Chemical compound OC(=O)CNC(=O)[C@H](CC(O)=O)NC(=O)[C@@H]1CCCN1 SGCZFWSQERRKBD-BQBZGAKWSA-N 0.000 description 10
- KPDRZQUWJKTMBP-DCAQKATOSA-N Pro-Asp-Leu Chemical compound CC(C)C[C@@H](C(=O)O)NC(=O)[C@H](CC(=O)O)NC(=O)[C@@H]1CCCN1 KPDRZQUWJKTMBP-DCAQKATOSA-N 0.000 description 10
- WFHYFCWBLSKEMS-KKUMJFAQSA-N Pro-Glu-Phe Chemical compound N([C@@H](CCC(=O)O)C(=O)N[C@@H](CC=1C=CC=CC=1)C(O)=O)C(=O)[C@@H]1CCCN1 WFHYFCWBLSKEMS-KKUMJFAQSA-N 0.000 description 10
- UEHYFUCOGHWASA-HJGDQZAQSA-N Pro-Glu-Thr Chemical compound C[C@@H](O)[C@@H](C(O)=O)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@@H]1CCCN1 UEHYFUCOGHWASA-HJGDQZAQSA-N 0.000 description 10
- CLNJSLSHKJECME-BQBZGAKWSA-N Pro-Gly-Ala Chemical compound OC(=O)[C@H](C)NC(=O)CNC(=O)[C@@H]1CCCN1 CLNJSLSHKJECME-BQBZGAKWSA-N 0.000 description 10
- WSRWHZRUOCACLJ-UWVGGRQHSA-N Pro-Gly-His Chemical compound C([C@@H](C(=O)O)NC(=O)CNC(=O)[C@H]1NCCC1)C1=CN=CN1 WSRWHZRUOCACLJ-UWVGGRQHSA-N 0.000 description 10
- KWMUAKQOVYCQJQ-ZPFDUUQYSA-N Pro-Ile-Glu Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CCC(=O)O)C(=O)O)NC(=O)[C@@H]1CCCN1 KWMUAKQOVYCQJQ-ZPFDUUQYSA-N 0.000 description 10
- FKVNLUZHSFCNGY-RVMXOQNASA-N Pro-Ile-Pro Chemical compound CC[C@H](C)[C@@H](C(=O)N1CCC[C@@H]1C(=O)O)NC(=O)[C@@H]2CCCN2 FKVNLUZHSFCNGY-RVMXOQNASA-N 0.000 description 10
- UREQLMJCKFLLHM-NAKRPEOUSA-N Pro-Ile-Ser Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CO)C(O)=O UREQLMJCKFLLHM-NAKRPEOUSA-N 0.000 description 10
- YXHYJEPDKSYPSQ-AVGNSLFASA-N Pro-Leu-Arg Chemical compound NC(N)=NCCC[C@@H](C(O)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@@H]1CCCN1 YXHYJEPDKSYPSQ-AVGNSLFASA-N 0.000 description 10
- HATVCTYBNCNMAA-AVGNSLFASA-N Pro-Leu-Met Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCSC)C(O)=O HATVCTYBNCNMAA-AVGNSLFASA-N 0.000 description 10
- GFHOSBYCLACKEK-GUBZILKMSA-N Pro-Pro-Asn Chemical compound [H]N1CCC[C@H]1C(=O)N1CCC[C@H]1C(=O)N[C@@H](CC(N)=O)C(O)=O GFHOSBYCLACKEK-GUBZILKMSA-N 0.000 description 10
- NAIPAPCKKRCMBL-JYJNAYRXSA-N Pro-Pro-Phe Chemical compound C([C@@H](C(=O)O)NC(=O)[C@H]1N(CCC1)C(=O)[C@H]1NCCC1)C1=CC=CC=C1 NAIPAPCKKRCMBL-JYJNAYRXSA-N 0.000 description 10
- ITUDDXVFGFEKPD-NAKRPEOUSA-N Pro-Ser-Ile Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](CO)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O ITUDDXVFGFEKPD-NAKRPEOUSA-N 0.000 description 10
- 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 10
- LZHHZYDPMZEMRX-STQMWFEESA-N Pro-Tyr-Gly Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(=O)NCC(O)=O LZHHZYDPMZEMRX-STQMWFEESA-N 0.000 description 10
- YHUBAXGAAYULJY-ULQDDVLXSA-N Pro-Tyr-Leu Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CC(C)C)C(O)=O YHUBAXGAAYULJY-ULQDDVLXSA-N 0.000 description 10
- IMNVAOPEMFDAQD-NHCYSSNCSA-N Pro-Val-Glu Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CCC(O)=O)C(O)=O IMNVAOPEMFDAQD-NHCYSSNCSA-N 0.000 description 10
- QGMLKFGTGXWAHF-IHRRRGAJSA-N Ser-Arg-Phe Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC1=CC=CC=C1)C(O)=O QGMLKFGTGXWAHF-IHRRRGAJSA-N 0.000 description 10
- XVAUJOAYHWWNQF-ZLUOBGJFSA-N Ser-Asn-Ala Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](C)C(O)=O XVAUJOAYHWWNQF-ZLUOBGJFSA-N 0.000 description 10
- ZFVFHHZBCVNLGD-GUBZILKMSA-N Ser-His-Gln Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CC1=CNC=N1)C(=O)N[C@@H](CCC(N)=O)C(O)=O ZFVFHHZBCVNLGD-GUBZILKMSA-N 0.000 description 10
- GZSZPKSBVAOGIE-CIUDSAMLSA-N Ser-Lys-Ala Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](C)C(O)=O GZSZPKSBVAOGIE-CIUDSAMLSA-N 0.000 description 10
- CKDXFSPMIDSMGV-GUBZILKMSA-N Ser-Pro-Val Chemical compound [H]N[C@@H](CO)C(=O)N1CCC[C@H]1C(=O)N[C@@H](C(C)C)C(O)=O CKDXFSPMIDSMGV-GUBZILKMSA-N 0.000 description 10
- BMKNXTJLHFIAAH-CIUDSAMLSA-N Ser-Ser-Leu Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CO)C(=O)N[C@@H](CC(C)C)C(O)=O BMKNXTJLHFIAAH-CIUDSAMLSA-N 0.000 description 10
- IAOHCSQDQDWRQU-GUBZILKMSA-N Ser-Val-Arg Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O IAOHCSQDQDWRQU-GUBZILKMSA-N 0.000 description 10
- JMZKMSTYXHFYAK-VEVYYDQMSA-N Thr-Arg-Asn Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CCCN=C(N)N)C(=O)N[C@@H](CC(=O)N)C(=O)O)N)O JMZKMSTYXHFYAK-VEVYYDQMSA-N 0.000 description 10
- LHUBVKCLOVALIA-HJGDQZAQSA-N Thr-Arg-Gln Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCC(N)=O)C(O)=O LHUBVKCLOVALIA-HJGDQZAQSA-N 0.000 description 10
- VASYSJHSMSBTDU-LKXGYXEUSA-N Thr-Asn-Cys Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CC(=O)N)C(=O)N[C@@H](CS)C(=O)O)N)O VASYSJHSMSBTDU-LKXGYXEUSA-N 0.000 description 10
- VBPDMBAFBRDZSK-HOUAVDHOSA-N Thr-Asn-Trp Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CC(=O)N)C(=O)N[C@@H](CC1=CNC2=CC=CC=C21)C(=O)O)N)O VBPDMBAFBRDZSK-HOUAVDHOSA-N 0.000 description 10
- XPNSAQMEAVSQRD-FBCQKBJTSA-N Thr-Gly-Gly Chemical compound C[C@@H](O)[C@H](N)C(=O)NCC(=O)NCC(O)=O XPNSAQMEAVSQRD-FBCQKBJTSA-N 0.000 description 10
- VTVVYQOXJCZVEB-WDCWCFNPSA-N Thr-Leu-Glu Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(O)=O)C(O)=O VTVVYQOXJCZVEB-WDCWCFNPSA-N 0.000 description 10
- MEJHFIOYJHTWMK-VOAKCMCISA-N Thr-Leu-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@@H](N)[C@@H](C)O MEJHFIOYJHTWMK-VOAKCMCISA-N 0.000 description 10
- MXDOAJQRJBMGMO-FJXKBIBVSA-N Thr-Pro-Gly Chemical compound C[C@@H](O)[C@H](N)C(=O)N1CCC[C@H]1C(=O)NCC(O)=O MXDOAJQRJBMGMO-FJXKBIBVSA-N 0.000 description 10
- BDENGIGFTNYZSJ-RCWTZXSCSA-N Thr-Pro-Met Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CCSC)C(O)=O BDENGIGFTNYZSJ-RCWTZXSCSA-N 0.000 description 10
- UJRIVCPPPMYCNA-HOCLYGCPSA-N Trp-Leu-Gly Chemical compound CC(C)C[C@@H](C(=O)NCC(=O)O)NC(=O)[C@H](CC1=CNC2=CC=CC=C21)N UJRIVCPPPMYCNA-HOCLYGCPSA-N 0.000 description 10
- XGFOXYJQBRTJPO-PJODQICGSA-N Trp-Pro-Ala Chemical compound [H]N[C@@H](CC1=CNC2=C1C=CC=C2)C(=O)N1CCC[C@H]1C(=O)N[C@@H](C)C(O)=O XGFOXYJQBRTJPO-PJODQICGSA-N 0.000 description 10
- YXSSXUIBUJGHJY-SFJXLCSZSA-N Trp-Thr-Phe Chemical compound C([C@H](NC(=O)[C@@H](NC(=O)[C@@H](N)CC=1C2=CC=CC=C2NC=1)[C@H](O)C)C(O)=O)C1=CC=CC=C1 YXSSXUIBUJGHJY-SFJXLCSZSA-N 0.000 description 10
- NLMXVDDEQFKQQU-CFMVVWHZSA-N Tyr-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)CC1=CC=C(O)C=C1 NLMXVDDEQFKQQU-CFMVVWHZSA-N 0.000 description 10
- VFJIWSJKZJTQII-SRVKXCTJSA-N Tyr-Asp-Ser Chemical compound [H]N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CO)C(O)=O VFJIWSJKZJTQII-SRVKXCTJSA-N 0.000 description 10
- UNUZEBFXGWVAOP-DZKIICNBSA-N Tyr-Glu-Val Chemical compound [H]N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](C(C)C)C(O)=O UNUZEBFXGWVAOP-DZKIICNBSA-N 0.000 description 10
- JHORGUYURUBVOM-KKUMJFAQSA-N Tyr-His-Ser Chemical compound [H]N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CC1=CNC=N1)C(=O)N[C@@H](CO)C(O)=O JHORGUYURUBVOM-KKUMJFAQSA-N 0.000 description 10
- WDGDKHLSDIOXQC-ACRUOGEOSA-N Tyr-Leu-Phe Chemical compound C([C@H](N)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC=1C=CC=CC=1)C(O)=O)C1=CC=C(O)C=C1 WDGDKHLSDIOXQC-ACRUOGEOSA-N 0.000 description 10
- KZOZXAYPVKKDIO-UFYCRDLUSA-N Tyr-Met-Tyr Chemical compound C([C@H](N)C(=O)N[C@@H](CCSC)C(=O)N[C@@H](CC=1C=CC(O)=CC=1)C(O)=O)C1=CC=C(O)C=C1 KZOZXAYPVKKDIO-UFYCRDLUSA-N 0.000 description 10
- ULUXAIYMVXLDQP-PMVMPFDFSA-N Tyr-Trp-His Chemical compound C1=CC=C2C(=C1)C(=CN2)C[C@@H](C(=O)N[C@@H](CC3=CN=CN3)C(=O)O)NC(=O)[C@H](CC4=CC=C(C=C4)O)N ULUXAIYMVXLDQP-PMVMPFDFSA-N 0.000 description 10
- PQPWEALFTLKSEB-DZKIICNBSA-N Tyr-Val-Glu Chemical compound [H]N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CCC(O)=O)C(O)=O PQPWEALFTLKSEB-DZKIICNBSA-N 0.000 description 10
- 108010064997 VPY tripeptide Proteins 0.000 description 10
- LABUITCFCAABSV-BPNCWPANSA-N Val-Ala-Tyr Chemical compound CC(C)[C@H](N)C(=O)N[C@@H](C)C(=O)N[C@H](C(O)=O)CC1=CC=C(O)C=C1 LABUITCFCAABSV-BPNCWPANSA-N 0.000 description 10
- LABUITCFCAABSV-UHFFFAOYSA-N Val-Ala-Tyr Natural products CC(C)C(N)C(=O)NC(C)C(=O)NC(C(O)=O)CC1=CC=C(O)C=C1 LABUITCFCAABSV-UHFFFAOYSA-N 0.000 description 10
- BYOHPUZJVXWHAE-BYULHYEWSA-N Val-Asn-Asn Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CC(=O)N)C(=O)N[C@@H](CC(=O)N)C(=O)O)N BYOHPUZJVXWHAE-BYULHYEWSA-N 0.000 description 10
- DBOXBUDEAJVKRE-LSJOCFKGSA-N Val-Asn-Val Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CC(=O)N)C(=O)N[C@@H](C(C)C)C(=O)O)N DBOXBUDEAJVKRE-LSJOCFKGSA-N 0.000 description 10
- XGJLNBNZNMVJRS-NRPADANISA-N Val-Glu-Ala Chemical compound CC(C)[C@H](N)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](C)C(O)=O XGJLNBNZNMVJRS-NRPADANISA-N 0.000 description 10
- 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 10
- BMOFUVHDBROBSE-DCAQKATOSA-N Val-Leu-Cys Chemical compound CC(C)C[C@@H](C(=O)N[C@@H](CS)C(=O)O)NC(=O)[C@H](C(C)C)N BMOFUVHDBROBSE-DCAQKATOSA-N 0.000 description 10
- SVFRYKBZHUGKLP-QXEWZRGKSA-N Val-Met-Asn Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CCSC)C(=O)N[C@@H](CC(=O)N)C(=O)O)N SVFRYKBZHUGKLP-QXEWZRGKSA-N 0.000 description 10
- SVLAAUGFIHSJPK-JYJNAYRXSA-N Val-Trp-Ser Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CC1=CNC2=CC=CC=C21)C(=O)N[C@@H](CO)C(=O)O)N SVLAAUGFIHSJPK-JYJNAYRXSA-N 0.000 description 10
- GUIYPEKUEMQBIK-JSGCOSHPSA-N Val-Tyr-Gly Chemical compound CC(C)[C@H](N)C(=O)N[C@@H](Cc1ccc(O)cc1)C(=O)NCC(O)=O GUIYPEKUEMQBIK-JSGCOSHPSA-N 0.000 description 10
- 108010008685 alanyl-glutamyl-aspartic acid Proteins 0.000 description 10
- 108010041407 alanylaspartic acid Proteins 0.000 description 10
- 108010047495 alanylglycine Proteins 0.000 description 10
- 108010070944 alanylhistidine Proteins 0.000 description 10
- 108010010430 asparagine-proline-alanine Proteins 0.000 description 10
- 108010093581 aspartyl-proline Proteins 0.000 description 10
- 108010038633 aspartylglutamate Proteins 0.000 description 10
- 108010047857 aspartylglycine Proteins 0.000 description 10
- 108010068265 aspartyltyrosine Proteins 0.000 description 10
- 108010078144 glutaminyl-glycine Proteins 0.000 description 10
- 108010033719 glycyl-histidyl-glycine Proteins 0.000 description 10
- 108010050848 glycylleucine Proteins 0.000 description 10
- 108010040030 histidinoalanine Proteins 0.000 description 10
- 108010041601 histidyl-aspartyl-glutamyl-leucine Proteins 0.000 description 10
- 108010083708 leucyl-aspartyl-valine Proteins 0.000 description 10
- 108010044311 leucyl-glycyl-glycine Proteins 0.000 description 10
- 108010005942 methionylglycine Proteins 0.000 description 10
- 108010012581 phenylalanylglutamate Proteins 0.000 description 10
- 108010051242 phenylalanylserine Proteins 0.000 description 10
- 108010020432 prolyl-prolylisoleucine Proteins 0.000 description 10
- 108010026333 seryl-proline Proteins 0.000 description 10
- 108010080629 tryptophan-leucine Proteins 0.000 description 10
- 108010051110 tyrosyl-lysine Proteins 0.000 description 10
- IBIDRSSEHFLGSD-UHFFFAOYSA-N valinyl-arginine Natural products CC(C)C(N)C(=O)NC(C(O)=O)CCCN=C(N)N IBIDRSSEHFLGSD-UHFFFAOYSA-N 0.000 description 10
- 235000014469 Bacillus subtilis Nutrition 0.000 description 9
- 210000004027 cell Anatomy 0.000 description 9
- 244000063299 Bacillus subtilis Species 0.000 description 7
- 239000000758 substrate Substances 0.000 description 7
- LGQPPBQRUBVTIF-JBDRJPRFSA-N Ala-Ala-Ile Chemical compound [H]N[C@@H](C)C(=O)N[C@@H](C)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O LGQPPBQRUBVTIF-JBDRJPRFSA-N 0.000 description 6
- QCTOLCVIGRLMQS-HRCADAONSA-N Arg-Tyr-Pro Chemical compound C1C[C@@H](N(C1)C(=O)[C@H](CC2=CC=C(C=C2)O)NC(=O)[C@H](CCCN=C(N)N)N)C(=O)O QCTOLCVIGRLMQS-HRCADAONSA-N 0.000 description 6
- XQLGNKLSPYCRMZ-HJWJTTGWSA-N Ile-Phe-Val Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](C(C)C)C(=O)O)N XQLGNKLSPYCRMZ-HJWJTTGWSA-N 0.000 description 6
- IVDFVBVIVLJJHR-LKXGYXEUSA-N Thr-Ser-Asp Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CO)C(=O)N[C@@H](CC(O)=O)C(O)=O IVDFVBVIVLJJHR-LKXGYXEUSA-N 0.000 description 6
- 229910021645 metal ion Inorganic materials 0.000 description 6
- 238000002703 mutagenesis Methods 0.000 description 6
- 231100000350 mutagenesis Toxicity 0.000 description 6
- OWEGMIWEEQEYGQ-UHFFFAOYSA-N 100676-05-9 Natural products OC1C(O)C(O)C(CO)OC1OCC1C(O)C(O)C(O)C(OC2C(OC(O)C(O)C2O)CO)O1 OWEGMIWEEQEYGQ-UHFFFAOYSA-N 0.000 description 5
- 101100363100 Chlorobium chlorochromatii (strain CaD3) rpsU gene Proteins 0.000 description 5
- 241001200922 Gagata Species 0.000 description 5
- GUBGYTABKSRVRQ-PICCSMPSSA-N Maltose Natural products O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@@H](CO)OC(O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-PICCSMPSSA-N 0.000 description 5
- 239000007788 liquid Substances 0.000 description 5
- 239000013612 plasmid Substances 0.000 description 5
- 239000000047 product Substances 0.000 description 5
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 5
- LSMDIAAALJJLRO-XQXXSGGOSA-N Ala-Thr-Glu Chemical compound [H]N[C@@H](C)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CCC(O)=O)C(O)=O LSMDIAAALJJLRO-XQXXSGGOSA-N 0.000 description 4
- 239000002609 medium Substances 0.000 description 4
- 238000000034 method Methods 0.000 description 4
- 235000018102 proteins Nutrition 0.000 description 4
- 102000004169 proteins and genes Human genes 0.000 description 4
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical group CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 description 3
- 102000018120 Recombinases Human genes 0.000 description 3
- 108010091086 Recombinases Proteins 0.000 description 3
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 3
- 239000007983 Tris buffer Substances 0.000 description 3
- 229910021529 ammonia Inorganic materials 0.000 description 3
- QGZKDVFQNNGYKY-UHFFFAOYSA-N ammonia Natural products N QGZKDVFQNNGYKY-UHFFFAOYSA-N 0.000 description 3
- 239000012153 distilled water Substances 0.000 description 3
- 238000002474 experimental method Methods 0.000 description 3
- 238000000855 fermentation Methods 0.000 description 3
- 230000004151 fermentation Effects 0.000 description 3
- 239000012530 fluid Substances 0.000 description 3
- 238000004128 high performance liquid chromatography Methods 0.000 description 3
- 230000001939 inductive effect Effects 0.000 description 3
- 238000011068 loading method Methods 0.000 description 3
- 239000000243 solution Substances 0.000 description 3
- 239000006228 supernatant Substances 0.000 description 3
- 238000012360 testing method Methods 0.000 description 3
- LENZDBCJOHFCAS-UHFFFAOYSA-N tris Chemical compound OCC(N)(CO)CO LENZDBCJOHFCAS-UHFFFAOYSA-N 0.000 description 3
- FRXSZNDVFUDTIR-UHFFFAOYSA-N 6-methoxy-1,2,3,4-tetrahydroquinoline Chemical compound N1CCCC2=CC(OC)=CC=C21 FRXSZNDVFUDTIR-UHFFFAOYSA-N 0.000 description 2
- GWWSUMLEWKQHLR-NUMRIWBASA-N Asp-Thr-Glu Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CCC(=O)O)C(=O)O)NC(=O)[C@H](CC(=O)O)N)O GWWSUMLEWKQHLR-NUMRIWBASA-N 0.000 description 2
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- 241000193755 Bacillus cereus Species 0.000 description 2
- 238000006555 catalytic reaction Methods 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 230000002255 enzymatic effect Effects 0.000 description 2
- 239000013604 expression vector Substances 0.000 description 2
- 230000002401 inhibitory effect Effects 0.000 description 2
- 230000008569 process Effects 0.000 description 2
- 238000002415 sodium dodecyl sulfate polyacrylamide gel electrophoresis Methods 0.000 description 2
- 241000894007 species Species 0.000 description 2
- SVBWNHOBPFJIRU-UHFFFAOYSA-N 1-O-alpha-D-Glucopyranosyl-D-fructose Natural products OC1C(O)C(O)C(CO)OC1OCC1(O)C(O)C(O)C(O)CO1 SVBWNHOBPFJIRU-UHFFFAOYSA-N 0.000 description 1
- DBTMGCOVALSLOR-UHFFFAOYSA-N 32-alpha-galactosyl-3-alpha-galactosyl-galactose Natural products OC1C(O)C(O)C(CO)OC1OC1C(O)C(OC2C(C(CO)OC(O)C2O)O)OC(CO)C1O DBTMGCOVALSLOR-UHFFFAOYSA-N 0.000 description 1
- PVXPPJIGRGXGCY-TZLCEDOOSA-N 6-O-alpha-D-glucopyranosyl-D-fructofuranose Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1OC[C@@H]1[C@@H](O)[C@H](O)C(O)(CO)O1 PVXPPJIGRGXGCY-TZLCEDOOSA-N 0.000 description 1
- RXVWSYJTUUKTEA-UHFFFAOYSA-N D-maltotriose Natural products OC1C(O)C(OC(C(O)CO)C(O)C(O)C=O)OC(CO)C1OC1C(O)C(O)C(O)C(CO)O1 RXVWSYJTUUKTEA-UHFFFAOYSA-N 0.000 description 1
- 241000192091 Deinococcus radiodurans Species 0.000 description 1
- 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 1
- 102000005744 Glycoside Hydrolases Human genes 0.000 description 1
- 108010031186 Glycoside Hydrolases Proteins 0.000 description 1
- 241001597008 Nomeidae Species 0.000 description 1
- 238000012408 PCR amplification Methods 0.000 description 1
- 244000062793 Sorghum vulgare Species 0.000 description 1
- 229920002472 Starch Polymers 0.000 description 1
- 229930006000 Sucrose Natural products 0.000 description 1
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 description 1
- 239000012505 Superdex™ Substances 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 230000004913 activation Effects 0.000 description 1
- 102000004139 alpha-Amylases Human genes 0.000 description 1
- 108090000637 alpha-Amylases Proteins 0.000 description 1
- 229940024171 alpha-amylase Drugs 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 238000009835 boiling Methods 0.000 description 1
- 239000006227 byproduct Substances 0.000 description 1
- 238000004364 calculation method Methods 0.000 description 1
- 239000003054 catalyst Substances 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 238000012512 characterization method Methods 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 230000004087 circulation Effects 0.000 description 1
- 238000001816 cooling Methods 0.000 description 1
- 239000002537 cosmetic Substances 0.000 description 1
- 230000018044 dehydration Effects 0.000 description 1
- 238000006297 dehydration reaction Methods 0.000 description 1
- 238000004925 denaturation Methods 0.000 description 1
- 230000036425 denaturation Effects 0.000 description 1
- 238000013461 design Methods 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 230000029087 digestion Effects 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- 150000002016 disaccharides Chemical class 0.000 description 1
- 238000001962 electrophoresis Methods 0.000 description 1
- 238000010828 elution Methods 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 239000000835 fiber Substances 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 238000001502 gel electrophoresis Methods 0.000 description 1
- 238000005227 gel permeation chromatography Methods 0.000 description 1
- 238000010353 genetic engineering Methods 0.000 description 1
- 239000008103 glucose Substances 0.000 description 1
- 229930182478 glucoside Natural products 0.000 description 1
- 150000008131 glucosides Chemical class 0.000 description 1
- 239000003292 glue Substances 0.000 description 1
- 125000003147 glycosyl group Chemical group 0.000 description 1
- 102000040353 glycosyl hydrolase 13 family Human genes 0.000 description 1
- 108091072133 glycosyl hydrolase 13 family Proteins 0.000 description 1
- 239000001963 growth medium Substances 0.000 description 1
- XLYOFNOQVPJJNP-ZSJDYOACSA-N heavy water Substances [2H]O[2H] XLYOFNOQVPJJNP-ZSJDYOACSA-N 0.000 description 1
- 230000002209 hydrophobic effect Effects 0.000 description 1
- 230000008676 import Effects 0.000 description 1
- 238000011534 incubation Methods 0.000 description 1
- 230000006698 induction Effects 0.000 description 1
- 239000002054 inoculum Substances 0.000 description 1
- 238000007689 inspection Methods 0.000 description 1
- 238000011835 investigation Methods 0.000 description 1
- 150000002500 ions Chemical class 0.000 description 1
- 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 1
- 238000004811 liquid chromatography Methods 0.000 description 1
- 125000003071 maltose group Chemical group 0.000 description 1
- FYGDTMLNYKFZSV-UHFFFAOYSA-N mannotriose Natural products OC1C(O)C(O)C(CO)OC1OC1C(CO)OC(OC2C(OC(O)C(O)C2O)CO)C(O)C1O FYGDTMLNYKFZSV-UHFFFAOYSA-N 0.000 description 1
- 238000011169 microbiological contamination Methods 0.000 description 1
- 238000000386 microscopy Methods 0.000 description 1
- 235000019713 millet Nutrition 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- 238000005457 optimization Methods 0.000 description 1
- 239000008363 phosphate buffer Substances 0.000 description 1
- 238000002264 polyacrylamide gel electrophoresis Methods 0.000 description 1
- 238000002360 preparation method Methods 0.000 description 1
- 239000003223 protective agent Substances 0.000 description 1
- 238000002331 protein detection Methods 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 238000001448 refractive index detection Methods 0.000 description 1
- 238000012163 sequencing technique Methods 0.000 description 1
- 238000012807 shake-flask culturing Methods 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 239000008107 starch Substances 0.000 description 1
- 235000019698 starch Nutrition 0.000 description 1
- 239000010902 straw Substances 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 239000005720 sucrose Substances 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- 210000004243 sweat Anatomy 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 238000012546 transfer Methods 0.000 description 1
- 230000009466 transformation Effects 0.000 description 1
- NMXLJRHBJVMYPD-IPFGBZKGSA-N trehalulose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@]1(O)CO[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 NMXLJRHBJVMYPD-IPFGBZKGSA-N 0.000 description 1
- 201000008827 tuberculosis Diseases 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
- 210000005253 yeast cell Anatomy 0.000 description 1
- FYGDTMLNYKFZSV-BYLHFPJWSA-N β-1,4-galactotrioside Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@H]1O[C@@H]1[C@H](CO)O[C@@H](O[C@@H]2[C@@H](O[C@@H](O)[C@H](O)[C@H]2O)CO)[C@H](O)[C@H]1O FYGDTMLNYKFZSV-BYLHFPJWSA-N 0.000 description 1
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
- C12N9/10—Transferases (2.)
- C12N9/1048—Glycosyltransferases (2.4)
- C12N9/1051—Hexosyltransferases (2.4.1)
-
- 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
- C12P19/00—Preparation of compounds containing saccharide radicals
- C12P19/12—Disaccharides
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Y—ENZYMES
- C12Y204/00—Glycosyltransferases (2.4)
- C12Y204/01—Hexosyltransferases (2.4.1)
- C12Y204/01245—Alpha,alpha-trehalose synthase (2.4.1.245)
Landscapes
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Zoology (AREA)
- Health & Medical Sciences (AREA)
- Wood Science & Technology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Genetics & Genomics (AREA)
- General Health & Medical Sciences (AREA)
- General Engineering & Computer Science (AREA)
- Biochemistry (AREA)
- Biotechnology (AREA)
- Microbiology (AREA)
- General Chemical & Material Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Medicinal Chemistry (AREA)
- Molecular Biology (AREA)
- Biomedical Technology (AREA)
- Enzymes And Modification Thereof (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention discloses a trehalose synthase mutant and a gene thereof. The enzyme is formed through mutation of mycobacterium smegmatis trehalose synthase. Particularly, amino acids at the 167th site, the 169th site, the 174th site, the 175th site and the 176th site of the emzyme are mutated into alanine, wherein the alanine mutated from the amino acids at the 169th site is freely selected to be combined with the alanine mutated from the amino acids at the 167th site, the 174th site, the 175th site and the 176th site. After mutation, compared with a wild type enzyme, enzyme activity is improved by 2.56 times at most.
Description
Technical field
The invention belongs to biological technical field, specifically, it is related to enzyme engineering field, a kind of mutation of trehalose synthetase
Body, gene, recombinant vector, transgenic cell and, application in catalyzing and synthesizing trehalose for the compositionss.
Background technology
Trehalose（Trehalose）, molecular formula is C12H22O11•2H2O, is two glucose molecules with α, α -1,1 glucosides
The disaccharide that key combines to form；Trehalose no hemiacetal hydroxyl, is nonreducing sugar, stable chemical nature.Because trehalose is in height
Warm, high and cold, the hyperosmosises and protecting film that the severe environmental conditions such as dehydration under can in cell surface form uniqueness is dried, effectively
Protected protein matter molecule invariance inactivates, and is therefore described as " sugar of life ".This characteristic is so that trehalose is except can conduct
Beyond the activity protecting agent of biological product and holding cytoactive, also there is great use in cosmetics and food, therefore
There is wide market prospect and using value.
At present, in all methods producing trehalose, simplest technique is：Trehalose synthetase（Trehalose
synthase）With maltose as substrate, glycosyl one step catalytic reaction is turned by intramolecular and directly generates trehalose.Reaction need not in
Intermediate step, low raw-material cost, process is simple, there is very high industrial application value；But trehalose synthetase also has and partly lacks
Point, mainly has that conversion ratio is low, by-product yield high, temperature and pH poor resistance etc..
Using genetic engineering and enzyme engineering means, improve the activity of trehalose synthetase, and its resistance characteristics, improve catalysis
In synthetic reaction, trehalose efficiency is to realize the only way that trehalose efficiently produces.
It was demonstrated that trehalose synthetase is glycosyl hydrolase 13 family in the further investigation to trehalose synthetase at present
（Glycoside hydrolase family 13, abbreviation Family GH l3, also referred to as alpha-amylase family）One of member.Should
The enzyme of family all has one typical (beta/alpha)8Barreled catalyst structure domain.Have been found that the trehalose of three kinds of separate sources closes
Become enzymatic structure be resolved out, respectively fromMycobacterium tuberculosis、 Deinococcusradiodurans、Mycobacterium smegmatisThree kinds of different bacterium, its PDB numbering is respectively：
4LXF、4TVU、3ZO.The comparison according to different structure for the inventor and the hydrophilic and hydrophobic of aminoacid, in the site attempting zones of different
After mutation, after finding mutation, enzyme is significantly improved compared to former wild type, completes the present invention program based on this.
Content of the invention
The technical problem to be solved is, existing comes from mycobacterium smegmatisMycobacterium smegmatisTrehalose synthetase activity too low, in order to improve the activity of this enzyme, the invention provides from shame dirt branch
BacillusMycobacterium smegmatisMutant of trehalose synthetase, the aminoacid sequence such as SEQ ID NO of this enzyme：
2nd, shown in 4,6,8 or 10.
In the present invention, inventor attempt zones of different carry out this enzyme site mutation it is found that the 167th, the
The amino acid mutation of 169, the 174th, the 175th and the 176th is alanine；After mutation enzyme have compared to former wild type bright
Aobvious raising, completes the present invention program based on this.
Specifically, the solution of the present invention is：
Will be from mycobacterium smegmatisMycobacterium smegmatisThe 167th of wild type trehalose synthetase,
The amino acid mutation of 169, the 174th, the 175th and the 176th is alanine；Wherein, optionally the 169th amino acids are dashed forward
It is changed into alanine and the amino acid mutation of the 167th, the 174th, the 175th and the 176th carries out mutation combination for alanine；
Obtain R169A, R169A-D174A, R169A-D174A-D167A, R169A-D174A-D167A-T175A, R169A-
Five mutants of D174A-D167A-T175A-E176A.
Preferably the 169th amino acids are sported alanine.
Accordingly, the nucleotide sequence of this enzyme mutant gene such as SEQ ID NO：:1st, shown in 3,5,7 or 9.
Recombinant vector of the present invention, includes above-mentioned gene order, imports this in existing carrier, plasmid
Gene order in invention, you can to realize.
Transgenic cell containing said gene, for example, escherichia coli, yeast cells, B. subtilis cell etc..Excellent
It is selected in escherichia coli, express said gene in bacillus subtilises.
Compositionss, wherein, using any of the above-described enzyme mutant as active component in said composition, are preferred for catalyzing and synthesizing
Trehalose.
Above-mentioned enzyme mutant, gene, recombinant vector, transgenic cell and compositionss answering in catalyzing and synthesizing trehalose
With.
The beneficial effects of the present invention is, R169A, R169A-D174A, R169A-D174A-D167A, R169A-D174A-
Five mutants of D167A-T175A, R169A-D174A-D167A-T175A-E176A, the enzyme activity after mutation is respectively than wild
The enzyme activity of type improves 1.07 times, 1.05 times, 2.56 times, 2.41 times, 1.94 times.
Specific embodiment
With reference to embodiment, the present invention will be further described.Listed embodiment only demonstrates and is used, and shows this
The spirit and scope of invention are not limited to the details in this and its modification case.
The preparation of embodiment 1 mutant of trehalose synthetase
According tomycobacterium smegmatisThe gene order of the trehalose synthetase in source, respectively successively design and according to
Secondary synthesis introduces R169A, R169A-D174A, R169A-D174A-D167A, R169A-D174A-D167A-T175A, R169A-
The primer of five kinds of mutants of D174A-D167A-T175A-E176A, divides five steps to carry out rite-directed mutagenesises to trehalose synthetase successively,
Measure DNA encoding sequence.
The Arg codon identifying the 169th is changed into Ala codon.
On the basis of previous step, the 174th Asp codon is changed into Ala codon.
On the basis of previous step, the 167th Asp codon is changed into Ala codon.
On the basis of previous step, the 175th Thr codon is changed into Ala codon.
On the basis of previous step, the 176th Glu codon is changed into Ala codon.
Mutant gene is connected suitable expression vector importing in escherichia coli expressed, obtain five kinds of differences and dash forward
The trehalose synthetase becoming.
Specifically,
The first step：With original plasmid TreS-pET22b as template, the rite-directed mutagenesis primer introducing R169A mutation is：
Forward primer：5-aggtatcccgacgcggccatcatcttcgtcgac-3
Reverse primer：5-gtcgacgaagatgatggccgcgtcgggatacct-3
Second step：With mutant R169A as template, the rite-directed mutagenesis primer introducing D174A mutation is：
Forward primer：5-gccatcatcttcgtcgccaccgaggagtcgaac-3
Reverse primer：5-gttcgactcctcggtggcgacgaagatgatggc-3
3rd step：With mutant R169A-D174A as template, the rite-directed mutagenesis primer introducing D167A mutation is：
Forward primer：5-agcgacaggtatcccgccgcggccatcatcttc-3
Reverse primer：5-gaagatgatggccgcggcgggatacctgtcgct-3
4th step：With mutant R169A-D174A-D167A as template, the rite-directed mutagenesis primer introducing T175A mutation is：
Forward primer：5-atcatcttcgtcgccgccgaggagtcgaactgg-3
Reverse primer：5-ccagttcgactcctcggcggcgacgaagatgat-3
5th step：With mutant R169A-D174A-D167A-T175A as template, introduce the rite-directed mutagenesis primer of E176A mutation
For：
Forward primer：5-atcttcgtcgccgccgcggagtcgaactggacg-3
Reverse primer：5-cgtccagttcgactccgcggcggcgacgaagat-3.
PCR reaction system is：5 X PS buffer10 μ L, dNTPsMix 4 μ L, forward primer (10 μM) 1 μ L, instead
To primer (1 0 μM) 1 μ L, template DNA 1 μ L, PrimerStar HS (5U/ μ L) 0.5 μ L, addition distilled water to 50 μ L.
PCR amplification condition is：98 DEG C of denaturations 5min, subsequent 28 circulations（98 DEG C of 15s, 58 DEG C of 30s, 68 DEG C
6min）, 68 DEG C of extension 10min.
PCR primer digests through Dpn I, converts bacillus coli DH 5 alpha competence, and competent cell is trained in LB solid medium
After supporting overnight（Ammonia benzyl resistance）, choose monoclonal and extract plasmid after the culture of LB fluid medium, after mutant plasmid sequencing is correct, turn
Change in expressive host e. coli bl21 (DE3) competent cell.
The expression and purification of embodiment 2 trehalose synthetase
The plasmid building pET22b-TreS is proceeded in E.coliBL21 (DE3) host and is expressed.Monoclonal is taken to inoculate
It is 37 DEG C of shaken cultivation 4h in the shaking table of 200 rpm in the LB fluid medium of 100 mg/L in 5 mL ammonia benzyl concentration.To train
Foster strain transfer is in the LB fluid medium of 100 mg/L to 1L ammonia benzyl concentration, cultivates to OD600=0.6 for 37 DEG C, is cooled to
30 DEG C of addition IPTG（Final concentration 0.2 mM）Abduction delivering 14 h, 6000 rpm be centrifuged 10 min collects thallines, frozen in -80 DEG C
In refrigerator.
Take the frozen thalline of 10 g, BufferA（25mM Tris、300Mm Nacl、pH8.0）Resuspended.Using high pressure cell
Broken instrument smudge cellses, after crushing, 16000 rpm are centrifuged 30 min, collect supernatant；Secondly, Ni-NTA affinity chromatograph：First use
BufferA balances Ni-NTA post, by the slow loading of broken supernatant after centrifugation to chromatographic column, uses BufferB（300mM
Tris、300Mm Nacl、pH8.0）Wash away unconjugated albumen.Then with 5%, 10%, 20%, 100% BufferB carries out eluting
And collect and pass liquid.Elution samples determine purity of protein using SDS-PAGE, merge highly purified protein eluate, and use 30
The Millipore super filter tube of kDa is concentrated into 2 mL at 4 DEG C with 2750 rpm；Finally, divided with Superdex 200 gel chromatography
From：First using the BufferC of a column volume（25mM Tris、300Mm Nacl、2Mm DTT、pH8.0）Balanced gel post.On
Sample and collect and use AKTA Purifier FPLC, the protein sample that 2 mL are concentrated according to the speed loading of 1 mL/min,
Using BufferC eluting, fraction collector is collected according to the speed of 1 mL/min.Sample after collection is carried out using SDS-PAGE
Protein Detection, merges highly purified albumen, according to 50 μ L/ pipe volume subpackages, is subsequently stored in -80 DEG C of ice using liquid nitrogen flash freezer
In case.
Expression in bacillus subtilises for embodiment 3 mutant of trehalose synthetase
（1）The structure of bacillus subtilises expression vector
Equimolecular biologic operation is connected by double digestion, glue reclaim, T4 ligase, five kinds of mutants of trehalose synthetase
It is cloned into respectively on pMA5 carrier, obtains five kinds of different pMA5-TreS, prepare the expression in bacillus subtilises.
（2）The expression of recombinant bacterium WB600 (pMA5-tres)
The recombinant vector building is imported bacillus subtilises B. subtilis WB600 competence, by recombinant bacterium
WB600 activates, and carries out dilution spread, then chooses the recombinant bacterium that a ring single bacterium colony is of moderate size and activates further in LB,
37 DEG C, 250r/min overnight incubation.The whether microbiological contamination of next day microscopy, more in an aseptic environment, it is inoculated in LB by 3% inoculum concentration
In culture medium, 37 DEG C of culture 24h.Take 1mL bacterium solution to be centrifuged 2min in 12000r/min, and use distilled water cyclic washing
2 times, finally with the resuspended thalline of 100 μ L distilled water.Same program is wild type trehalose synthetase WB600 (pMA5-
Tres) comparison and blank bacterium WB600 comparison.Respectively take recombinant bacterium and blank bacterium suspension 10 μ L, delay with 10 μ L loadings respectively
Rush liquid mixing, boiling water bath 10min, carry out PAGE gel electrophoresis.Observe expression in millet straw bacillus cereuss for each mutant
Situation.Electrophoresis result shows, expression effect in bacillus subtilises is consistent with wild type trehalose synthetase for mutant.
The mensure of embodiment 4 trehalose synthetase enzyme activity
（1）Enzyme-activity unit defines：At 25 DEG C, the amount that 1min generates the trehalose synthetase needed for 1 μm of oL trehalose is not had to be one
Enzyme activity unit.
（2）Enzyme activity determination step：Take maltose reagent 0.54g, be dissolved in the phosphate buffer of 10ml, pH=7.4 so as to
Final concentration of 150mmol/L.800 μ l maltose solutions and 200 μ l crude enzyme liquids mix（Every group three parallel）, 1ml reaction system
At 25 DEG C, react 30min, boil 10min enzyme denaturing.After its cooling, 5000r/min is centrifuged 10min, takes supernatant, measures life
The content of trehalose becoming.
（3）High effective liquid chromatography for measuring product and substrate：
Condition determination：Mobile phase is acetonitrile：Water=75：25；Flow velocity：0.6ml/min；Chromatographic column selects nh 2 column（35 DEG C of column temperature）；
From ShodexRI101 refractive index detection device.
（4）The shake-flask culture 16h of wild type trehalose synthetase and mutant enzyme, through high performance liquid chromatography detection and meter
Calculation draws enzyme activity, is listed in table 1.
Table 1 wild type trehalose synthetase and the enzyme activity of mutant enzyme
Note：Enzyme enzyme activity/U ml-1Enzyme enzyme activity/U ml-1
Test result indicate that, compared to wild type, the numbering of expression in escherichia coli be 1 R169A, in escherichia coli table
Reach numbering be 2 R169A-D174A, in bacillus subtilises expression numbering be 3 R169A-D174A-D167A, in hay
In bacillus cereuss, expression numbering is 4 R169A-D174A-D167A-T175A, to number in expression in escherichia coli be 5
R169A-D174A-D167A-T175A-E176A mutant enzyme, enzyme activity be respectively increased 1.07 times, 1.05 times, 2.56 times, 2.41
Again, 1.94 times.
Embodiment 5 is this example demonstrates that the characterization analysis of mutant
All reactions are all carried out according to standard reaction system：Wherein, mutant enzyme 1,2,3,4,5, corresponding mutant is respectively
R169A、R169A-D174A、R169A-D174A-D167A、R169A-D174A-D167A-T175A、R169A-D174A-
D167A-T175A-E176A.
（1）The optimum pH of enzyme measures：Delay in a series of PBS of differences pH value (5.5,6.0,6.5,7.0,7.5,8.0)
Rush the impact measuring in liquid to recombinase enzyme activity.Product content highest pH value is the optimum pH of this enzyme.
（2）Optimal reactive temperature measures：Respectively measure recombinase series of temperature (10 DEG C, 15 DEG C, 20 DEG C, 25 DEG C, 30
DEG C, 35 DEG C, 40 DEG C, the optimal reactive temperature to determine enzyme for the enzyme activity at 45 DEG C.
（3）Under optimum temperature and pH value, measure Fe3+、Cu2+、Zn2+、Fe2+、Ni2+、Mn2+Etc. each metal ion species to enzyme
The impact lived, final concentration of 5 mM of metal ion in reaction system, with the PBS that is not added with other metal ions for comparison
(100% enzyme activity).
（4）Under optimum reaction conditionses, measure the product composition of substrate for enzymatic activity of recombinating different action times.
（5）Under optimum reaction conditionses, (substrate includes trehalose, sucrose, fiber two to the substrate specificity of mensure recombinase
Sugar, starch, isomaltulose, trehalulose, maltotriose etc.).After sample reacts 5 h under optimal condition, detected with HPLC
Product.
Test result indicate that, wild enzyme, the optimum pH of the enzyme of each mutant respectively are 7.2,7.3,7.2,
7.0、7.0、6.9；Optimal reactive temperature respectively is 28 DEG C, 28 DEG C, 27.5 DEG C, 28 DEG C, 29 DEG C, 29 DEG C；Metal ion pair
The impact of mutant enzyme, experiment shows Ni2+、Fe3+、Mn2+Deng 8 metal ion species to wild type trehalose synthetase and its mutation
The enzyme activity of body does not all have activation, and the inhibitory action to enzyme activity of most metal ion is also inconspicuous, wherein Cu2+
The enzyme activity of ion pair wild type and mutant all has strong inhibitory action；In terms of substrate specificity, according to the inspection of HPLC
Survey data, the most suitable substrate of wild type and mutant enzyme is maltose.
Comprehensive above zymetology qualitative experiment result shows, the zymologic property of each mutant is all similar to wild enzyme.
Fermentation condition optimization in escherichia coli for embodiment 6 mutant of trehalose synthetase
Using single factor experiment and orthogonal test, study bacterium when adding derivant in mutant of trehalose synthetase sweat dense
OD600 nm, inducing temperature, induction time, trehalose synthetase conversion 4 factors of trehalose time are to fermentation yield and conversion
The impact of rate, determines optimum fermentation condition in escherichia coli for the mutant of trehalose synthetase and conversion condition.Result shows,
The impact to conversion ratio of transformation time and inducing temperature is larger, adds when Escherichia coli Growth to bacterium OD600 nm value is for 0.8 and lures
Lead agent, 37 DEG C of holding 14 h of inducing temperature, in conversion process, 37 DEG C, pH7.2, time be conversion ratio highest during 3h.Here is
Under the conditions of good, the conversion ratio that different mutant of trehalose synthetase convert maltose generation trehalose reaches as high as 69.29%.
Sequence table
<110>Nanjing University of Technology
Shanghai Organic Chemistry Institute, Chinese Academy of Sciences
<120>Mutant of trehalose synthetase and its gene
<130> xb16100802
<160> 10
<170> PatentIn version 3.3
<210> 1
<211> 1782
<212> DNA
<213> Artificial
<220>
<223>Mutant enzyme 1
<220>
<221> CDS
<222> (1)..(1782)
<400> 1
atg gag gag cac acg cag ggc agc cat gtc gag gcg ggg atc gtc gag 48
Met Glu Glu His Thr Gln Gly Ser His Val Glu Ala Gly Ile Val Glu
1 5 10 15
cat ccg aac gcc gag gac ttc ggt cat gcg cgc acg ctt ccc acc gac 96
His Pro Asn Ala Glu Asp Phe Gly His Ala Arg Thr Leu Pro Thr Asp
20 25 30
acc aac tgg ttc aag cac gcg gtg ttc tat gag gtg ctg gtg cgg gcg 144
Thr Asn Trp Phe Lys His Ala Val Phe Tyr Glu Val Leu Val Arg Ala
35 40 45
ttc tac gac tcg aac gcc gac ggt atc ggt gat ctg cgc ggc ctc acc 192
Phe Tyr Asp Ser Asn Ala Asp Gly Ile Gly Asp Leu Arg Gly Leu Thr
50 55 60
gaa aaa ctc gac tac atc aag tgg ctc ggg gtc gac tgc ctg tgg ctg 240
Glu Lys Leu Asp Tyr Ile Lys Trp Leu Gly Val Asp Cys Leu Trp Leu
65 70 75 80
ccg ccg ttc tac gat tcg ccg ctg cgc gac ggc ggt tac gac atc cgc 288
Pro Pro Phe Tyr Asp Ser Pro Leu Arg Asp Gly Gly Tyr Asp Ile Arg
85 90 95
gac ttc tac aag gtg ctg ccc gag ttc ggc acg gtc gac gac ttc gtc 336
Asp Phe Tyr Lys Val Leu Pro Glu Phe Gly Thr Val Asp Asp Phe Val
100 105 110
acc ctg ctg gac gcg gcc cac cgc agg ggc atc cgc atc atc acc gac 384
Thr Leu Leu Asp Ala Ala His Arg Arg Gly Ile Arg Ile Ile Thr Asp
115 120 125
ctg gtg atg aac cac acc tcc gat cag cac gag tgg ttc cag gaa tcc 432
Leu Val Met Asn His Thr Ser Asp Gln His Glu Trp Phe Gln Glu Ser
130 135 140
cgg cac aat ccc gac ggc ccc tac ggc gac ttc tac gtc tgg agc gac 480
Arg His Asn Pro Asp Gly Pro Tyr Gly Asp Phe Tyr Val Trp Ser Asp
145 150 155 160
acc agc gac agg tat ccc gac gcg gcc atc atc ttc gtc gac acc gag 528
Thr Ser Asp Arg Tyr Pro Asp Ala Ala Ile Ile Phe Val Asp Thr Glu
165 170 175
gag tcg aac tgg acg ttc gac ccc gtc cgc agg cag ttc tac tgg cac 576
Glu Ser Asn Trp Thr Phe Asp Pro Val Arg Arg Gln Phe Tyr Trp His
180 185 190
cgc ttc ttc agc cac cag ccc gac ctc aac tac gac aac ccg gct gtg 624
Arg Phe Phe Ser His Gln Pro Asp Leu Asn Tyr Asp Asn Pro Ala Val
195 200 205
cag gag gcg atg ctc gac gtg ctg agg ttc tgg ctg gac ctc ggt atc 672
Gln Glu Ala Met Leu Asp Val Leu Arg Phe Trp Leu Asp Leu Gly Ile
210 215 220
gac ggt ttc cgt ctg gac gcg gtg ccg tac ctg ttc gag cgc gag ggc 720
Asp Gly Phe Arg Leu Asp Ala Val Pro Tyr Leu Phe Glu Arg Glu Gly
225 230 235 240
acc aac tgc gag aac ctg ccc gag acc cac gcg ttc ctc aag cgg tgc 768
Thr Asn Cys Glu Asn Leu Pro Glu Thr His Ala Phe Leu Lys Arg Cys
245 250 255
cgc aag gcg atc gac gac gag tac ccg ggg cgc gtg ctg ctg gcc gag 816
Arg Lys Ala Ile Asp Asp Glu Tyr Pro Gly Arg Val Leu Leu Ala Glu
260 265 270
gcc aat cag tgg ccc gcc gac gtc gtc gcg tac ttc ggc gac ccg gac 864
Ala Asn Gln Trp Pro Ala Asp Val Val Ala Tyr Phe Gly Asp Pro Asp
275 280 285
acc ggc ggc gac gaa tgc cac atg gcg ttc cac ttc ccg ctg atg cca 912
Thr Gly Gly Asp Glu Cys His Met Ala Phe His Phe Pro Leu Met Pro
290 295 300
cgg atc ttc atg gcc gtg cgg cgc gag tcc cgc ttc ccg atc tcg gag 960
Arg Ile Phe Met Ala Val Arg Arg Glu Ser Arg Phe Pro Ile Ser Glu
305 310 315 320
atc ctg gcg cag acc ccg ccg atc ccc gac acc gcg cag tgg ggc atc 1008
Ile Leu Ala Gln Thr Pro Pro Ile Pro Asp Thr Ala Gln Trp Gly Ile
325 330 335
ttc ctg cgc aac cac gac gag ctc acg ctc gag atg gtc acc gac gaa 1056
Phe Leu Arg Asn His Asp Glu Leu Thr Leu Glu Met Val Thr Asp Glu
340 345 350
gaa cgt gac tac atg tac gcc gag tac gcc aag gac ccg cgg atg aag 1104
Glu Arg Asp Tyr Met Tyr Ala Glu Tyr Ala Lys Asp Pro Arg Met Lys
355 360 365
gcc aac gtc ggc atc cgg cgc agg ctc gca ccg ctg ttg gag aac gac 1152
Ala Asn Val Gly Ile Arg Arg Arg Leu Ala Pro Leu Leu Glu Asn Asp
370 375 380
cgc aac cag atc gag ttg ttc acc gcg ctg ctg ctc tcg ctg ccg ggg 1200
Arg Asn Gln Ile Glu Leu Phe Thr Ala Leu Leu Leu Ser Leu Pro Gly
385 390 395 400
tcg ccg gtg ctg tac tac ggt gac gag ata ggt atg ggg gac atc atc 1248
Ser Pro Val Leu Tyr Tyr Gly Asp Glu Ile Gly Met Gly Asp Ile Ile
405 410 415
tgg ctc ggc gac cgc gac agc gtg cgt acg ccc atg cag tgg acc ccc 1296
Trp Leu Gly Asp Arg Asp Ser Val Arg Thr Pro Met Gln Trp Thr Pro
420 425 430
gac cgc aat gcg ggc ttc tcc aag gcg act ccg ggc cgg ctc tac ctg 1344
Asp Arg Asn Ala Gly Phe Ser Lys Ala Thr Pro Gly Arg Leu Tyr Leu
435 440 445
ccg ccc aac cag gac gcg gtg tac ggc tac cac tct gtg aac gtc gag 1392
Pro Pro Asn Gln Asp Ala Val Tyr Gly Tyr His Ser Val Asn Val Glu
450 455 460
gcg caa ctc gac agc tcc agc tcg ctg ttg aac tgg acg cgc aac atg 1440
Ala Gln Leu Asp Ser Ser Ser Ser Leu Leu Asn Trp Thr Arg Asn Met
465 470 475 480
ctg gcc gtg cgc agc cgc cac gat gcc ttc gca gtc ggc acg ttc cgc 1488
Leu Ala Val Arg Ser Arg His Asp Ala Phe Ala Val Gly Thr Phe Arg
485 490 495
gaa ctg ggc ggc tcc aat ccg tcg gtg ctg gcg tac atc cgg gag gtc 1536
Glu Leu Gly Gly Ser Asn Pro Ser Val Leu Ala Tyr Ile Arg Glu Val
500 505 510
acg cgc caa cag ggc gac ggc ggc gcg aag acc gat gcc gtc ctg tgt 1584
Thr Arg Gln Gln Gly Asp Gly Gly Ala Lys Thr Asp Ala Val Leu Cys
515 520 525
gtc aac aac ctg tcc cgc ttc ccc cag ccc atc gag ttg aat ctc cag 1632
Val Asn Asn Leu Ser Arg Phe Pro Gln Pro Ile Glu Leu Asn Leu Gln
530 535 540
cag tgg gcc ggt tac ata ccc gtc gag atg acc ggc tac gtc gag ttc 1680
Gln Trp Ala Gly Tyr Ile Pro Val Glu Met Thr Gly Tyr Val Glu Phe
545 550 555 560
ccg agt atc ggc cag ttg ccg tac ctg ctc acc ctg ccc ggt cac ggg 1728
Pro Ser Ile Gly Gln Leu Pro Tyr Leu Leu Thr Leu Pro Gly His Gly
565 570 575
ttc tac tgg ttc cag ctt cga gaa ccc gat cca gaa ccg gga gcg cag 1776
Phe Tyr Trp Phe Gln Leu Arg Glu Pro Asp Pro Glu Pro Gly Ala Gln
580 585 590
caa tga 1782
Gln
<210> 2
<211> 593
<212> PRT
<213> Artificial
<220>
<223> Synthetic Construct
<400> 2
Met Glu Glu His Thr Gln Gly Ser His Val Glu Ala Gly Ile Val Glu
1 5 10 15
His Pro Asn Ala Glu Asp Phe Gly His Ala Arg Thr Leu Pro Thr Asp
20 25 30
Thr Asn Trp Phe Lys His Ala Val Phe Tyr Glu Val Leu Val Arg Ala
35 40 45
Phe Tyr Asp Ser Asn Ala Asp Gly Ile Gly Asp Leu Arg Gly Leu Thr
50 55 60
Glu Lys Leu Asp Tyr Ile Lys Trp Leu Gly Val Asp Cys Leu Trp Leu
65 70 75 80
Pro Pro Phe Tyr Asp Ser Pro Leu Arg Asp Gly Gly Tyr Asp Ile Arg
85 90 95
Asp Phe Tyr Lys Val Leu Pro Glu Phe Gly Thr Val Asp Asp Phe Val
100 105 110
Thr Leu Leu Asp Ala Ala His Arg Arg Gly Ile Arg Ile Ile Thr Asp
115 120 125
Leu Val Met Asn His Thr Ser Asp Gln His Glu Trp Phe Gln Glu Ser
130 135 140
Arg His Asn Pro Asp Gly Pro Tyr Gly Asp Phe Tyr Val Trp Ser Asp
145 150 155 160
Thr Ser Asp Arg Tyr Pro Asp Ala Ala Ile Ile Phe Val Asp Thr Glu
165 170 175
Glu Ser Asn Trp Thr Phe Asp Pro Val Arg Arg Gln Phe Tyr Trp His
180 185 190
Arg Phe Phe Ser His Gln Pro Asp Leu Asn Tyr Asp Asn Pro Ala Val
195 200 205
Gln Glu Ala Met Leu Asp Val Leu Arg Phe Trp Leu Asp Leu Gly Ile
210 215 220
Asp Gly Phe Arg Leu Asp Ala Val Pro Tyr Leu Phe Glu Arg Glu Gly
225 230 235 240
Thr Asn Cys Glu Asn Leu Pro Glu Thr His Ala Phe Leu Lys Arg Cys
245 250 255
Arg Lys Ala Ile Asp Asp Glu Tyr Pro Gly Arg Val Leu Leu Ala Glu
260 265 270
Ala Asn Gln Trp Pro Ala Asp Val Val Ala Tyr Phe Gly Asp Pro Asp
275 280 285
Thr Gly Gly Asp Glu Cys His Met Ala Phe His Phe Pro Leu Met Pro
290 295 300
Arg Ile Phe Met Ala Val Arg Arg Glu Ser Arg Phe Pro Ile Ser Glu
305 310 315 320
Ile Leu Ala Gln Thr Pro Pro Ile Pro Asp Thr Ala Gln Trp Gly Ile
325 330 335
Phe Leu Arg Asn His Asp Glu Leu Thr Leu Glu Met Val Thr Asp Glu
340 345 350
Glu Arg Asp Tyr Met Tyr Ala Glu Tyr Ala Lys Asp Pro Arg Met Lys
355 360 365
Ala Asn Val Gly Ile Arg Arg Arg Leu Ala Pro Leu Leu Glu Asn Asp
370 375 380
Arg Asn Gln Ile Glu Leu Phe Thr Ala Leu Leu Leu Ser Leu Pro Gly
385 390 395 400
Ser Pro Val Leu Tyr Tyr Gly Asp Glu Ile Gly Met Gly Asp Ile Ile
405 410 415
Trp Leu Gly Asp Arg Asp Ser Val Arg Thr Pro Met Gln Trp Thr Pro
420 425 430
Asp Arg Asn Ala Gly Phe Ser Lys Ala Thr Pro Gly Arg Leu Tyr Leu
435 440 445
Pro Pro Asn Gln Asp Ala Val Tyr Gly Tyr His Ser Val Asn Val Glu
450 455 460
Ala Gln Leu Asp Ser Ser Ser Ser Leu Leu Asn Trp Thr Arg Asn Met
465 470 475 480
Leu Ala Val Arg Ser Arg His Asp Ala Phe Ala Val Gly Thr Phe Arg
485 490 495
Glu Leu Gly Gly Ser Asn Pro Ser Val Leu Ala Tyr Ile Arg Glu Val
500 505 510
Thr Arg Gln Gln Gly Asp Gly Gly Ala Lys Thr Asp Ala Val Leu Cys
515 520 525
Val Asn Asn Leu Ser Arg Phe Pro Gln Pro Ile Glu Leu Asn Leu Gln
530 535 540
Gln Trp Ala Gly Tyr Ile Pro Val Glu Met Thr Gly Tyr Val Glu Phe
545 550 555 560
Pro Ser Ile Gly Gln Leu Pro Tyr Leu Leu Thr Leu Pro Gly His Gly
565 570 575
Phe Tyr Trp Phe Gln Leu Arg Glu Pro Asp Pro Glu Pro Gly Ala Gln
580 585 590
Gln
<210> 3
<211> 1782
<212> DNA
<213> Artificial
<220>
<223>Mutant enzyme 2
<220>
<221> CDS
<222> (1)..(1782)
<400> 3
atg gag gag cac acg cag ggc agc cat gtc gag gcg ggg atc gtc gag 48
Met Glu Glu His Thr Gln Gly Ser His Val Glu Ala Gly Ile Val Glu
1 5 10 15
cat ccg aac gcc gag gac ttc ggt cat gcg cgc acg ctt ccc acc gac 96
His Pro Asn Ala Glu Asp Phe Gly His Ala Arg Thr Leu Pro Thr Asp
20 25 30
acc aac tgg ttc aag cac gcg gtg ttc tat gag gtg ctg gtg cgg gcg 144
Thr Asn Trp Phe Lys His Ala Val Phe Tyr Glu Val Leu Val Arg Ala
35 40 45
ttc tac gac tcg aac gcc gac ggt atc ggt gat ctg cgc ggc ctc acc 192
Phe Tyr Asp Ser Asn Ala Asp Gly Ile Gly Asp Leu Arg Gly Leu Thr
50 55 60
gaa aaa ctc gac tac atc aag tgg ctc ggg gtc gac tgc ctg tgg ctg 240
Glu Lys Leu Asp Tyr Ile Lys Trp Leu Gly Val Asp Cys Leu Trp Leu
65 70 75 80
ccg ccg ttc tac gat tcg ccg ctg cgc gac ggc ggt tac gac atc cgc 288
Pro Pro Phe Tyr Asp Ser Pro Leu Arg Asp Gly Gly Tyr Asp Ile Arg
85 90 95
gac ttc tac aag gtg ctg ccc gag ttc ggc acg gtc gac gac ttc gtc 336
Asp Phe Tyr Lys Val Leu Pro Glu Phe Gly Thr Val Asp Asp Phe Val
100 105 110
acc ctg ctg gac gcg gcc cac cgc agg ggc atc cgc atc atc acc gac 384
Thr Leu Leu Asp Ala Ala His Arg Arg Gly Ile Arg Ile Ile Thr Asp
115 120 125
ctg gtg atg aac cac acc tcc gat cag cac gag tgg ttc cag gaa tcc 432
Leu Val Met Asn His Thr Ser Asp Gln His Glu Trp Phe Gln Glu Ser
130 135 140
cgg cac aat ccc gac ggc ccc tac ggc gac ttc tac gtc tgg agc gac 480
Arg His Asn Pro Asp Gly Pro Tyr Gly Asp Phe Tyr Val Trp Ser Asp
145 150 155 160
acc agc gac agg tat ccc gac gcg gcc atc atc ttc gtc gcc acc gag 528
Thr Ser Asp Arg Tyr Pro Asp Ala Ala Ile Ile Phe Val Ala Thr Glu
165 170 175
gag tcg aac tgg acg ttc gac ccc gtc cgc agg cag ttc tac tgg cac 576
Glu Ser Asn Trp Thr Phe Asp Pro Val Arg Arg Gln Phe Tyr Trp His
180 185 190
cgc ttc ttc agc cac cag ccc gac ctc aac tac gac aac ccg gct gtg 624
Arg Phe Phe Ser His Gln Pro Asp Leu Asn Tyr Asp Asn Pro Ala Val
195 200 205
cag gag gcg atg ctc gac gtg ctg agg ttc tgg ctg gac ctc ggt atc 672
Gln Glu Ala Met Leu Asp Val Leu Arg Phe Trp Leu Asp Leu Gly Ile
210 215 220
gac ggt ttc cgt ctg gac gcg gtg ccg tac ctg ttc gag cgc gag ggc 720
Asp Gly Phe Arg Leu Asp Ala Val Pro Tyr Leu Phe Glu Arg Glu Gly
225 230 235 240
acc aac tgc gag aac ctg ccc gag acc cac gcg ttc ctc aag cgg tgc 768
Thr Asn Cys Glu Asn Leu Pro Glu Thr His Ala Phe Leu Lys Arg Cys
245 250 255
cgc aag gcg atc gac gac gag tac ccg ggg cgc gtg ctg ctg gcc gag 816
Arg Lys Ala Ile Asp Asp Glu Tyr Pro Gly Arg Val Leu Leu Ala Glu
260 265 270
gcc aat cag tgg ccc gcc gac gtc gtc gcg tac ttc ggc gac ccg gac 864
Ala Asn Gln Trp Pro Ala Asp Val Val Ala Tyr Phe Gly Asp Pro Asp
275 280 285
acc ggc ggc gac gaa tgc cac atg gcg ttc cac ttc ccg ctg atg cca 912
Thr Gly Gly Asp Glu Cys His Met Ala Phe His Phe Pro Leu Met Pro
290 295 300
cgg atc ttc atg gcc gtg cgg cgc gag tcc cgc ttc ccg atc tcg gag 960
Arg Ile Phe Met Ala Val Arg Arg Glu Ser Arg Phe Pro Ile Ser Glu
305 310 315 320
atc ctg gcg cag acc ccg ccg atc ccc gac acc gcg cag tgg ggc atc 1008
Ile Leu Ala Gln Thr Pro Pro Ile Pro Asp Thr Ala Gln Trp Gly Ile
325 330 335
ttc ctg cgc aac cac gac gag ctc acg ctc gag atg gtc acc gac gaa 1056
Phe Leu Arg Asn His Asp Glu Leu Thr Leu Glu Met Val Thr Asp Glu
340 345 350
gaa cgt gac tac atg tac gcc gag tac gcc aag gac ccg cgg atg aag 1104
Glu Arg Asp Tyr Met Tyr Ala Glu Tyr Ala Lys Asp Pro Arg Met Lys
355 360 365
gcc aac gtc ggc atc cgg cgc agg ctc gca ccg ctg ttg gag aac gac 1152
Ala Asn Val Gly Ile Arg Arg Arg Leu Ala Pro Leu Leu Glu Asn Asp
370 375 380
cgc aac cag atc gag ttg ttc acc gcg ctg ctg ctc tcg ctg ccg ggg 1200
Arg Asn Gln Ile Glu Leu Phe Thr Ala Leu Leu Leu Ser Leu Pro Gly
385 390 395 400
tcg ccg gtg ctg tac tac ggt gac gag ata ggt atg ggg gac atc atc 1248
Ser Pro Val Leu Tyr Tyr Gly Asp Glu Ile Gly Met Gly Asp Ile Ile
405 410 415
tgg ctc ggc gac cgc gac agc gtg cgt acg ccc atg cag tgg acc ccc 1296
Trp Leu Gly Asp Arg Asp Ser Val Arg Thr Pro Met Gln Trp Thr Pro
420 425 430
gac cgc aat gcg ggc ttc tcc aag gcg act ccg ggc cgg ctc tac ctg 1344
Asp Arg Asn Ala Gly Phe Ser Lys Ala Thr Pro Gly Arg Leu Tyr Leu
435 440 445
ccg ccc aac cag gac gcg gtg tac ggc tac cac tct gtg aac gtc gag 1392
Pro Pro Asn Gln Asp Ala Val Tyr Gly Tyr His Ser Val Asn Val Glu
450 455 460
gcg caa ctc gac agc tcc agc tcg ctg ttg aac tgg acg cgc aac atg 1440
Ala Gln Leu Asp Ser Ser Ser Ser Leu Leu Asn Trp Thr Arg Asn Met
465 470 475 480
ctg gcc gtg cgc agc cgc cac gat gcc ttc gca gtc ggc acg ttc cgc 1488
Leu Ala Val Arg Ser Arg His Asp Ala Phe Ala Val Gly Thr Phe Arg
485 490 495
gaa ctg ggc ggc tcc aat ccg tcg gtg ctg gcg tac atc cgg gag gtc 1536
Glu Leu Gly Gly Ser Asn Pro Ser Val Leu Ala Tyr Ile Arg Glu Val
500 505 510
acg cgc caa cag ggc gac ggc ggc gcg aag acc gat gcc gtc ctg tgt 1584
Thr Arg Gln Gln Gly Asp Gly Gly Ala Lys Thr Asp Ala Val Leu Cys
515 520 525
gtc aac aac ctg tcc cgc ttc ccc cag ccc atc gag ttg aat ctc cag 1632
Val Asn Asn Leu Ser Arg Phe Pro Gln Pro Ile Glu Leu Asn Leu Gln
530 535 540
cag tgg gcc ggt tac ata ccc gtc gag atg acc ggc tac gtc gag ttc 1680
Gln Trp Ala Gly Tyr Ile Pro Val Glu Met Thr Gly Tyr Val Glu Phe
545 550 555 560
ccg agt atc ggc cag ttg ccg tac ctg ctc acc ctg ccc ggt cac ggg 1728
Pro Ser Ile Gly Gln Leu Pro Tyr Leu Leu Thr Leu Pro Gly His Gly
565 570 575
ttc tac tgg ttc cag ctt cga gaa ccc gat cca gaa ccg gga gcg cag 1776
Phe Tyr Trp Phe Gln Leu Arg Glu Pro Asp Pro Glu Pro Gly Ala Gln
580 585 590
caa tga 1782
Gln
<210> 4
<211> 593
<212> PRT
<213> Artificial
<220>
<223> Synthetic Construct
<400> 4
Met Glu Glu His Thr Gln Gly Ser His Val Glu Ala Gly Ile Val Glu
1 5 10 15
His Pro Asn Ala Glu Asp Phe Gly His Ala Arg Thr Leu Pro Thr Asp
20 25 30
Thr Asn Trp Phe Lys His Ala Val Phe Tyr Glu Val Leu Val Arg Ala
35 40 45
Phe Tyr Asp Ser Asn Ala Asp Gly Ile Gly Asp Leu Arg Gly Leu Thr
50 55 60
Glu Lys Leu Asp Tyr Ile Lys Trp Leu Gly Val Asp Cys Leu Trp Leu
65 70 75 80
Pro Pro Phe Tyr Asp Ser Pro Leu Arg Asp Gly Gly Tyr Asp Ile Arg
85 90 95
Asp Phe Tyr Lys Val Leu Pro Glu Phe Gly Thr Val Asp Asp Phe Val
100 105 110
Thr Leu Leu Asp Ala Ala His Arg Arg Gly Ile Arg Ile Ile Thr Asp
115 120 125
Leu Val Met Asn His Thr Ser Asp Gln His Glu Trp Phe Gln Glu Ser
130 135 140
Arg His Asn Pro Asp Gly Pro Tyr Gly Asp Phe Tyr Val Trp Ser Asp
145 150 155 160
Thr Ser Asp Arg Tyr Pro Asp Ala Ala Ile Ile Phe Val Ala Thr Glu
165 170 175
Glu Ser Asn Trp Thr Phe Asp Pro Val Arg Arg Gln Phe Tyr Trp His
180 185 190
Arg Phe Phe Ser His Gln Pro Asp Leu Asn Tyr Asp Asn Pro Ala Val
195 200 205
Gln Glu Ala Met Leu Asp Val Leu Arg Phe Trp Leu Asp Leu Gly Ile
210 215 220
Asp Gly Phe Arg Leu Asp Ala Val Pro Tyr Leu Phe Glu Arg Glu Gly
225 230 235 240
Thr Asn Cys Glu Asn Leu Pro Glu Thr His Ala Phe Leu Lys Arg Cys
245 250 255
Arg Lys Ala Ile Asp Asp Glu Tyr Pro Gly Arg Val Leu Leu Ala Glu
260 265 270
Ala Asn Gln Trp Pro Ala Asp Val Val Ala Tyr Phe Gly Asp Pro Asp
275 280 285
Thr Gly Gly Asp Glu Cys His Met Ala Phe His Phe Pro Leu Met Pro
290 295 300
Arg Ile Phe Met Ala Val Arg Arg Glu Ser Arg Phe Pro Ile Ser Glu
305 310 315 320
Ile Leu Ala Gln Thr Pro Pro Ile Pro Asp Thr Ala Gln Trp Gly Ile
325 330 335
Phe Leu Arg Asn His Asp Glu Leu Thr Leu Glu Met Val Thr Asp Glu
340 345 350
Glu Arg Asp Tyr Met Tyr Ala Glu Tyr Ala Lys Asp Pro Arg Met Lys
355 360 365
Ala Asn Val Gly Ile Arg Arg Arg Leu Ala Pro Leu Leu Glu Asn Asp
370 375 380
Arg Asn Gln Ile Glu Leu Phe Thr Ala Leu Leu Leu Ser Leu Pro Gly
385 390 395 400
Ser Pro Val Leu Tyr Tyr Gly Asp Glu Ile Gly Met Gly Asp Ile Ile
405 410 415
Trp Leu Gly Asp Arg Asp Ser Val Arg Thr Pro Met Gln Trp Thr Pro
420 425 430
Asp Arg Asn Ala Gly Phe Ser Lys Ala Thr Pro Gly Arg Leu Tyr Leu
435 440 445
Pro Pro Asn Gln Asp Ala Val Tyr Gly Tyr His Ser Val Asn Val Glu
450 455 460
Ala Gln Leu Asp Ser Ser Ser Ser Leu Leu Asn Trp Thr Arg Asn Met
465 470 475 480
Leu Ala Val Arg Ser Arg His Asp Ala Phe Ala Val Gly Thr Phe Arg
485 490 495
Glu Leu Gly Gly Ser Asn Pro Ser Val Leu Ala Tyr Ile Arg Glu Val
500 505 510
Thr Arg Gln Gln Gly Asp Gly Gly Ala Lys Thr Asp Ala Val Leu Cys
515 520 525
Val Asn Asn Leu Ser Arg Phe Pro Gln Pro Ile Glu Leu Asn Leu Gln
530 535 540
Gln Trp Ala Gly Tyr Ile Pro Val Glu Met Thr Gly Tyr Val Glu Phe
545 550 555 560
Pro Ser Ile Gly Gln Leu Pro Tyr Leu Leu Thr Leu Pro Gly His Gly
565 570 575
Phe Tyr Trp Phe Gln Leu Arg Glu Pro Asp Pro Glu Pro Gly Ala Gln
580 585 590
Gln
<210> 5
<211> 1782
<212> DNA
<213> Artificial
<220>
<223>Mutant enzyme 3
<220>
<221> CDS
<222> (1)..(1782)
<400> 5
atg gag gag cac acg cag ggc agc cat gtc gag gcg ggg atc gtc gag 48
Met Glu Glu His Thr Gln Gly Ser His Val Glu Ala Gly Ile Val Glu
1 5 10 15
cat ccg aac gcc gag gac ttc ggt cat gcg cgc acg ctt ccc acc gac 96
His Pro Asn Ala Glu Asp Phe Gly His Ala Arg Thr Leu Pro Thr Asp
20 25 30
acc aac tgg ttc aag cac gcg gtg ttc tat gag gtg ctg gtg cgg gcg 144
Thr Asn Trp Phe Lys His Ala Val Phe Tyr Glu Val Leu Val Arg Ala
35 40 45
ttc tac gac tcg aac gcc gac ggt atc ggt gat ctg cgc ggc ctc acc 192
Phe Tyr Asp Ser Asn Ala Asp Gly Ile Gly Asp Leu Arg Gly Leu Thr
50 55 60
gaa aaa ctc gac tac atc aag tgg ctc ggg gtc gac tgc ctg tgg ctg 240
Glu Lys Leu Asp Tyr Ile Lys Trp Leu Gly Val Asp Cys Leu Trp Leu
65 70 75 80
ccg ccg ttc tac gat tcg ccg ctg cgc gac ggc ggt tac gac atc cgc 288
Pro Pro Phe Tyr Asp Ser Pro Leu Arg Asp Gly Gly Tyr Asp Ile Arg
85 90 95
gac ttc tac aag gtg ctg ccc gag ttc ggc acg gtc gac gac ttc gtc 336
Asp Phe Tyr Lys Val Leu Pro Glu Phe Gly Thr Val Asp Asp Phe Val
100 105 110
acc ctg ctg gac gcg gcc cac cgc agg ggc atc cgc atc atc acc gac 384
Thr Leu Leu Asp Ala Ala His Arg Arg Gly Ile Arg Ile Ile Thr Asp
115 120 125
ctg gtg atg aac cac acc tcc gat cag cac gag tgg ttc cag gaa tcc 432
Leu Val Met Asn His Thr Ser Asp Gln His Glu Trp Phe Gln Glu Ser
130 135 140
cgg cac aat ccc gac ggc ccc tac ggc gac ttc tac gtc tgg agc gac 480
Arg His Asn Pro Asp Gly Pro Tyr Gly Asp Phe Tyr Val Trp Ser Asp
145 150 155 160
acc agc gac agg tat ccc gcc gcg gcc atc atc ttc gtc gcc acc gag 528
Thr Ser Asp Arg Tyr Pro Ala Ala Ala Ile Ile Phe Val Ala Thr Glu
165 170 175
gag tcg aac tgg acg ttc gac ccc gtc cgc agg cag ttc tac tgg cac 576
Glu Ser Asn Trp Thr Phe Asp Pro Val Arg Arg Gln Phe Tyr Trp His
180 185 190
cgc ttc ttc agc cac cag ccc gac ctc aac tac gac aac ccg gct gtg 624
Arg Phe Phe Ser His Gln Pro Asp Leu Asn Tyr Asp Asn Pro Ala Val
195 200 205
cag gag gcg atg ctc gac gtg ctg agg ttc tgg ctg gac ctc ggt atc 672
Gln Glu Ala Met Leu Asp Val Leu Arg Phe Trp Leu Asp Leu Gly Ile
210 215 220
gac ggt ttc cgt ctg gac gcg gtg ccg tac ctg ttc gag cgc gag ggc 720
Asp Gly Phe Arg Leu Asp Ala Val Pro Tyr Leu Phe Glu Arg Glu Gly
225 230 235 240
acc aac tgc gag aac ctg ccc gag acc cac gcg ttc ctc aag cgg tgc 768
Thr Asn Cys Glu Asn Leu Pro Glu Thr His Ala Phe Leu Lys Arg Cys
245 250 255
cgc aag gcg atc gac gac gag tac ccg ggg cgc gtg ctg ctg gcc gag 816
Arg Lys Ala Ile Asp Asp Glu Tyr Pro Gly Arg Val Leu Leu Ala Glu
260 265 270
gcc aat cag tgg ccc gcc gac gtc gtc gcg tac ttc ggc gac ccg gac 864
Ala Asn Gln Trp Pro Ala Asp Val Val Ala Tyr Phe Gly Asp Pro Asp
275 280 285
acc ggc ggc gac gaa tgc cac atg gcg ttc cac ttc ccg ctg atg cca 912
Thr Gly Gly Asp Glu Cys His Met Ala Phe His Phe Pro Leu Met Pro
290 295 300
cgg atc ttc atg gcc gtg cgg cgc gag tcc cgc ttc ccg atc tcg gag 960
Arg Ile Phe Met Ala Val Arg Arg Glu Ser Arg Phe Pro Ile Ser Glu
305 310 315 320
atc ctg gcg cag acc ccg ccg atc ccc gac acc gcg cag tgg ggc atc 1008
Ile Leu Ala Gln Thr Pro Pro Ile Pro Asp Thr Ala Gln Trp Gly Ile
325 330 335
ttc ctg cgc aac cac gac gag ctc acg ctc gag atg gtc acc gac gaa 1056
Phe Leu Arg Asn His Asp Glu Leu Thr Leu Glu Met Val Thr Asp Glu
340 345 350
gaa cgt gac tac atg tac gcc gag tac gcc aag gac ccg cgg atg aag 1104
Glu Arg Asp Tyr Met Tyr Ala Glu Tyr Ala Lys Asp Pro Arg Met Lys
355 360 365
gcc aac gtc ggc atc cgg cgc agg ctc gca ccg ctg ttg gag aac gac 1152
Ala Asn Val Gly Ile Arg Arg Arg Leu Ala Pro Leu Leu Glu Asn Asp
370 375 380
cgc aac cag atc gag ttg ttc acc gcg ctg ctg ctc tcg ctg ccg ggg 1200
Arg Asn Gln Ile Glu Leu Phe Thr Ala Leu Leu Leu Ser Leu Pro Gly
385 390 395 400
tcg ccg gtg ctg tac tac ggt gac gag ata ggt atg ggg gac atc atc 1248
Ser Pro Val Leu Tyr Tyr Gly Asp Glu Ile Gly Met Gly Asp Ile Ile
405 410 415
tgg ctc ggc gac cgc gac agc gtg cgt acg ccc atg cag tgg acc ccc 1296
Trp Leu Gly Asp Arg Asp Ser Val Arg Thr Pro Met Gln Trp Thr Pro
420 425 430
gac cgc aat gcg ggc ttc tcc aag gcg act ccg ggc cgg ctc tac ctg 1344
Asp Arg Asn Ala Gly Phe Ser Lys Ala Thr Pro Gly Arg Leu Tyr Leu
435 440 445
ccg ccc aac cag gac gcg gtg tac ggc tac cac tct gtg aac gtc gag 1392
Pro Pro Asn Gln Asp Ala Val Tyr Gly Tyr His Ser Val Asn Val Glu
450 455 460
gcg caa ctc gac agc tcc agc tcg ctg ttg aac tgg acg cgc aac atg 1440
Ala Gln Leu Asp Ser Ser Ser Ser Leu Leu Asn Trp Thr Arg Asn Met
465 470 475 480
ctg gcc gtg cgc agc cgc cac gat gcc ttc gca gtc ggc acg ttc cgc 1488
Leu Ala Val Arg Ser Arg His Asp Ala Phe Ala Val Gly Thr Phe Arg
485 490 495
gaa ctg ggc ggc tcc aat ccg tcg gtg ctg gcg tac atc cgg gag gtc 1536
Glu Leu Gly Gly Ser Asn Pro Ser Val Leu Ala Tyr Ile Arg Glu Val
500 505 510
acg cgc caa cag ggc gac ggc ggc gcg aag acc gat gcc gtc ctg tgt 1584
Thr Arg Gln Gln Gly Asp Gly Gly Ala Lys Thr Asp Ala Val Leu Cys
515 520 525
gtc aac aac ctg tcc cgc ttc ccc cag ccc atc gag ttg aat ctc cag 1632
Val Asn Asn Leu Ser Arg Phe Pro Gln Pro Ile Glu Leu Asn Leu Gln
530 535 540
cag tgg gcc ggt tac ata ccc gtc gag atg acc ggc tac gtc gag ttc 1680
Gln Trp Ala Gly Tyr Ile Pro Val Glu Met Thr Gly Tyr Val Glu Phe
545 550 555 560
ccg agt atc ggc cag ttg ccg tac ctg ctc acc ctg ccc ggt cac ggg 1728
Pro Ser Ile Gly Gln Leu Pro Tyr Leu Leu Thr Leu Pro Gly His Gly
565 570 575
ttc tac tgg ttc cag ctt cga gaa ccc gat cca gaa ccg gga gcg cag 1776
Phe Tyr Trp Phe Gln Leu Arg Glu Pro Asp Pro Glu Pro Gly Ala Gln
580 585 590
caa tga 1782
Gln
<210> 6
<211> 593
<212> PRT
<213> Artificial
<220>
<223> Synthetic Construct
<400> 6
Met Glu Glu His Thr Gln Gly Ser His Val Glu Ala Gly Ile Val Glu
1 5 10 15
His Pro Asn Ala Glu Asp Phe Gly His Ala Arg Thr Leu Pro Thr Asp
20 25 30
Thr Asn Trp Phe Lys His Ala Val Phe Tyr Glu Val Leu Val Arg Ala
35 40 45
Phe Tyr Asp Ser Asn Ala Asp Gly Ile Gly Asp Leu Arg Gly Leu Thr
50 55 60
Glu Lys Leu Asp Tyr Ile Lys Trp Leu Gly Val Asp Cys Leu Trp Leu
65 70 75 80
Pro Pro Phe Tyr Asp Ser Pro Leu Arg Asp Gly Gly Tyr Asp Ile Arg
85 90 95
Asp Phe Tyr Lys Val Leu Pro Glu Phe Gly Thr Val Asp Asp Phe Val
100 105 110
Thr Leu Leu Asp Ala Ala His Arg Arg Gly Ile Arg Ile Ile Thr Asp
115 120 125
Leu Val Met Asn His Thr Ser Asp Gln His Glu Trp Phe Gln Glu Ser
130 135 140
Arg His Asn Pro Asp Gly Pro Tyr Gly Asp Phe Tyr Val Trp Ser Asp
145 150 155 160
Thr Ser Asp Arg Tyr Pro Ala Ala Ala Ile Ile Phe Val Ala Thr Glu
165 170 175
Glu Ser Asn Trp Thr Phe Asp Pro Val Arg Arg Gln Phe Tyr Trp His
180 185 190
Arg Phe Phe Ser His Gln Pro Asp Leu Asn Tyr Asp Asn Pro Ala Val
195 200 205
Gln Glu Ala Met Leu Asp Val Leu Arg Phe Trp Leu Asp Leu Gly Ile
210 215 220
Asp Gly Phe Arg Leu Asp Ala Val Pro Tyr Leu Phe Glu Arg Glu Gly
225 230 235 240
Thr Asn Cys Glu Asn Leu Pro Glu Thr His Ala Phe Leu Lys Arg Cys
245 250 255
Arg Lys Ala Ile Asp Asp Glu Tyr Pro Gly Arg Val Leu Leu Ala Glu
260 265 270
Ala Asn Gln Trp Pro Ala Asp Val Val Ala Tyr Phe Gly Asp Pro Asp
275 280 285
Thr Gly Gly Asp Glu Cys His Met Ala Phe His Phe Pro Leu Met Pro
290 295 300
Arg Ile Phe Met Ala Val Arg Arg Glu Ser Arg Phe Pro Ile Ser Glu
305 310 315 320
Ile Leu Ala Gln Thr Pro Pro Ile Pro Asp Thr Ala Gln Trp Gly Ile
325 330 335
Phe Leu Arg Asn His Asp Glu Leu Thr Leu Glu Met Val Thr Asp Glu
340 345 350
Glu Arg Asp Tyr Met Tyr Ala Glu Tyr Ala Lys Asp Pro Arg Met Lys
355 360 365
Ala Asn Val Gly Ile Arg Arg Arg Leu Ala Pro Leu Leu Glu Asn Asp
370 375 380
Arg Asn Gln Ile Glu Leu Phe Thr Ala Leu Leu Leu Ser Leu Pro Gly
385 390 395 400
Ser Pro Val Leu Tyr Tyr Gly Asp Glu Ile Gly Met Gly Asp Ile Ile
405 410 415
Trp Leu Gly Asp Arg Asp Ser Val Arg Thr Pro Met Gln Trp Thr Pro
420 425 430
Asp Arg Asn Ala Gly Phe Ser Lys Ala Thr Pro Gly Arg Leu Tyr Leu
435 440 445
Pro Pro Asn Gln Asp Ala Val Tyr Gly Tyr His Ser Val Asn Val Glu
450 455 460
Ala Gln Leu Asp Ser Ser Ser Ser Leu Leu Asn Trp Thr Arg Asn Met
465 470 475 480
Leu Ala Val Arg Ser Arg His Asp Ala Phe Ala Val Gly Thr Phe Arg
485 490 495
Glu Leu Gly Gly Ser Asn Pro Ser Val Leu Ala Tyr Ile Arg Glu Val
500 505 510
Thr Arg Gln Gln Gly Asp Gly Gly Ala Lys Thr Asp Ala Val Leu Cys
515 520 525
Val Asn Asn Leu Ser Arg Phe Pro Gln Pro Ile Glu Leu Asn Leu Gln
530 535 540
Gln Trp Ala Gly Tyr Ile Pro Val Glu Met Thr Gly Tyr Val Glu Phe
545 550 555 560
Pro Ser Ile Gly Gln Leu Pro Tyr Leu Leu Thr Leu Pro Gly His Gly
565 570 575
Phe Tyr Trp Phe Gln Leu Arg Glu Pro Asp Pro Glu Pro Gly Ala Gln
580 585 590
Gln
<210> 7
<211> 1782
<212> DNA
<213> Artificial
<220>
<223>Mutant enzyme 4
<220>
<221> CDS
<222> (1)..(1782)
<400> 7
atg gag gag cac acg cag ggc agc cat gtc gag gcg ggg atc gtc gag 48
Met Glu Glu His Thr Gln Gly Ser His Val Glu Ala Gly Ile Val Glu
1 5 10 15
cat ccg aac gcc gag gac ttc ggt cat gcg cgc acg ctt ccc acc gac 96
His Pro Asn Ala Glu Asp Phe Gly His Ala Arg Thr Leu Pro Thr Asp
20 25 30
acc aac tgg ttc aag cac gcg gtg ttc tat gag gtg ctg gtg cgg gcg 144
Thr Asn Trp Phe Lys His Ala Val Phe Tyr Glu Val Leu Val Arg Ala
35 40 45
ttc tac gac tcg aac gcc gac ggt atc ggt gat ctg cgc ggc ctc acc 192
Phe Tyr Asp Ser Asn Ala Asp Gly Ile Gly Asp Leu Arg Gly Leu Thr
50 55 60
gaa aaa ctc gac tac atc aag tgg ctc ggg gtc gac tgc ctg tgg ctg 240
Glu Lys Leu Asp Tyr Ile Lys Trp Leu Gly Val Asp Cys Leu Trp Leu
65 70 75 80
ccg ccg ttc tac gat tcg ccg ctg cgc gac ggc ggt tac gac atc cgc 288
Pro Pro Phe Tyr Asp Ser Pro Leu Arg Asp Gly Gly Tyr Asp Ile Arg
85 90 95
gac ttc tac aag gtg ctg ccc gag ttc ggc acg gtc gac gac ttc gtc 336
Asp Phe Tyr Lys Val Leu Pro Glu Phe Gly Thr Val Asp Asp Phe Val
100 105 110
acc ctg ctg gac gcg gcc cac cgc agg ggc atc cgc atc atc acc gac 384
Thr Leu Leu Asp Ala Ala His Arg Arg Gly Ile Arg Ile Ile Thr Asp
115 120 125
ctg gtg atg aac cac acc tcc gat cag cac gag tgg ttc cag gaa tcc 432
Leu Val Met Asn His Thr Ser Asp Gln His Glu Trp Phe Gln Glu Ser
130 135 140
cgg cac aat ccc gac ggc ccc tac ggc gac ttc tac gtc tgg agc gac 480
Arg His Asn Pro Asp Gly Pro Tyr Gly Asp Phe Tyr Val Trp Ser Asp
145 150 155 160
acc agc gac agg tat ccc gcc gcg gcc atc atc ttc gtc gcc gcc gag 528
Thr Ser Asp Arg Tyr Pro Ala Ala Ala Ile Ile Phe Val Ala Ala Glu
165 170 175
gag tcg aac tgg acg ttc gac ccc gtc cgc agg cag ttc tac tgg cac 576
Glu Ser Asn Trp Thr Phe Asp Pro Val Arg Arg Gln Phe Tyr Trp His
180 185 190
cgc ttc ttc agc cac cag ccc gac ctc aac tac gac aac ccg gct gtg 624
Arg Phe Phe Ser His Gln Pro Asp Leu Asn Tyr Asp Asn Pro Ala Val
195 200 205
cag gag gcg atg ctc gac gtg ctg agg ttc tgg ctg gac ctc ggt atc 672
Gln Glu Ala Met Leu Asp Val Leu Arg Phe Trp Leu Asp Leu Gly Ile
210 215 220
gac ggt ttc cgt ctg gac gcg gtg ccg tac ctg ttc gag cgc gag ggc 720
Asp Gly Phe Arg Leu Asp Ala Val Pro Tyr Leu Phe Glu Arg Glu Gly
225 230 235 240
acc aac tgc gag aac ctg ccc gag acc cac gcg ttc ctc aag cgg tgc 768
Thr Asn Cys Glu Asn Leu Pro Glu Thr His Ala Phe Leu Lys Arg Cys
245 250 255
cgc aag gcg atc gac gac gag tac ccg ggg cgc gtg ctg ctg gcc gag 816
Arg Lys Ala Ile Asp Asp Glu Tyr Pro Gly Arg Val Leu Leu Ala Glu
260 265 270
gcc aat cag tgg ccc gcc gac gtc gtc gcg tac ttc ggc gac ccg gac 864
Ala Asn Gln Trp Pro Ala Asp Val Val Ala Tyr Phe Gly Asp Pro Asp
275 280 285
acc ggc ggc gac gaa tgc cac atg gcg ttc cac ttc ccg ctg atg cca 912
Thr Gly Gly Asp Glu Cys His Met Ala Phe His Phe Pro Leu Met Pro
290 295 300
cgg atc ttc atg gcc gtg cgg cgc gag tcc cgc ttc ccg atc tcg gag 960
Arg Ile Phe Met Ala Val Arg Arg Glu Ser Arg Phe Pro Ile Ser Glu
305 310 315 320
atc ctg gcg cag acc ccg ccg atc ccc gac acc gcg cag tgg ggc atc 1008
Ile Leu Ala Gln Thr Pro Pro Ile Pro Asp Thr Ala Gln Trp Gly Ile
325 330 335
ttc ctg cgc aac cac gac gag ctc acg ctc gag atg gtc acc gac gaa 1056
Phe Leu Arg Asn His Asp Glu Leu Thr Leu Glu Met Val Thr Asp Glu
340 345 350
gaa cgt gac tac atg tac gcc gag tac gcc aag gac ccg cgg atg aag 1104
Glu Arg Asp Tyr Met Tyr Ala Glu Tyr Ala Lys Asp Pro Arg Met Lys
355 360 365
gcc aac gtc ggc atc cgg cgc agg ctc gca ccg ctg ttg gag aac gac 1152
Ala Asn Val Gly Ile Arg Arg Arg Leu Ala Pro Leu Leu Glu Asn Asp
370 375 380
cgc aac cag atc gag ttg ttc acc gcg ctg ctg ctc tcg ctg ccg ggg 1200
Arg Asn Gln Ile Glu Leu Phe Thr Ala Leu Leu Leu Ser Leu Pro Gly
385 390 395 400
tcg ccg gtg ctg tac tac ggt gac gag ata ggt atg ggg gac atc atc 1248
Ser Pro Val Leu Tyr Tyr Gly Asp Glu Ile Gly Met Gly Asp Ile Ile
405 410 415
tgg ctc ggc gac cgc gac agc gtg cgt acg ccc atg cag tgg acc ccc 1296
Trp Leu Gly Asp Arg Asp Ser Val Arg Thr Pro Met Gln Trp Thr Pro
420 425 430
gac cgc aat gcg ggc ttc tcc aag gcg act ccg ggc cgg ctc tac ctg 1344
Asp Arg Asn Ala Gly Phe Ser Lys Ala Thr Pro Gly Arg Leu Tyr Leu
435 440 445
ccg ccc aac cag gac gcg gtg tac ggc tac cac tct gtg aac gtc gag 1392
Pro Pro Asn Gln Asp Ala Val Tyr Gly Tyr His Ser Val Asn Val Glu
450 455 460
gcg caa ctc gac agc tcc agc tcg ctg ttg aac tgg acg cgc aac atg 1440
Ala Gln Leu Asp Ser Ser Ser Ser Leu Leu Asn Trp Thr Arg Asn Met
465 470 475 480
ctg gcc gtg cgc agc cgc cac gat gcc ttc gca gtc ggc acg ttc cgc 1488
Leu Ala Val Arg Ser Arg His Asp Ala Phe Ala Val Gly Thr Phe Arg
485 490 495
gaa ctg ggc ggc tcc aat ccg tcg gtg ctg gcg tac atc cgg gag gtc 1536
Glu Leu Gly Gly Ser Asn Pro Ser Val Leu Ala Tyr Ile Arg Glu Val
500 505 510
acg cgc caa cag ggc gac ggc ggc gcg aag acc gat gcc gtc ctg tgt 1584
Thr Arg Gln Gln Gly Asp Gly Gly Ala Lys Thr Asp Ala Val Leu Cys
515 520 525
gtc aac aac ctg tcc cgc ttc ccc cag ccc atc gag ttg aat ctc cag 1632
Val Asn Asn Leu Ser Arg Phe Pro Gln Pro Ile Glu Leu Asn Leu Gln
530 535 540
cag tgg gcc ggt tac ata ccc gtc gag atg acc ggc tac gtc gag ttc 1680
Gln Trp Ala Gly Tyr Ile Pro Val Glu Met Thr Gly Tyr Val Glu Phe
545 550 555 560
ccg agt atc ggc cag ttg ccg tac ctg ctc acc ctg ccc ggt cac ggg 1728
Pro Ser Ile Gly Gln Leu Pro Tyr Leu Leu Thr Leu Pro Gly His Gly
565 570 575
ttc tac tgg ttc cag ctt cga gaa ccc gat cca gaa ccg gga gcg cag 1776
Phe Tyr Trp Phe Gln Leu Arg Glu Pro Asp Pro Glu Pro Gly Ala Gln
580 585 590
caa tga 1782
Gln
<210> 8
<211> 593
<212> PRT
<213> Artificial
<220>
<223> Synthetic Construct
<400> 8
Met Glu Glu His Thr Gln Gly Ser His Val Glu Ala Gly Ile Val Glu
1 5 10 15
His Pro Asn Ala Glu Asp Phe Gly His Ala Arg Thr Leu Pro Thr Asp
20 25 30
Thr Asn Trp Phe Lys His Ala Val Phe Tyr Glu Val Leu Val Arg Ala
35 40 45
Phe Tyr Asp Ser Asn Ala Asp Gly Ile Gly Asp Leu Arg Gly Leu Thr
50 55 60
Glu Lys Leu Asp Tyr Ile Lys Trp Leu Gly Val Asp Cys Leu Trp Leu
65 70 75 80
Pro Pro Phe Tyr Asp Ser Pro Leu Arg Asp Gly Gly Tyr Asp Ile Arg
85 90 95
Asp Phe Tyr Lys Val Leu Pro Glu Phe Gly Thr Val Asp Asp Phe Val
100 105 110
Thr Leu Leu Asp Ala Ala His Arg Arg Gly Ile Arg Ile Ile Thr Asp
115 120 125
Leu Val Met Asn His Thr Ser Asp Gln His Glu Trp Phe Gln Glu Ser
130 135 140
Arg His Asn Pro Asp Gly Pro Tyr Gly Asp Phe Tyr Val Trp Ser Asp
145 150 155 160
Thr Ser Asp Arg Tyr Pro Ala Ala Ala Ile Ile Phe Val Ala Ala Glu
165 170 175
Glu Ser Asn Trp Thr Phe Asp Pro Val Arg Arg Gln Phe Tyr Trp His
180 185 190
Arg Phe Phe Ser His Gln Pro Asp Leu Asn Tyr Asp Asn Pro Ala Val
195 200 205
Gln Glu Ala Met Leu Asp Val Leu Arg Phe Trp Leu Asp Leu Gly Ile
210 215 220
Asp Gly Phe Arg Leu Asp Ala Val Pro Tyr Leu Phe Glu Arg Glu Gly
225 230 235 240
Thr Asn Cys Glu Asn Leu Pro Glu Thr His Ala Phe Leu Lys Arg Cys
245 250 255
Arg Lys Ala Ile Asp Asp Glu Tyr Pro Gly Arg Val Leu Leu Ala Glu
260 265 270
Ala Asn Gln Trp Pro Ala Asp Val Val Ala Tyr Phe Gly Asp Pro Asp
275 280 285
Thr Gly Gly Asp Glu Cys His Met Ala Phe His Phe Pro Leu Met Pro
290 295 300
Arg Ile Phe Met Ala Val Arg Arg Glu Ser Arg Phe Pro Ile Ser Glu
305 310 315 320
Ile Leu Ala Gln Thr Pro Pro Ile Pro Asp Thr Ala Gln Trp Gly Ile
325 330 335
Phe Leu Arg Asn His Asp Glu Leu Thr Leu Glu Met Val Thr Asp Glu
340 345 350
Glu Arg Asp Tyr Met Tyr Ala Glu Tyr Ala Lys Asp Pro Arg Met Lys
355 360 365
Ala Asn Val Gly Ile Arg Arg Arg Leu Ala Pro Leu Leu Glu Asn Asp
370 375 380
Arg Asn Gln Ile Glu Leu Phe Thr Ala Leu Leu Leu Ser Leu Pro Gly
385 390 395 400
Ser Pro Val Leu Tyr Tyr Gly Asp Glu Ile Gly Met Gly Asp Ile Ile
405 410 415
Trp Leu Gly Asp Arg Asp Ser Val Arg Thr Pro Met Gln Trp Thr Pro
420 425 430
Asp Arg Asn Ala Gly Phe Ser Lys Ala Thr Pro Gly Arg Leu Tyr Leu
435 440 445
Pro Pro Asn Gln Asp Ala Val Tyr Gly Tyr His Ser Val Asn Val Glu
450 455 460
Ala Gln Leu Asp Ser Ser Ser Ser Leu Leu Asn Trp Thr Arg Asn Met
465 470 475 480
Leu Ala Val Arg Ser Arg His Asp Ala Phe Ala Val Gly Thr Phe Arg
485 490 495
Glu Leu Gly Gly Ser Asn Pro Ser Val Leu Ala Tyr Ile Arg Glu Val
500 505 510
Thr Arg Gln Gln Gly Asp Gly Gly Ala Lys Thr Asp Ala Val Leu Cys
515 520 525
Val Asn Asn Leu Ser Arg Phe Pro Gln Pro Ile Glu Leu Asn Leu Gln
530 535 540
Gln Trp Ala Gly Tyr Ile Pro Val Glu Met Thr Gly Tyr Val Glu Phe
545 550 555 560
Pro Ser Ile Gly Gln Leu Pro Tyr Leu Leu Thr Leu Pro Gly His Gly
565 570 575
Phe Tyr Trp Phe Gln Leu Arg Glu Pro Asp Pro Glu Pro Gly Ala Gln
580 585 590
Gln
<210> 9
<211> 1782
<212> DNA
<213> Artificial
<220>
<223>Mutant enzyme 5
<220>
<221> CDS
<222> (1)..(1782)
<400> 9
atg gag gag cac acg cag ggc agc cat gtc gag gcg ggg atc gtc gag 48
Met Glu Glu His Thr Gln Gly Ser His Val Glu Ala Gly Ile Val Glu
1 5 10 15
cat ccg aac gcc gag gac ttc ggt cat gcg cgc acg ctt ccc acc gac 96
His Pro Asn Ala Glu Asp Phe Gly His Ala Arg Thr Leu Pro Thr Asp
20 25 30
acc aac tgg ttc aag cac gcg gtg ttc tat gag gtg ctg gtg cgg gcg 144
Thr Asn Trp Phe Lys His Ala Val Phe Tyr Glu Val Leu Val Arg Ala
35 40 45
ttc tac gac tcg aac gcc gac ggt atc ggt gat ctg cgc ggc ctc acc 192
Phe Tyr Asp Ser Asn Ala Asp Gly Ile Gly Asp Leu Arg Gly Leu Thr
50 55 60
gaa aaa ctc gac tac atc aag tgg ctc ggg gtc gac tgc ctg tgg ctg 240
Glu Lys Leu Asp Tyr Ile Lys Trp Leu Gly Val Asp Cys Leu Trp Leu
65 70 75 80
ccg ccg ttc tac gat tcg ccg ctg cgc gac ggc ggt tac gac atc cgc 288
Pro Pro Phe Tyr Asp Ser Pro Leu Arg Asp Gly Gly Tyr Asp Ile Arg
85 90 95
gac ttc tac aag gtg ctg ccc gag ttc ggc acg gtc gac gac ttc gtc 336
Asp Phe Tyr Lys Val Leu Pro Glu Phe Gly Thr Val Asp Asp Phe Val
100 105 110
acc ctg ctg gac gcg gcc cac cgc agg ggc atc cgc atc atc acc gac 384
Thr Leu Leu Asp Ala Ala His Arg Arg Gly Ile Arg Ile Ile Thr Asp
115 120 125
ctg gtg atg aac cac acc tcc gat cag cac gag tgg ttc cag gaa tcc 432
Leu Val Met Asn His Thr Ser Asp Gln His Glu Trp Phe Gln Glu Ser
130 135 140
cgg cac aat ccc gac ggc ccc tac ggc gac ttc tac gtc tgg agc gac 480
Arg His Asn Pro Asp Gly Pro Tyr Gly Asp Phe Tyr Val Trp Ser Asp
145 150 155 160
acc agc gac agg tat ccc gcc gcg gcc atc atc ttc gtc gcc gcc gcg 528
Thr Ser Asp Arg Tyr Pro Ala Ala Ala Ile Ile Phe Val Ala Ala Ala
165 170 175
gag tcg aac tgg acg ttc gac ccc gtc cgc agg cag ttc tac tgg cac 576
Glu Ser Asn Trp Thr Phe Asp Pro Val Arg Arg Gln Phe Tyr Trp His
180 185 190
cgc ttc ttc agc cac cag ccc gac ctc aac tac gac aac ccg gct gtg 624
Arg Phe Phe Ser His Gln Pro Asp Leu Asn Tyr Asp Asn Pro Ala Val
195 200 205
cag gag gcg atg ctc gac gtg ctg agg ttc tgg ctg gac ctc ggt atc 672
Gln Glu Ala Met Leu Asp Val Leu Arg Phe Trp Leu Asp Leu Gly Ile
210 215 220
gac ggt ttc cgt ctg gac gcg gtg ccg tac ctg ttc gag cgc gag ggc 720
Asp Gly Phe Arg Leu Asp Ala Val Pro Tyr Leu Phe Glu Arg Glu Gly
225 230 235 240
acc aac tgc gag aac ctg ccc gag acc cac gcg ttc ctc aag cgg tgc 768
Thr Asn Cys Glu Asn Leu Pro Glu Thr His Ala Phe Leu Lys Arg Cys
245 250 255
cgc aag gcg atc gac gac gag tac ccg ggg cgc gtg ctg ctg gcc gag 816
Arg Lys Ala Ile Asp Asp Glu Tyr Pro Gly Arg Val Leu Leu Ala Glu
260 265 270
gcc aat cag tgg ccc gcc gac gtc gtc gcg tac ttc ggc gac ccg gac 864
Ala Asn Gln Trp Pro Ala Asp Val Val Ala Tyr Phe Gly Asp Pro Asp
275 280 285
acc ggc ggc gac gaa tgc cac atg gcg ttc cac ttc ccg ctg atg cca 912
Thr Gly Gly Asp Glu Cys His Met Ala Phe His Phe Pro Leu Met Pro
290 295 300
cgg atc ttc atg gcc gtg cgg cgc gag tcc cgc ttc ccg atc tcg gag 960
Arg Ile Phe Met Ala Val Arg Arg Glu Ser Arg Phe Pro Ile Ser Glu
305 310 315 320
atc ctg gcg cag acc ccg ccg atc ccc gac acc gcg cag tgg ggc atc 1008
Ile Leu Ala Gln Thr Pro Pro Ile Pro Asp Thr Ala Gln Trp Gly Ile
325 330 335
ttc ctg cgc aac cac gac gag ctc acg ctc gag atg gtc acc gac gaa 1056
Phe Leu Arg Asn His Asp Glu Leu Thr Leu Glu Met Val Thr Asp Glu
340 345 350
gaa cgt gac tac atg tac gcc gag tac gcc aag gac ccg cgg atg aag 1104
Glu Arg Asp Tyr Met Tyr Ala Glu Tyr Ala Lys Asp Pro Arg Met Lys
355 360 365
gcc aac gtc ggc atc cgg cgc agg ctc gca ccg ctg ttg gag aac gac 1152
Ala Asn Val Gly Ile Arg Arg Arg Leu Ala Pro Leu Leu Glu Asn Asp
370 375 380
cgc aac cag atc gag ttg ttc acc gcg ctg ctg ctc tcg ctg ccg ggg 1200
Arg Asn Gln Ile Glu Leu Phe Thr Ala Leu Leu Leu Ser Leu Pro Gly
385 390 395 400
tcg ccg gtg ctg tac tac ggt gac gag ata ggt atg ggg gac atc atc 1248
Ser Pro Val Leu Tyr Tyr Gly Asp Glu Ile Gly Met Gly Asp Ile Ile
405 410 415
tgg ctc ggc gac cgc gac agc gtg cgt acg ccc atg cag tgg acc ccc 1296
Trp Leu Gly Asp Arg Asp Ser Val Arg Thr Pro Met Gln Trp Thr Pro
420 425 430
gac cgc aat gcg ggc ttc tcc aag gcg act ccg ggc cgg ctc tac ctg 1344
Asp Arg Asn Ala Gly Phe Ser Lys Ala Thr Pro Gly Arg Leu Tyr Leu
435 440 445
ccg ccc aac cag gac gcg gtg tac ggc tac cac tct gtg aac gtc gag 1392
Pro Pro Asn Gln Asp Ala Val Tyr Gly Tyr His Ser Val Asn Val Glu
450 455 460
gcg caa ctc gac agc tcc agc tcg ctg ttg aac tgg acg cgc aac atg 1440
Ala Gln Leu Asp Ser Ser Ser Ser Leu Leu Asn Trp Thr Arg Asn Met
465 470 475 480
ctg gcc gtg cgc agc cgc cac gat gcc ttc gca gtc ggc acg ttc cgc 1488
Leu Ala Val Arg Ser Arg His Asp Ala Phe Ala Val Gly Thr Phe Arg
485 490 495
gaa ctg ggc ggc tcc aat ccg tcg gtg ctg gcg tac atc cgg gag gtc 1536
Glu Leu Gly Gly Ser Asn Pro Ser Val Leu Ala Tyr Ile Arg Glu Val
500 505 510
acg cgc caa cag ggc gac ggc ggc gcg aag acc gat gcc gtc ctg tgt 1584
Thr Arg Gln Gln Gly Asp Gly Gly Ala Lys Thr Asp Ala Val Leu Cys
515 520 525
gtc aac aac ctg tcc cgc ttc ccc cag ccc atc gag ttg aat ctc cag 1632
Val Asn Asn Leu Ser Arg Phe Pro Gln Pro Ile Glu Leu Asn Leu Gln
530 535 540
cag tgg gcc ggt tac ata ccc gtc gag atg acc ggc tac gtc gag ttc 1680
Gln Trp Ala Gly Tyr Ile Pro Val Glu Met Thr Gly Tyr Val Glu Phe
545 550 555 560
ccg agt atc ggc cag ttg ccg tac ctg ctc acc ctg ccc ggt cac ggg 1728
Pro Ser Ile Gly Gln Leu Pro Tyr Leu Leu Thr Leu Pro Gly His Gly
565 570 575
ttc tac tgg ttc cag ctt cga gaa ccc gat cca gaa ccg gga gcg cag 1776
Phe Tyr Trp Phe Gln Leu Arg Glu Pro Asp Pro Glu Pro Gly Ala Gln
580 585 590
caa tga 1782
Gln
<210> 10
<211> 593
<212> PRT
<213> Artificial
<220>
<223> Synthetic Construct
<400> 10
Met Glu Glu His Thr Gln Gly Ser His Val Glu Ala Gly Ile Val Glu
1 5 10 15
His Pro Asn Ala Glu Asp Phe Gly His Ala Arg Thr Leu Pro Thr Asp
20 25 30
Thr Asn Trp Phe Lys His Ala Val Phe Tyr Glu Val Leu Val Arg Ala
35 40 45
Phe Tyr Asp Ser Asn Ala Asp Gly Ile Gly Asp Leu Arg Gly Leu Thr
50 55 60
Glu Lys Leu Asp Tyr Ile Lys Trp Leu Gly Val Asp Cys Leu Trp Leu
65 70 75 80
Pro Pro Phe Tyr Asp Ser Pro Leu Arg Asp Gly Gly Tyr Asp Ile Arg
85 90 95
Asp Phe Tyr Lys Val Leu Pro Glu Phe Gly Thr Val Asp Asp Phe Val
100 105 110
Thr Leu Leu Asp Ala Ala His Arg Arg Gly Ile Arg Ile Ile Thr Asp
115 120 125
Leu Val Met Asn His Thr Ser Asp Gln His Glu Trp Phe Gln Glu Ser
130 135 140
Arg His Asn Pro Asp Gly Pro Tyr Gly Asp Phe Tyr Val Trp Ser Asp
145 150 155 160
Thr Ser Asp Arg Tyr Pro Ala Ala Ala Ile Ile Phe Val Ala Ala Ala
165 170 175
Glu Ser Asn Trp Thr Phe Asp Pro Val Arg Arg Gln Phe Tyr Trp His
180 185 190
Arg Phe Phe Ser His Gln Pro Asp Leu Asn Tyr Asp Asn Pro Ala Val
195 200 205
Gln Glu Ala Met Leu Asp Val Leu Arg Phe Trp Leu Asp Leu Gly Ile
210 215 220
Asp Gly Phe Arg Leu Asp Ala Val Pro Tyr Leu Phe Glu Arg Glu Gly
225 230 235 240
Thr Asn Cys Glu Asn Leu Pro Glu Thr His Ala Phe Leu Lys Arg Cys
245 250 255
Arg Lys Ala Ile Asp Asp Glu Tyr Pro Gly Arg Val Leu Leu Ala Glu
260 265 270
Ala Asn Gln Trp Pro Ala Asp Val Val Ala Tyr Phe Gly Asp Pro Asp
275 280 285
Thr Gly Gly Asp Glu Cys His Met Ala Phe His Phe Pro Leu Met Pro
290 295 300
Arg Ile Phe Met Ala Val Arg Arg Glu Ser Arg Phe Pro Ile Ser Glu
305 310 315 320
Ile Leu Ala Gln Thr Pro Pro Ile Pro Asp Thr Ala Gln Trp Gly Ile
325 330 335
Phe Leu Arg Asn His Asp Glu Leu Thr Leu Glu Met Val Thr Asp Glu
340 345 350
Glu Arg Asp Tyr Met Tyr Ala Glu Tyr Ala Lys Asp Pro Arg Met Lys
355 360 365
Ala Asn Val Gly Ile Arg Arg Arg Leu Ala Pro Leu Leu Glu Asn Asp
370 375 380
Arg Asn Gln Ile Glu Leu Phe Thr Ala Leu Leu Leu Ser Leu Pro Gly
385 390 395 400
Ser Pro Val Leu Tyr Tyr Gly Asp Glu Ile Gly Met Gly Asp Ile Ile
405 410 415
Trp Leu Gly Asp Arg Asp Ser Val Arg Thr Pro Met Gln Trp Thr Pro
420 425 430
Asp Arg Asn Ala Gly Phe Ser Lys Ala Thr Pro Gly Arg Leu Tyr Leu
435 440 445
Pro Pro Asn Gln Asp Ala Val Tyr Gly Tyr His Ser Val Asn Val Glu
450 455 460
Ala Gln Leu Asp Ser Ser Ser Ser Leu Leu Asn Trp Thr Arg Asn Met
465 470 475 480
Leu Ala Val Arg Ser Arg His Asp Ala Phe Ala Val Gly Thr Phe Arg
485 490 495
Glu Leu Gly Gly Ser Asn Pro Ser Val Leu Ala Tyr Ile Arg Glu Val
500 505 510
Thr Arg Gln Gln Gly Asp Gly Gly Ala Lys Thr Asp Ala Val Leu Cys
515 520 525
Val Asn Asn Leu Ser Arg Phe Pro Gln Pro Ile Glu Leu Asn Leu Gln
530 535 540
Gln Trp Ala Gly Tyr Ile Pro Val Glu Met Thr Gly Tyr Val Glu Phe
545 550 555 560
Pro Ser Ile Gly Gln Leu Pro Tyr Leu Leu Thr Leu Pro Gly His Gly
565 570 575
Phe Tyr Trp Phe Gln Leu Arg Glu Pro Asp Pro Glu Pro Gly Ala Gln
580 585 590
Gln
Claims (6)
1. derive from mycobacterium smegmatismycobacterium smegmatisMutant of trehalose synthetase, its feature exists
In the aminoacid sequence such as SEQ ID NO of this enzyme：2nd, shown in 4,6,8 or 10.
2. mutant of trehalose synthetase gene is it is characterised in that the nucleotide sequence such as SEQ ID NO of this gene：1、3、5、7
Or shown in 9.
3. recombinant vector is it is characterised in that this recombinant vector contains the gene described in claim 2.
4. transgenic cell is it is characterised in that this transgenic cell contains the gene described in claim 2.
5. compositionss are it is characterised in that said composition is using the enzyme described in claim 1 as active component.
6. the enzyme mutant described in claim 1, the gene described in claim 2, the recombinant vector described in claim 3, power
Profit requires application in catalyzing and synthesizing trehalose for the compositionss described in transgenic cell and claim 5 described in 4.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610876031.2A CN106434586B (en) | 2016-10-08 | 2016-10-08 | Trehalose synthetase mutant and gene thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610876031.2A CN106434586B (en) | 2016-10-08 | 2016-10-08 | Trehalose synthetase mutant and gene thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN106434586A true CN106434586A (en) | 2017-02-22 |
| CN106434586B CN106434586B (en) | 2019-12-10 |
Family
ID=58171835
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201610876031.2A Active CN106434586B (en) | 2016-10-08 | 2016-10-08 | Trehalose synthetase mutant and gene thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN106434586B (en) |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107384888A (en) * | 2017-08-04 | 2017-11-24 | 齐鲁工业大学 | A kind of high temperature resistant trehalose synthase and its application by mutation transformation |
| CN108048439A (en) * | 2017-11-20 | 2018-05-18 | 齐鲁工业大学 | A kind of preparation method and application of saltant type trehalose synthase |
-
2016
- 2016-10-08 CN CN201610876031.2A patent/CN106434586B/en active Active
Non-Patent Citations (3)
| Title |
|---|
| YUNG-LIN WANG: "Structures of trehalose synthase from Deinococcus radiodurans reveal that a closed conformation is involved in catalysis of the intramolecular isomerization", 《ACTA CRYSTALLOGRAPHICA SECTION D BIOLOGICAL CRYSTALLOGRAPHY》 * |
| 李镭: "耻垢分枝杆菌海藻糖合成酶基因TreS的克隆及其在大肠杆菌中的表达", 《食品与生物技术学报》 * |
| 王东生: "海藻糖合成酶结构和功能的研究进展", 《化学与生物工程》 * |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107384888A (en) * | 2017-08-04 | 2017-11-24 | 齐鲁工业大学 | A kind of high temperature resistant trehalose synthase and its application by mutation transformation |
| CN107384888B (en) * | 2017-08-04 | 2020-06-30 | 齐鲁工业大学 | Mutation-modified high-temperature-resistant trehalose synthase and application thereof |
| CN108048439A (en) * | 2017-11-20 | 2018-05-18 | 齐鲁工业大学 | A kind of preparation method and application of saltant type trehalose synthase |
| CN108048439B (en) * | 2017-11-20 | 2021-02-26 | 齐鲁工业大学 | Preparation method and application of mutant trehalose synthase |
Also Published As
| Publication number | Publication date |
|---|---|
| CN106434586B (en) | 2019-12-10 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN112080480B (en) | Glycosyltransferase mutants and uses thereof | |
| CN111647579B (en) | Thermolabile exoinulase mutant MutQ23 delta 9 and preparation and application thereof | |
| Li et al. | Cloning and characterization of a sucrose isomerase from Erwinia rhapontici NX-5 for isomaltulose hyperproduction | |
| CN109609530B (en) | Trehalose synthetase and application thereof in trehalose production | |
| CN109423486B (en) | Novel UDP-glycosyltransferase and use thereof | |
| US20140178961A1 (en) | Constructs and strains for fixing carbon dioxide and methods for preparing the same | |
| Mu et al. | Characterization of a thermostable glucose isomerase with an acidic pH optimum from Acidothermus cellulolyticus | |
| CN113151198B (en) | Gamma-glutamine synthetase mutant, coding gene, amino acid sequence and application thereof | |
| CN104774813B (en) | A kind of leucine dehydrogenase and its preparation method and application | |
| CN103571862B (en) | A kind of preparation method of alkaline pullulanase and application | |
| CN102965361B (en) | Pullulanase XWPu2 and gene thereof | |
| CN106434586A (en) | Trehalose synthase mutant and gene thereof | |
| CN111139229B (en) | Novel GDSL family lipid hydrolase EII-2 and encoding gene and application thereof | |
| CN109988778A (en) | A kind of saccharose phosphorylation enzyme gene and its application | |
| CN108753746B (en) | Maltooligosyl trehalose synthase mutant with improved thermal stability | |
| CN106701629B (en) | Pseudomonas fluorescens, pseudomonas fluorescens lipase LIPASEBJ10 and application thereof | |
| CN105969751B (en) | Beta-glucosidase gene and application thereof | |
| Wu et al. | Molecular cloning of maltooligosyltrehalose trehalohydrolase gene from Nostoc flagelliforme and trehalose-related response to stresses | |
| CN107974441A (en) | Endoglucanase, its encoding gene cel5A-h37 and its application | |
| CN109628429B (en) | Extreme-halophilic surfactant-resistant non-calcium ion-dependent alpha-amylase and gene and application thereof | |
| CN101629180A (en) | Thermophilic multifunctional amylase genes, thermophilic multifunctional amylases and application thereof | |
| Xue et al. | Directed evolution of the transglutaminase from Streptomyces mobaraensis and its enhanced expression in Escherichia coli | |
| US20110020896A1 (en) | Mutant dna polymerases and their genes | |
| US20190249158A1 (en) | Maltooligosyl Trehalose Trehalohydrolase (MTHase) Mutant and Application Thereof | |
| CN110982831B (en) | Application of gene AlgL23 with cold adaptability |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |