产气荚膜梭菌甘油脱水酶基因及其1,3－丙二醇的生产方法

摘要

本发明涉及一种全新的甘油脱水酶基因的克隆及其在相关宿主中的表达，该酶的基因系克隆自产气荚膜梭菌(clostridium perfringens)，它能够在正常的生化反应条件下将甘油转化为3－羟基丙醛。在本发明的基础上，可进一步把1，3－丙二醇氧化还原酶等相关酶基因一起重组到大肠杆菌中，获得一个新的代谢工程菌，这个代谢工程菌可以通过发酵将木薯淀粉的水解物、葡萄糖或甘油转化为1，3－丙二醇。

主权项

1、一种甘油脱水酶基因，其特征在于：该甘油脱水酶基因是产气荚膜梭菌(clostridiumperfringens)中获得，是所述的核苷酸序列包含在SEQ IDNO.1所述的核苷酸序列中，其基因的DNA序列和氨基酸序列如下所示：产气荚膜梭菌甘油脱水酶基因的DNA序列： 1 GTGAAATCTA AAAGATTCCA AGTATTATCA GAACGTCCTG TAAACCAAGA 51 TGGACTTATA GGAGAGTGGG CTGATGAAGG CTTAATAGCT TTAGATAGTC 101 CAAATGATCC AAAATCATCA ATAAAAATAG AAAATGGAAT AATTACTGAA 151 TTAGACGGCA GATCAAGAGA TGAGTTTGAT ATGATAGATA AATTTATAGC 201 AGAGTACGCT ATAAATATAG AAGACGCAGA AGCATCTATG AAACTTTCAT 251 CTAAAGAAAT AGCAAGAAGA TTAGTTGATA TAAATGTTAG TAGAGATGAA 301 ATAGTAAAAA TCACTACTTC AATAACACCA ATGAAGGCTG TAGAAGTTAT 351 TCAAGAAATG AACGTTGTTG AAATGATGAT GGCTCTTCAA AAAATGAGAG 401 CAAGAAGAAC ACCTGCTAAC CAATGTCACG TTACTAACGT AAAAGACAAC 451 CCAGTTCAAA TAGCAGCAGA TGCTGCAGAG GCTGCTTTAA GAGGATTCGC 501 AGAGCAAGAA ACTACAGTAG GTATAGTTAG ATATGCACCT TTTAATGCAT 551 TAGCTATCTT AGTAGGTTCA CAAGTAGGTA GAGGAGGAGT TTTAACTCAA 601 TGTGCAGTTG AGGAAGCTAC TGAACTTGAC CTAGGAATGA GAGGACTTAC 651 AAGTTATGCA GAAACAGTTT CAGTTTATGG AACAGAATCA GTATTTACAG 701 ATGGAGATGA TACTCCATGG TCAAAAGCAT TCTTAGCATC AGCTTATGCT 751 TCAAGAGGAC TTAAGATGAG ATTTACATCA GGTTCAGGTT CAGAAGCATT 801 AATGGGATAC TCAGAAGGTA GATCAATGCT TTACTTAGAA TCAAGATGTA 851 TATATATAAC TAAGGGAGCT GGAGTTCAAG GATTACAAAA TGGTGCAGTT 901 AGTTGTATAG GTATGACAGG AGCAGTTCCA TCAGGAATAA GAGCAGTTCT 951 TGGAGAAAAC TTAATAGCTG CAATGCTTGA TATAGAGGTT GCATCAGCAA1001 ATGACCAAAC ATTCTCACAC TCAGACATAA GAAGAACAGC AAGAATGTTA1051 ATGCAAATGC TTCCAGGAAC AGACTTCATA TTCTCAGGAT ATAGTGCAGT1101 TCCAAACTAC GATAACATGT TTGCTGGATC AAACTTTGAT GCAGAAGACT1151 TTGATGATTA CAACATACTT CAAAGAGACT TAAAAGTTGA CGGTGGATTA1201 AGACCAGTTA CAGAAGAAGA AACTATAAAG GTTAGAAATA AAGCTGCTAA1251 ATGCATACAA ATAATCTTTA GAGAATTAGG ATTCCCAGAA GTTACTGATG1301 AAGAAGTAGA AGCTGCAACT TACTGTCACG GAAGTAAGGA AATGCCAAAC1351 AGAAATGTAG TTGAAGATTT AAAGGCTGCA GAAGAAATGT TAGAAAGAAG1401 AATAACAGGA TTAGATATAA TAAAAGCTTT AAGCAAAAAT GGTATGGAAG1451 ATATAGCAAA CAACTTATTA AACATGCTTA AGCAAAGAGT TACTGGAGAT1501 TATCTTCAAA CTTCAGCAAT TTTAGATAAA GATTTCAATG TTATAAGTGC1551 TGTTAATGAT GTAAATGACT ATATGGGACC TGGAACAGGA TATAGACTAG1601 ATGGTCAAAG ATGGGAAGAA ATCAAAAAAG TTCCTACAGT AATGAGACCA1651 GAGGATATAG AGTAGGGGGA GAATGACAAT GGAAGAAAGA ACTTTTATAC1701 CAGAAATAAC AGTTGAAGAA GTTGGAGAAG CTAAGGTTGG TTTAAGATCA1751 GATGAAGTTG TTATAGGATT AGCACCTGCA TTTTTAAAAT ATCAAAATAA1801 AACAATAGTA GATGTTCCTC ATACTGAAAC TTTACTTGAA ATTATAGCAG1851 GTATTGAAGA AGAAGGATTA CATGCAAGAG TAGTTAGAAT TTTAAGAACA1901 TCTGATGTAT CTTTTATAGC TCATGATGCA GCTTGTTTAA GTGGATCAGG1951 AATAGGTATA GGAATACAAT CAAAAGGTAC TACAGTAATA CATCAAAAAG2001 ATTTATTACC ATTAAATAAC TTAGAATTAT TCTCACAAGC TCCGCTTTTA2051 ACTACAGAAA CATATAGATT AATCGGAAAA AATGCTGCTA AATATGCTAA2101 AGGTGAGTCA CCAACACCAG TACCAGTAAA AAATGACCAA ATGGTTAGAC2151 CTAAATTCAT GGCAAAGGCA GCTTTATTAC ACATAAAAGA AACTAAACAC2201 GTTGAACCAG GTAAAAAGCC AGTTCAATTA GAAGTTAAGT TTTAATTTAG2251 AAAGGATTGA AATCATGGAA AATAAGAGAA TGACTGCTGC AGATTATCCT2301 TTAACTTCTA AGAGAAAAGG AGATATAAAA ACTCCTACAG GAAAAGCTTT2351 AGAAGATATA ACTTTAGAAA AAGTTTTAAG TGGAGAAATC AATGCAGATG2401 ATATAAGAAT TTCACCAGAA ACTTTAGAAA TGCAAGCTCA AATAGCTGAA2451 AGCATGAACA GAGATGCTAT AGCTAGAAAC TTTAGAAGAG CTGCGGAGCT2501 TATAAGAGTT CCAGACGATA GAATATTAGA GATGTACAAT GCTTTAAGAC2551 CATATCGTTC AACAAAAGAA GATTTATTCA AAATAGCTGA TGAACTTGAA2601 ACAAAATATG ATGCTAAAGT TAATGCTGAT TTTGTTAGAG AAGCAGCTGA2651 AGTATATGAA ACTAGAAATA AATTAAGAAT AGAAGAG产气荚膜梭菌甘油脱水酶的氨基酸序列：第一个亚基为554个氨基酸VKSKRFQVLS ERPVNQDGLI GEWADEGLIA LDSPNDPKSS IKIENGIITELDGRSRDEFD MIDKFIAEYA INIEDAEASM KLSSKEIARR LVDINVSRDEIVKITTSITP MKAVEVIQEM NVVEMMMALQ KMRARRTPAN QCHVTNVKDNPVQIAADAAE AALRGFAEQE TTVGIVRYAP FNALAILVGS QVGRGGVLTQCAVEEATELD LGMRGLTSYA ETVSVYGTES VFTDGDDTPW SKAFLASAYASRGLKMRFTS GSGSEALMGY SEGRSMLYLE SRCIYITKGA GVQGLQNGAVSCIGMTGAVP SGIRAVLGEN LIAAMLDIEV ASANDQTFSH SDIRRTARMLMQMLPGTDFI FSGYSAVPNY DNMFAGSNFD AEDFDDYNIL QRDLKVDGGLRPVTEEETIK VRNKAAKCIQ IIFRELGFPE VTDEEVEAAT YCHGSKEMPNRNVVEDLKAA EEMLERRITG LDIIKALSKN GMEDIANNLL NMLKQRVTGDYLQTSAILDK DFNVISAVND VNDYMGPGTG YRLDGQRWEE IKKVPTVMRPEDTE第二个亚基为190个氨基酸MTMEERTFIP EITVEEVGEA KVGLRSDEVV IGLAPAFLKY QNKTIVDVPHTETLLEIIAG IEEEGLHARV VRILRTSDVS FIAHDAACLS GSGIGIGIQSKGTTVIHQKD LLPLNNLELF SQAPLLTTET YRLIGKNAAK YAKGESPTPVPVKNDQMVRP KFMAKAALLH IKETKHVEPG KKPVQLEVKF第三个亚基为141个氨基酸MENKRMTAAD YPLTSKRKGD IKTPTGKALE DITLEKVLSG EINADDIRISPETLEMQAQI AESMNRDAIA RNFRRAAELI RVPDDRILEM YNALRPYRSTKEDLFKIADE LETKYDAKVN ADFVREAAEV YETRNKLRIE E。