JP5805768B2 - スクロースとグリセロールとを同時に利用する新規コハク酸生成変異微生物及びこれを利用したコハク酸製造方法 - Google Patents
スクロースとグリセロールとを同時に利用する新規コハク酸生成変異微生物及びこれを利用したコハク酸製造方法 Download PDFInfo
- Publication number
- JP5805768B2 JP5805768B2 JP2013527009A JP2013527009A JP5805768B2 JP 5805768 B2 JP5805768 B2 JP 5805768B2 JP 2013527009 A JP2013527009 A JP 2013527009A JP 2013527009 A JP2013527009 A JP 2013527009A JP 5805768 B2 JP5805768 B2 JP 5805768B2
- Authority
- JP
- Japan
- Prior art keywords
- succinic acid
- glycerol
- sucrose
- producing
- production
- 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.)
- Active
Links
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 title claims description 246
- KDYFGRWQOYBRFD-UHFFFAOYSA-N succinic acid Chemical compound OC(=O)CCC(O)=O KDYFGRWQOYBRFD-UHFFFAOYSA-N 0.000 title claims description 233
- 239000001384 succinic acid Substances 0.000 title claims description 114
- 238000004519 manufacturing process Methods 0.000 title claims description 72
- 229930006000 Sucrose Natural products 0.000 title claims description 66
- 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 title claims description 66
- 239000005720 sucrose Substances 0.000 title claims description 66
- 244000005700 microbiome Species 0.000 title claims description 51
- 238000000034 method Methods 0.000 claims description 51
- 108090000623 proteins and genes Proteins 0.000 claims description 35
- 230000007246 mechanism Effects 0.000 claims description 31
- 239000006227 byproduct Substances 0.000 claims description 28
- 241000894006 Bacteria Species 0.000 claims description 12
- 210000004767 rumen Anatomy 0.000 claims description 12
- 229930091371 Fructose Natural products 0.000 claims description 11
- 239000005715 Fructose Substances 0.000 claims description 11
- RFSUNEUAIZKAJO-ARQDHWQXSA-N Fructose Chemical compound OC[C@H]1O[C@](O)(CO)[C@@H](O)[C@@H]1O RFSUNEUAIZKAJO-ARQDHWQXSA-N 0.000 claims description 11
- 238000012258 culturing Methods 0.000 claims description 11
- 230000003313 weakening effect Effects 0.000 claims description 9
- 108091000080 Phosphotransferase Proteins 0.000 claims description 6
- KDYFGRWQOYBRFD-NUQCWPJISA-N butanedioic acid Chemical compound O[14C](=O)CC[14C](O)=O KDYFGRWQOYBRFD-NUQCWPJISA-N 0.000 claims description 6
- 102000020233 phosphotransferase Human genes 0.000 claims description 6
- 241000722955 Anaerobiospirillum Species 0.000 claims description 3
- 241001293415 Mannheimia Species 0.000 claims description 3
- 150000007524 organic acids Chemical class 0.000 claims description 3
- 241000131104 Actinobacillus sp. Species 0.000 claims description 2
- 239000001963 growth medium Substances 0.000 claims description 2
- 230000035772 mutation Effects 0.000 claims description 2
- 230000015556 catabolic process Effects 0.000 claims 2
- 210000004027 cell Anatomy 0.000 description 30
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 24
- 229910052799 carbon Inorganic materials 0.000 description 24
- 239000002609 medium Substances 0.000 description 24
- 239000002207 metabolite Substances 0.000 description 18
- 230000001965 increasing effect Effects 0.000 description 16
- 230000004060 metabolic process Effects 0.000 description 16
- 108020004414 DNA Proteins 0.000 description 15
- 238000000855 fermentation Methods 0.000 description 14
- 230000004151 fermentation Effects 0.000 description 14
- 101100337058 Rattus norvegicus Glp1r gene Proteins 0.000 description 13
- 239000000243 solution Substances 0.000 description 13
- XPYBSIWDXQFNMH-UHFFFAOYSA-N D-fructose 1,6-bisphosphate Natural products OP(=O)(O)OCC(O)C(O)C(O)C(=O)COP(O)(O)=O XPYBSIWDXQFNMH-UHFFFAOYSA-N 0.000 description 12
- RNBGYGVWRKECFJ-ZXXMMSQZSA-N alpha-D-fructofuranose 1,6-bisphosphate Chemical compound O[C@H]1[C@H](O)[C@](O)(COP(O)(O)=O)O[C@@H]1COP(O)(O)=O RNBGYGVWRKECFJ-ZXXMMSQZSA-N 0.000 description 12
- 230000001925 catabolic effect Effects 0.000 description 12
- RNBGYGVWRKECFJ-UHFFFAOYSA-N fructose-1,6-phosphate Natural products OC1C(O)C(O)(COP(O)(O)=O)OC1COP(O)(O)=O RNBGYGVWRKECFJ-UHFFFAOYSA-N 0.000 description 12
- 238000013518 transcription Methods 0.000 description 12
- 230000035897 transcription Effects 0.000 description 12
- CURLTUGMZLYLDI-UHFFFAOYSA-N Carbon dioxide Chemical compound O=C=O CURLTUGMZLYLDI-UHFFFAOYSA-N 0.000 description 10
- 101150006566 fruA gene Proteins 0.000 description 10
- 230000014509 gene expression Effects 0.000 description 10
- 230000012010 growth Effects 0.000 description 10
- 230000008850 allosteric inhibition Effects 0.000 description 9
- 239000013598 vector Substances 0.000 description 9
- 230000005764 inhibitory process Effects 0.000 description 8
- AWUCVROLDVIAJX-GSVOUGTGSA-N sn-glycerol 3-phosphate Chemical compound OC[C@@H](O)COP(O)(O)=O AWUCVROLDVIAJX-GSVOUGTGSA-N 0.000 description 8
- 239000000126 substance Substances 0.000 description 8
- 230000001629 suppression Effects 0.000 description 8
- 102000057621 Glycerol kinases Human genes 0.000 description 7
- 108700016170 Glycerol kinases Proteins 0.000 description 7
- 102000030621 adenylate cyclase Human genes 0.000 description 7
- 108060000200 adenylate cyclase Proteins 0.000 description 7
- 230000001939 inductive effect Effects 0.000 description 7
- 239000007788 liquid Substances 0.000 description 7
- DTBNBXWJWCWCIK-UHFFFAOYSA-K phosphonatoenolpyruvate Chemical compound [O-]C(=O)C(=C)OP([O-])([O-])=O DTBNBXWJWCWCIK-UHFFFAOYSA-K 0.000 description 7
- IVOMOUWHDPKRLL-KQYNXXCUSA-N Cyclic adenosine monophosphate Chemical compound C([C@H]1O2)OP(O)(=O)O[C@H]1[C@@H](O)[C@@H]2N1C(N=CN=C2N)=C2N=C1 IVOMOUWHDPKRLL-KQYNXXCUSA-N 0.000 description 6
- LCTONWCANYUPML-UHFFFAOYSA-N Pyruvic acid Chemical compound CC(=O)C(O)=O LCTONWCANYUPML-UHFFFAOYSA-N 0.000 description 6
- IVOMOUWHDPKRLL-UHFFFAOYSA-N UNPD107823 Natural products O1C2COP(O)(=O)OC2C(O)C1N1C(N=CN=C2N)=C2N=C1 IVOMOUWHDPKRLL-UHFFFAOYSA-N 0.000 description 6
- 230000027455 binding Effects 0.000 description 6
- 229960005091 chloramphenicol Drugs 0.000 description 6
- WIIZWVCIJKGZOK-RKDXNWHRSA-N chloramphenicol Chemical compound ClC(Cl)C(=O)N[C@H](CO)[C@H](O)C1=CC=C([N+]([O-])=O)C=C1 WIIZWVCIJKGZOK-RKDXNWHRSA-N 0.000 description 6
- 229940095074 cyclic amp Drugs 0.000 description 6
- 230000002018 overexpression Effects 0.000 description 6
- 229930029653 phosphoenolpyruvate Natural products 0.000 description 6
- 239000007787 solid Substances 0.000 description 6
- 102000004190 Enzymes Human genes 0.000 description 5
- 108090000790 Enzymes Proteins 0.000 description 5
- 102100025169 Max-binding protein MNT Human genes 0.000 description 5
- 239000001569 carbon dioxide Substances 0.000 description 5
- 229910002092 carbon dioxide Inorganic materials 0.000 description 5
- 230000037353 metabolic pathway Effects 0.000 description 5
- 238000002360 preparation method Methods 0.000 description 5
- 239000000047 product Substances 0.000 description 5
- 230000002829 reductive effect Effects 0.000 description 5
- 108091006107 transcriptional repressors Proteins 0.000 description 5
- 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 4
- 101710161955 Mannitol-specific phosphotransferase enzyme IIA component Proteins 0.000 description 4
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 4
- 230000003115 biocidal effect Effects 0.000 description 4
- 108091006374 cAMP receptor proteins Proteins 0.000 description 4
- 230000010261 cell growth Effects 0.000 description 4
- 238000011161 development Methods 0.000 description 4
- 239000012634 fragment Substances 0.000 description 4
- 239000008103 glucose Substances 0.000 description 4
- 230000002401 inhibitory effect Effects 0.000 description 4
- 239000002054 inoculum Substances 0.000 description 4
- 239000012528 membrane Substances 0.000 description 4
- 230000000813 microbial effect Effects 0.000 description 4
- 239000000203 mixture Substances 0.000 description 4
- 238000010899 nucleation Methods 0.000 description 4
- 239000002994 raw material Substances 0.000 description 4
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 3
- 241000588724 Escherichia coli Species 0.000 description 3
- 239000003242 anti bacterial agent Substances 0.000 description 3
- 238000012136 culture method Methods 0.000 description 3
- 238000012217 deletion Methods 0.000 description 3
- 230000037430 deletion Effects 0.000 description 3
- 101150056064 glpK gene Proteins 0.000 description 3
- 101150040073 glpK2 gene Proteins 0.000 description 3
- 238000009499 grossing Methods 0.000 description 3
- 239000002243 precursor Substances 0.000 description 3
- 230000008569 process Effects 0.000 description 3
- 238000000746 purification Methods 0.000 description 3
- 229940107700 pyruvic acid Drugs 0.000 description 3
- 238000004064 recycling Methods 0.000 description 3
- 238000012552 review Methods 0.000 description 3
- 238000000926 separation method Methods 0.000 description 3
- YBJHBAHKTGYVGT-ZKWXMUAHSA-N (+)-Biotin Chemical compound N1C(=O)N[C@@H]2[C@H](CCCCC(=O)O)SC[C@@H]21 YBJHBAHKTGYVGT-ZKWXMUAHSA-N 0.000 description 2
- 108020004465 16S ribosomal RNA Proteins 0.000 description 2
- YEJRWHAVMIAJKC-UHFFFAOYSA-N 4-Butyrolactone Chemical compound O=C1CCCO1 YEJRWHAVMIAJKC-UHFFFAOYSA-N 0.000 description 2
- VHUUQVKOLVNVRT-UHFFFAOYSA-N Ammonium hydroxide Chemical compound [NH4+].[OH-] VHUUQVKOLVNVRT-UHFFFAOYSA-N 0.000 description 2
- CIWBSHSKHKDKBQ-JLAZNSOCSA-N Ascorbic acid Chemical compound OC[C@H](O)[C@H]1OC(=O)C(O)=C1O CIWBSHSKHKDKBQ-JLAZNSOCSA-N 0.000 description 2
- 241000053954 Basfia Species 0.000 description 2
- NBSCHQHZLSJFNQ-GASJEMHNSA-N D-Glucose 6-phosphate Chemical compound OC1O[C@H](COP(O)(O)=O)[C@@H](O)[C@H](O)[C@H]1O NBSCHQHZLSJFNQ-GASJEMHNSA-N 0.000 description 2
- SRBFZHDQGSBBOR-IOVATXLUSA-N D-xylopyranose Chemical compound O[C@@H]1COC(O)[C@H](O)[C@H]1O SRBFZHDQGSBBOR-IOVATXLUSA-N 0.000 description 2
- 241001646716 Escherichia coli K-12 Species 0.000 description 2
- VFRROHXSMXFLSN-UHFFFAOYSA-N Glc6P Natural products OP(=O)(O)OCC(O)C(O)C(O)C(O)C=O VFRROHXSMXFLSN-UHFFFAOYSA-N 0.000 description 2
- CKLJMWTZIZZHCS-REOHCLBHSA-N L-aspartic acid Chemical compound OC(=O)[C@@H](N)CC(O)=O CKLJMWTZIZZHCS-REOHCLBHSA-N 0.000 description 2
- GUBGYTABKSRVRQ-QKKXKWKRSA-N Lactose Natural products OC[C@H]1O[C@@H](O[C@H]2[C@H](O)[C@@H](O)C(O)O[C@@H]2CO)[C@H](O)[C@@H](O)[C@H]1O GUBGYTABKSRVRQ-QKKXKWKRSA-N 0.000 description 2
- PVNIIMVLHYAWGP-UHFFFAOYSA-N Niacin Chemical compound OC(=O)C1=CC=CN=C1 PVNIIMVLHYAWGP-UHFFFAOYSA-N 0.000 description 2
- ONIBWKKTOPOVIA-UHFFFAOYSA-N Proline Natural products OC(=O)C1CCCN1 ONIBWKKTOPOVIA-UHFFFAOYSA-N 0.000 description 2
- WYURNTSHIVDZCO-UHFFFAOYSA-N Tetrahydrofuran Chemical compound C1CCOC1 WYURNTSHIVDZCO-UHFFFAOYSA-N 0.000 description 2
- 241000029538 [Mannheimia] succiniciproducens Species 0.000 description 2
- OIRDTQYFTABQOQ-KQYNXXCUSA-N adenosine Chemical compound C1=NC=2C(N)=NC=NC=2N1[C@@H]1O[C@H](CO)[C@@H](O)[C@H]1O OIRDTQYFTABQOQ-KQYNXXCUSA-N 0.000 description 2
- WNLRTRBMVRJNCN-UHFFFAOYSA-N adipic acid Chemical compound OC(=O)CCCCC(O)=O WNLRTRBMVRJNCN-UHFFFAOYSA-N 0.000 description 2
- QGZKDVFQNNGYKY-UHFFFAOYSA-N ammonia Natural products N QGZKDVFQNNGYKY-UHFFFAOYSA-N 0.000 description 2
- 229960000723 ampicillin Drugs 0.000 description 2
- AVKUERGKIZMTKX-NJBDSQKTSA-N ampicillin Chemical compound C1([C@@H](N)C(=O)N[C@H]2[C@H]3SC([C@@H](N3C2=O)C(O)=O)(C)C)=CC=CC=C1 AVKUERGKIZMTKX-NJBDSQKTSA-N 0.000 description 2
- 235000003704 aspartic acid Nutrition 0.000 description 2
- 230000008901 benefit Effects 0.000 description 2
- WQZGKKKJIJFFOK-VFUOTHLCSA-N beta-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 description 2
- OQFSQFPPLPISGP-UHFFFAOYSA-N beta-carboxyaspartic acid Natural products OC(=O)C(N)C(C(O)=O)C(O)=O OQFSQFPPLPISGP-UHFFFAOYSA-N 0.000 description 2
- 230000001588 bifunctional effect Effects 0.000 description 2
- 239000003225 biodiesel Substances 0.000 description 2
- 230000006696 biosynthetic metabolic pathway Effects 0.000 description 2
- WERYXYBDKMZEQL-UHFFFAOYSA-N butane-1,4-diol Chemical compound OCCCCO WERYXYBDKMZEQL-UHFFFAOYSA-N 0.000 description 2
- 150000001720 carbohydrates Chemical class 0.000 description 2
- 238000006243 chemical reaction Methods 0.000 description 2
- 230000003247 decreasing effect Effects 0.000 description 2
- 238000010790 dilution Methods 0.000 description 2
- 239000012895 dilution Substances 0.000 description 2
- 238000004520 electroporation Methods 0.000 description 2
- BTCSSZJGUNDROE-UHFFFAOYSA-N gamma-aminobutyric acid Chemical compound NCCCC(O)=O BTCSSZJGUNDROE-UHFFFAOYSA-N 0.000 description 2
- 238000010353 genetic engineering Methods 0.000 description 2
- 150000002314 glycerols Chemical class 0.000 description 2
- UYTPUPDQBNUYGX-UHFFFAOYSA-N guanine Chemical compound O=C1NC(N)=NC2=C1N=CN2 UYTPUPDQBNUYGX-UHFFFAOYSA-N 0.000 description 2
- 229940093915 gynecological organic acid Drugs 0.000 description 2
- 230000006801 homologous recombination Effects 0.000 description 2
- 238000002744 homologous recombination Methods 0.000 description 2
- JVTAAEKCZFNVCJ-UHFFFAOYSA-N lactic acid Chemical compound CC(O)C(O)=O JVTAAEKCZFNVCJ-UHFFFAOYSA-N 0.000 description 2
- 239000008101 lactose Substances 0.000 description 2
- 230000001404 mediated effect Effects 0.000 description 2
- 238000012269 metabolic engineering Methods 0.000 description 2
- BDAGIHXWWSANSR-UHFFFAOYSA-N methanoic acid Natural products OC=O BDAGIHXWWSANSR-UHFFFAOYSA-N 0.000 description 2
- 229930027945 nicotinamide-adenine dinucleotide Natural products 0.000 description 2
- 235000005985 organic acids Nutrition 0.000 description 2
- 230000037361 pathway Effects 0.000 description 2
- 239000008188 pellet Substances 0.000 description 2
- 108010055837 phosphocarrier protein HPr Proteins 0.000 description 2
- 230000009467 reduction Effects 0.000 description 2
- 239000011780 sodium chloride Substances 0.000 description 2
- -1 sucrose Chemical class 0.000 description 2
- OGNSCSPNOLGXSM-UHFFFAOYSA-N (+/-)-DABA Natural products NCCC(N)C(O)=O OGNSCSPNOLGXSM-UHFFFAOYSA-N 0.000 description 1
- HFVMEOPYDLEHBR-UHFFFAOYSA-N (2-fluorophenyl)-phenylmethanol Chemical compound C=1C=CC=C(F)C=1C(O)C1=CC=CC=C1 HFVMEOPYDLEHBR-UHFFFAOYSA-N 0.000 description 1
- JAHNSTQSQJOJLO-UHFFFAOYSA-N 2-(3-fluorophenyl)-1h-imidazole Chemical compound FC1=CC=CC(C=2NC=CN=2)=C1 JAHNSTQSQJOJLO-UHFFFAOYSA-N 0.000 description 1
- OSWFIVFLDKOXQC-UHFFFAOYSA-N 4-(3-methoxyphenyl)aniline Chemical compound COC1=CC=CC(C=2C=CC(N)=CC=2)=C1 OSWFIVFLDKOXQC-UHFFFAOYSA-N 0.000 description 1
- 108010092060 Acetate kinase Proteins 0.000 description 1
- 241000606750 Actinobacillus Species 0.000 description 1
- GUBGYTABKSRVRQ-XLOQQCSPSA-N Alpha-Lactose 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](O)[C@H]1O GUBGYTABKSRVRQ-XLOQQCSPSA-N 0.000 description 1
- DCXYFEDJOCDNAF-UHFFFAOYSA-N Asparagine Natural products OC(=O)C(N)CC(N)=O DCXYFEDJOCDNAF-UHFFFAOYSA-N 0.000 description 1
- 239000002126 C01EB10 - Adenosine Substances 0.000 description 1
- 108091026890 Coding region Proteins 0.000 description 1
- 230000004568 DNA-binding Effects 0.000 description 1
- YPZRHBJKEMOYQH-UYBVJOGSSA-N FADH2 Chemical compound C1=NC2=C(N)N=CN=C2N1[C@@H]([C@H](O)[C@@H]1O)O[C@@H]1COP(O)(=O)OP(O)(=O)OC[C@@H](O)[C@@H](O)[C@@H](O)CN1C(NC(=O)NC2=O)=C2NC2=C1C=C(C)C(C)=C2 YPZRHBJKEMOYQH-UYBVJOGSSA-N 0.000 description 1
- VZCYOOQTPOCHFL-OWOJBTEDSA-N Fumaric acid Chemical compound OC(=O)\C=C\C(O)=O VZCYOOQTPOCHFL-OWOJBTEDSA-N 0.000 description 1
- QNAYBMKLOCPYGJ-REOHCLBHSA-N L-alanine Chemical compound C[C@H](N)C(O)=O QNAYBMKLOCPYGJ-REOHCLBHSA-N 0.000 description 1
- DCXYFEDJOCDNAF-REOHCLBHSA-N L-asparagine Chemical compound OC(=O)[C@@H](N)CC(N)=O DCXYFEDJOCDNAF-REOHCLBHSA-N 0.000 description 1
- 102000003855 L-lactate dehydrogenase Human genes 0.000 description 1
- 108700023483 L-lactate dehydrogenases Proteins 0.000 description 1
- FFEARJCKVFRZRR-BYPYZUCNSA-N L-methionine Chemical compound CSCC[C@H](N)C(O)=O FFEARJCKVFRZRR-BYPYZUCNSA-N 0.000 description 1
- 101100433987 Latilactobacillus sakei subsp. sakei (strain 23K) ackA1 gene Proteins 0.000 description 1
- OFOBLEOULBTSOW-UHFFFAOYSA-N Malonic acid Chemical compound OC(=O)CC(O)=O OFOBLEOULBTSOW-UHFFFAOYSA-N 0.000 description 1
- ACFIXJIJDZMPPO-NNYOXOHSSA-N NADPH Chemical compound C1=CCC(C(=O)N)=CN1[C@H]1[C@H](O)[C@H](O)[C@@H](COP(O)(=O)OP(O)(=O)OC[C@@H]2[C@H]([C@@H](OP(O)(O)=O)[C@@H](O2)N2C3=NC=NC(N)=C3N=C2)O)O1 ACFIXJIJDZMPPO-NNYOXOHSSA-N 0.000 description 1
- 108700023175 Phosphate acetyltransferases Proteins 0.000 description 1
- LCTONWCANYUPML-UHFFFAOYSA-M Pyruvate Chemical compound CC(=O)C([O-])=O LCTONWCANYUPML-UHFFFAOYSA-M 0.000 description 1
- MTCFGRXMJLQNBG-UHFFFAOYSA-N Serine Natural products OCC(N)C(O)=O MTCFGRXMJLQNBG-UHFFFAOYSA-N 0.000 description 1
- 101100398785 Streptococcus agalactiae serotype V (strain ATCC BAA-611 / 2603 V/R) ldhD gene Proteins 0.000 description 1
- JZRWCGZRTZMZEH-UHFFFAOYSA-N Thiamine Natural products CC1=C(CCO)SC=[N+]1CC1=CN=C(C)N=C1N JZRWCGZRTZMZEH-UHFFFAOYSA-N 0.000 description 1
- 101100386830 Zymomonas mobilis subsp. mobilis (strain ATCC 31821 / ZM4 / CP4) ddh gene Proteins 0.000 description 1
- 238000002835 absorbance Methods 0.000 description 1
- 238000009825 accumulation Methods 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 101150006213 ackA gene Proteins 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 239000012190 activator Substances 0.000 description 1
- 229960005305 adenosine Drugs 0.000 description 1
- 239000001361 adipic acid Substances 0.000 description 1
- 235000011037 adipic acid Nutrition 0.000 description 1
- 238000013019 agitation Methods 0.000 description 1
- 235000004279 alanine Nutrition 0.000 description 1
- 230000003281 allosteric effect Effects 0.000 description 1
- 229940088710 antibiotic agent Drugs 0.000 description 1
- PYMYPHUHKUWMLA-UHFFFAOYSA-N arabinose Natural products OCC(O)C(O)C(O)C=O PYMYPHUHKUWMLA-UHFFFAOYSA-N 0.000 description 1
- 235000010323 ascorbic acid Nutrition 0.000 description 1
- 229960005070 ascorbic acid Drugs 0.000 description 1
- 239000011668 ascorbic acid Substances 0.000 description 1
- 235000009582 asparagine Nutrition 0.000 description 1
- 229960001230 asparagine Drugs 0.000 description 1
- 230000002238 attenuated effect Effects 0.000 description 1
- 230000001580 bacterial effect Effects 0.000 description 1
- SRBFZHDQGSBBOR-UHFFFAOYSA-N beta-D-Pyranose-Lyxose Natural products OC1COC(O)C(O)C1O SRBFZHDQGSBBOR-UHFFFAOYSA-N 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 229960002685 biotin Drugs 0.000 description 1
- 235000020958 biotin Nutrition 0.000 description 1
- 239000011616 biotin Substances 0.000 description 1
- 125000005340 bisphosphate group Chemical group 0.000 description 1
- 230000000740 bleeding effect Effects 0.000 description 1
- 230000000903 blocking effect Effects 0.000 description 1
- 229940041514 candida albicans extract Drugs 0.000 description 1
- 125000003178 carboxy group Chemical group [H]OC(*)=O 0.000 description 1
- 238000004113 cell culture Methods 0.000 description 1
- 239000006285 cell suspension Substances 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 210000000349 chromosome Anatomy 0.000 description 1
- 239000010779 crude oil Substances 0.000 description 1
- 235000018417 cysteine Nutrition 0.000 description 1
- XUJNEKJLAYXESH-UHFFFAOYSA-N cysteine Natural products SCC(N)C(O)=O XUJNEKJLAYXESH-UHFFFAOYSA-N 0.000 description 1
- 230000007423 decrease Effects 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 238000009585 enzyme analysis Methods 0.000 description 1
- 229940071087 ethylenediamine disuccinate Drugs 0.000 description 1
- 239000013604 expression vector Substances 0.000 description 1
- 230000002349 favourable effect Effects 0.000 description 1
- 235000019253 formic acid Nutrition 0.000 description 1
- 229960003692 gamma aminobutyric acid Drugs 0.000 description 1
- 238000012224 gene deletion Methods 0.000 description 1
- 230000002068 genetic effect Effects 0.000 description 1
- 230000007614 genetic variation Effects 0.000 description 1
- 238000012826 global research Methods 0.000 description 1
- 101150071897 glpF gene Proteins 0.000 description 1
- 125000002791 glucosyl group Chemical group C1([C@H](O)[C@@H](O)[C@H](O)[C@H](O1)CO)* 0.000 description 1
- 238000004128 high performance liquid chromatography Methods 0.000 description 1
- HNDVDQJCIGZPNO-UHFFFAOYSA-N histidine Natural products OC(=O)C(N)CC1=CN=CN1 HNDVDQJCIGZPNO-UHFFFAOYSA-N 0.000 description 1
- 239000012510 hollow fiber Substances 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
- 230000000415 inactivating effect Effects 0.000 description 1
- 238000001802 infusion Methods 0.000 description 1
- 238000011081 inoculation Methods 0.000 description 1
- ZKLLSNQJRLJIGT-UYFOZJQFSA-N keto-D-fructose 1-phosphate Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)C(=O)COP(O)(O)=O ZKLLSNQJRLJIGT-UYFOZJQFSA-N 0.000 description 1
- 239000004310 lactic acid Substances 0.000 description 1
- 235000014655 lactic acid Nutrition 0.000 description 1
- 101150026107 ldh1 gene Proteins 0.000 description 1
- 101150041530 ldha gene Proteins 0.000 description 1
- 238000004811 liquid chromatography Methods 0.000 description 1
- 229940049920 malate Drugs 0.000 description 1
- BJEPYKJPYRNKOW-UHFFFAOYSA-N malic acid Chemical compound OC(=O)C(O)CC(O)=O BJEPYKJPYRNKOW-UHFFFAOYSA-N 0.000 description 1
- 230000002503 metabolic effect Effects 0.000 description 1
- 230000037323 metabolic rate Effects 0.000 description 1
- 229930182817 methionine Natural products 0.000 description 1
- LVHBHZANLOWSRM-UHFFFAOYSA-N methylenebutanedioic acid Natural products OC(=O)CC(=C)C(O)=O LVHBHZANLOWSRM-UHFFFAOYSA-N 0.000 description 1
- BOPGDPNILDQYTO-NNYOXOHSSA-N nicotinamide-adenine dinucleotide Chemical compound C1=CCC(C(=O)N)=CN1[C@H]1[C@H](O)[C@H](O)[C@@H](COP(O)(=O)OP(O)(=O)OC[C@@H]2[C@H]([C@@H](O)[C@@H](O2)N2C3=NC=NC(N)=C3N=C2)O)O1 BOPGDPNILDQYTO-NNYOXOHSSA-N 0.000 description 1
- 235000001968 nicotinic acid Nutrition 0.000 description 1
- 229960003512 nicotinic acid Drugs 0.000 description 1
- 239000011664 nicotinic acid Substances 0.000 description 1
- 230000008520 organization Effects 0.000 description 1
- KHPXUQMNIQBQEV-UHFFFAOYSA-N oxaloacetic acid Chemical compound OC(=O)CC(=O)C(O)=O KHPXUQMNIQBQEV-UHFFFAOYSA-N 0.000 description 1
- 229940014662 pantothenate Drugs 0.000 description 1
- 239000011713 pantothenic acid Substances 0.000 description 1
- 230000036961 partial effect Effects 0.000 description 1
- 235000018102 proteins Nutrition 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- LXNHXLLTXMVWPM-UHFFFAOYSA-N pyridoxine Chemical compound CC1=NC=C(CO)C(CO)=C1O LXNHXLLTXMVWPM-UHFFFAOYSA-N 0.000 description 1
- 235000019171 pyridoxine hydrochloride Nutrition 0.000 description 1
- 239000011764 pyridoxine hydrochloride Substances 0.000 description 1
- 229940076788 pyruvate Drugs 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 238000012163 sequencing technique Methods 0.000 description 1
- 230000002226 simultaneous effect Effects 0.000 description 1
- 229960000268 spectinomycin Drugs 0.000 description 1
- UNFWWIHTNXNPBV-WXKVUWSESA-N spectinomycin Chemical compound O([C@@H]1[C@@H](NC)[C@@H](O)[C@H]([C@@H]([C@H]1O1)O)NC)[C@]2(O)[C@H]1O[C@H](C)CC2=O UNFWWIHTNXNPBV-WXKVUWSESA-N 0.000 description 1
- 238000001228 spectrum Methods 0.000 description 1
- 235000000346 sugar Nutrition 0.000 description 1
- 150000008163 sugars Chemical class 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- YLQBMQCUIZJEEH-UHFFFAOYSA-N tetrahydrofuran Natural products C=1C=COC=1 YLQBMQCUIZJEEH-UHFFFAOYSA-N 0.000 description 1
- 235000019157 thiamine Nutrition 0.000 description 1
- KYMBYSLLVAOCFI-UHFFFAOYSA-N thiamine Chemical compound CC1=C(CCO)SCN1CC1=CN=C(C)N=C1N KYMBYSLLVAOCFI-UHFFFAOYSA-N 0.000 description 1
- 229960003495 thiamine Drugs 0.000 description 1
- 239000011721 thiamine Substances 0.000 description 1
- VZCYOOQTPOCHFL-UHFFFAOYSA-N trans-butenedioic acid Natural products OC(=O)C=CC(O)=O VZCYOOQTPOCHFL-UHFFFAOYSA-N 0.000 description 1
- 230000002103 transcriptional effect Effects 0.000 description 1
- 230000037426 transcriptional repression Effects 0.000 description 1
- 238000012546 transfer Methods 0.000 description 1
- 230000007306 turnover Effects 0.000 description 1
- 238000012795 verification Methods 0.000 description 1
- 229940011671 vitamin b6 Drugs 0.000 description 1
- 239000012138 yeast extract Substances 0.000 description 1
Classifications
-
- 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
- C12P7/00—Preparation of oxygen-containing organic compounds
- C12P7/40—Preparation of oxygen-containing organic compounds containing a carboxyl group including Peroxycarboxylic acids
- C12P7/44—Polycarboxylic acids
- C12P7/46—Dicarboxylic acids having four or less carbon atoms, e.g. fumaric acid, maleic acid
-
- 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
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
-
- 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
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
- C12N1/205—Bacterial isolates
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/11—DNA or RNA fragments; Modified forms thereof; Non-coding nucleic acids having a biological activity
- C12N15/52—Genes encoding for enzymes or proenzymes
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02E—REDUCTION OF GREENHOUSE GAS [GHG] EMISSIONS, RELATED TO ENERGY GENERATION, TRANSMISSION OR DISTRIBUTION
- Y02E50/00—Technologies for the production of fuel of non-fossil origin
- Y02E50/10—Biofuels, e.g. bio-diesel
Landscapes
- Engineering & Computer Science (AREA)
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Genetics & Genomics (AREA)
- Organic Chemistry (AREA)
- Wood Science & Technology (AREA)
- Zoology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Biotechnology (AREA)
- General Engineering & Computer Science (AREA)
- Biomedical Technology (AREA)
- Microbiology (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- Molecular Biology (AREA)
- Medicinal Chemistry (AREA)
- Tropical Medicine & Parasitology (AREA)
- Virology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Physics & Mathematics (AREA)
- Biophysics (AREA)
- Plant Pathology (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Description
異化代謝産物抑制機構のメカニズムを詳しく調べると、大きく転写レベルの阻害とアロステリック阻害の二つに分けられる(Deutscher et al., Microbiol. Mol. Biol. Rev., 70:939, 2006; Gorke et al., Nature reviews, 6:613, 2008; Pettigrew et al., J. Bacteriol., 178:2846, 1996; Zwaig et al., J. Bacteriol., 102:753, 1970)。先に転写レベルでは、GlpRレギュレーターの作用でグリセロールを細胞内に輸送し、利用するのに用いられる遺伝子の転写(transcription)が阻害される。アロステリック阻害とは、グリセロール代謝で核心的な機能を担うグリセロールキナーゼにフルクトース1,6−ビスホスフェート(FBP)やEIIA等が結合して、その活性を落とすことをいう。スクロースとグリセロールとが同時にある環境では、GlpRレギュレーターによる転写レベルの阻害と、FBPとEIIAによるアロステリック阻害が、同時に起きて、グリセロール代謝が全般的に阻害される。従って、このような異化代謝産物抑制機構を弱化できる戦略は、GlpRによる転写レベルの阻害を弱化し、FBPやEIIAによるアロステリック阻害を受けないようにすることである。
コハク酸生成微生物のゲノムからfruA遺伝子(プルクトースホスホトランスフェラーゼをコードする遺伝子)を相同組換え方法で除去するために、遺伝子交換ベクターを次のように作製した。先にM.サクシニシプロデュセンスMBEL55E(KCTC0769BP)のゲノムDNAを鋳型として、配列番号1と2、配列番号3と4、配列番号5と6のプライマーを用いてfruA左側相同性部位(HL)、クロラムフェニコール(Cm)を抵抗性遺伝子として含むPCR切片、及びfruA右側相同性部位(HR)各々のPCR切片を取得した。その後、配列番号1と6のプライマーを用いて前記三つのPCR切片を鋳型として、オーバーラッピングPCRを行って、このように得られたDNAの切片をSacIとPstIで切断し、これをpSacHR06ベクターに導入して、pSacHR06fruAベクターを完成した。
配列番号1:5'-ATCGCGGATCCGGTGGAAACCCTCGGTTTATT
配列番号2:5'-AATCTGCTCTGATGCGCAGCTAAAACCTGGTGCAATA
配列番号3:5'-CCAGGTTTTAGCTGCGCATCAGAGCAGATTGTACTGAGAG
配列番号4:5'-AATTACACTTGAAACCCTGATTCTGTGGATAACCGTATTAC
配列番号5:5'-ATCCACAGAATCAGGGTTTCAAGTGTAATTGGCGGAG
配列番号6:5'-TCGACGCGTCGACTTCATCTAACCCCAACGCTTG
実施例1で作製されたfruA遺伝子除去用ベクターpSacHR06fruAを利用して、M.サクシニシプロデュセンスPALK(KCTC10973BP)のゲノムにおいてfruAを欠失させて変異菌株を作製した。
配列番号7:5'-CTATGATTTGGTCGGGCGTTT
配列番号8:5'-TGAATGGCAGAAATTCGAAA
配列番号9:5'-TGATCTTCCGTCACAGGTAT
配列番号10:5''TTGACCGCACTTTGCACATC
大腸菌由来グリセロールキナーゼをコードする遺伝子(glpK22)を含むDNAを取得するために大腸菌K−12 MG1655(ATCC 47076; American Type Culture Collection, Manassas, VA, USA)のゲノムDNAを鋳型として、配列番号11と12、配列番号13と14、配列番号15と16のプライマーを用いて各々PCRを行った。このように得られた三つのPCR産物を鋳型として再度PCRを行うことで、一つのPCR産物を取得し、このPCR産物にはコード配列の913番目の塩基対が本来大腸菌K−12 MG1655のglpK遺伝子が持っていたグアニンがアデノシンに転化されているglpK22遺伝子が含まれているが、本遺伝子から発現するグリセロールキナーゼ(glycerol kinase)は変異前のglpKに比べてFBPとIIAによるアロステリック阻害を相対的に少なく受けることが知られている(Pettigrew et al., J. Bacteriol., 178:2846, 1996)。
配列番号11:5'-ACTCCGGAATTCAACGCACTGACAATCTCACTT
配列番号12:5'-CAGCGAAGCTTTTTGGGTAGAAAGTATAAAGACAGAATCACA
配列番号13:5'-TTTATACTTTCTACCCAAAAAGCTTCGCTGTAATATG
配列番号14:5'-CCATAAACACCGCACTTTCCAACGCATAGTTCACTTC
配列番号15:5'-AACTATGCGTTGGAAAGTGCGGTGTTTATGGCAGG
配列番号16:5'-ACCTGCGAGCTCATTATTGATGTGTGCGGGGT
配列番号17:5'-TAGCTATGTGTTAAAGCCTT
配列番号18:5'-ACCAGGGCACCACCAGCTCC
配列番号19:5'-CCGATTACACCAACGCCTCT
配列番号20:5'-ATGTGGTTCCGGCATTTACC
配列番号21:5'-TTATGCCGCATCCGGTAGTCCC
前記実施例2及び3で作製されたM.サクシニシプロデュセンスPALFK(KCTC11694BP)、またはPALKG(KCTC11693BP)菌株を10g/Lスクロースを含んだ10mLのMH5培地(リットル当り、2.5g酵母エキス(yeast extract)、2.5gポリペプトン(polypeptone)、1gのNaCl、0.02gのCaCl2・2H2O、0.2gのMgCl2・6H2O、8.7gのK2HPO4、及び10.0gのNaHCO3)に播種して嫌気条件下39℃で8時間培養した後、これをまた同じ培地300mLに移して培養した。発酵は、前記培養液を2.5Lの合成培地(リットル当り、1gのNaCl、2gの(NH4)2HPO4、0.02gのCaCl2・2H2O、0.2gのMgCl2・6H2O、8.709gのK2HPO4、0.5gシステイン(cysteine)、0.5gメチオニン(methionine)、0.5gアラニン(alanine)、0.5gアスパラギン(asparagine)、0.5gアスパラギン酸(aspartic acid)、2.46gプロリン(proline)、0.5gセリン(serine)、0.005gニコチン酸(nicotinic acid)、0.005gCa−パントセネート(Ca-pantothenate)、0.005gピリドキシンHCl(pyridoxine・HCl)、0.005gチアミン(thiamine)、0.005gアスコルビン酸(ascorbic acid)及び0.005gビオチン(biotin))を含有した微バイオリアクター(Bioflo 3000, New Brunswick Scientific Co., Edison, NJ, USA)に播種して行い、発酵条件は、22.25g/L(65mM)初期スクロース濃度、4.6g/L(50mM)初期グリセロール濃度で、温度39℃ 200rpm条件で0.2vvm(二酸化炭素体積/培養器内稼動体積(working volume)/分)速度で純粋二酸化炭素を供給しながら発酵させた。発酵中pHはアンモニア水を用いて6.5に調整し、抗生剤は、PALKの場合、スペクチノマイシン50μg/mL、PALFKの場合、クロラムフェニコール6.8μg/mL、PALKGの場合、アンピシリン25μg/mLを各々添加した。高濃度のコハク酸産生のために、炭素源が完全に消耗した場合には、必要時700g/Lのスクロース及びグリセロール溶液を半連続的に添加した。前記と同様の方法でコハク酸産生用変異菌株であるM.サクシニシプロデュセンスPALK(KCTC10973BP)をPALFK菌株の性能と比較のために培養した。培養液内の細胞濃度は、分光光度計を利用して測定し、予め測定した分光光度計の吸光度と乾燥細胞の重量検証線を利用して細胞濃度を計算した。発酵過程中バイオリアクターから周期的にサンプルを採取し、採取された試料は13,000rpmで10分間遠心分離した後、上澄液の各種代謝産物及びコハク酸濃度、スクロース及びグリセロール濃度を液体クロマトグラフィー(HPLC)で分析した。
本実施例では高収率単一コハク酸産生菌株であるM.サクシニシプロデュセンスPALFKの生産性向上方法を確認した。
受託番号:KCTC11693BP
受託日時:20100510
受託機関名:韓国生命工学研究院
受託番号:KCTC11694BP
受託日時:20100510
Claims (7)
- コハク酸生成微生物から、フルクトースホスホトランスフェラーゼをコードする遺伝子を欠失させて、スクロースによるグリセロールの異化代謝産物抑制機構を弱化させることによって得られる、コハク酸産生にスクロースとグリセロールとを同時に利用可能な変異微生物を嫌気的条件で培養し、グリセロールとスクロースとを同時に摂取してコハク酸を生産する段階及び前記培養液からコハク酸を回収する段階を含むコハク酸の製造方法。
- 副産物として、他の有機酸の生成量は、コハク酸生成重量の1%以下であることを特徴とする請求項1に記載のコハク酸の製造方法。
- 前記コハク酸生成微生物は、ルーメンバクテリアであることを特徴とする請求項1又は2に記載の方法。
- 前記ルーメンバクテリアは、マンヘミア属(Mannheimia sp.)、アクチノバチルス属(Actinobacillus sp.)及びアネロビオスピリルム(Anaerobiospirillum sp.)からなる群から選択されることを特徴とする請求項3に記載の方法。
- 前記マンヘミア属菌株は、マンヘミア・サクシニシプロデュセンスPALK(KCTC10973BP)であることを特徴とする請求項4に記載の方法。
- 前記変異微生物は、マンヘミア・サクシニシプロデュセンスPALFK(KCTC11694BP)であることを特徴とする請求項1に記載の方法。
- 下記段階を含むコハク酸の製造方法:
(a)コハク酸生成微生物から、フルクトースホスホトランスフェラーゼをコードする遺伝子を欠失させて、スクロースによるグリセロールの異化代謝産物抑制機構を弱化させることによって得られる、コハク酸産生にスクロースとグリセロールとを同時に利用可能な変異微生物を嫌気的条件で培養し、グリセロールとスクロースとを同時に摂取してコハク酸を生産する段階;
(b)前記培養液内の細胞濃度を高濃度に濃縮させる段階;
(c)前記高濃度の細胞を培養する段階;及び
(d)前記培養液からコハク酸を回収する段階。
Applications Claiming Priority (3)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
KR1020100084327A KR101221557B1 (ko) | 2010-08-30 | 2010-08-30 | 수크로오즈와 글리세롤을 동시에 이용하는 신규 숙신산 생성 변이 미생물 및 이를 이용한 숙신산 제조방법 |
KR10-2010-0084327 | 2010-08-30 | ||
PCT/KR2011/006382 WO2012030130A2 (ko) | 2010-08-30 | 2011-08-30 | 수크로오즈와 글리세롤을 동시에 이용하는 신규 숙신산 생성 변이 미생물 및 이를 이용한 숙신산 제조방법 |
Publications (2)
Publication Number | Publication Date |
---|---|
JP2013539368A JP2013539368A (ja) | 2013-10-24 |
JP5805768B2 true JP5805768B2 (ja) | 2015-11-10 |
Family
ID=45773370
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
JP2013527009A Active JP5805768B2 (ja) | 2010-08-30 | 2011-08-30 | スクロースとグリセロールとを同時に利用する新規コハク酸生成変異微生物及びこれを利用したコハク酸製造方法 |
Country Status (15)
Country | Link |
---|---|
US (1) | US8691516B2 (ja) |
EP (2) | EP2993225B1 (ja) |
JP (1) | JP5805768B2 (ja) |
KR (1) | KR101221557B1 (ja) |
CN (2) | CN105154381A (ja) |
AU (1) | AU2011296791B2 (ja) |
BR (1) | BR112013004652B1 (ja) |
CA (1) | CA2809942C (ja) |
DK (1) | DK2612905T3 (ja) |
ES (2) | ES2602780T3 (ja) |
MX (1) | MX344549B (ja) |
MY (1) | MY183660A (ja) |
NZ (1) | NZ607585A (ja) |
RU (1) | RU2537003C2 (ja) |
WO (1) | WO2012030130A2 (ja) |
Families Citing this family (8)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
KR100780324B1 (ko) | 2006-07-28 | 2007-11-29 | 한국과학기술원 | 신규 순수 숙신산 생성 변이 미생물 및 이를 이용한 숙신산제조방법 |
US9657316B2 (en) | 2012-08-27 | 2017-05-23 | Genomatica, Inc. | Microorganisms and methods for enhancing the availability of reducing equivalents in the presence of methanol, and for producing 1,4-butanediol related thereto |
US10150976B2 (en) | 2012-12-17 | 2018-12-11 | Genomatica, Inc. | Microorganisms and methods for enhancing the availability of reducing equivalents in the presence of methanol, and for producing adipate, 6-aminocaproate, hexamethylenediamine or caprolactam related thereto |
CA2938372A1 (en) * | 2014-02-07 | 2015-08-13 | Basf Se | Modified microorganism for improved production of fine chemicals on sucrose |
WO2015140226A1 (en) * | 2014-03-19 | 2015-09-24 | Basf Se | The use of glycerol with limited feed of carbohydrates for fermentation |
US10287571B2 (en) * | 2014-05-08 | 2019-05-14 | Basf Se | Genetically modified microorganism for improved production of fine chemicals on sucrose |
KR102129379B1 (ko) | 2018-10-10 | 2020-07-02 | 한국과학기술원 | 고활성의 말산 탈수소효소가 도입된 숙신산 생성용 변이 미생물 및 이를 이용한 숙신산 제조방법 |
CN110747147B (zh) * | 2019-11-29 | 2021-05-28 | 江南大学 | 一株衣康酸胁迫抗性提高的大肠杆菌 |
Family Cites Families (14)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US6159738A (en) | 1998-04-28 | 2000-12-12 | University Of Chicago | Method for construction of bacterial strains with increased succinic acid production |
KR100372218B1 (ko) * | 2000-06-29 | 2003-02-14 | 바이오인포메틱스 주식회사 | 유기산을 생산하는 균주 및 이를 이용한 유기산의 생산방법 |
DE602004022584D1 (de) * | 2003-11-27 | 2009-09-24 | Korea Advanced Inst Sci & Tech | Neue kaumagenbakterienvarianten sowie verfahren zur herstellung von bernsteinsäure unter verwendung davon |
JPWO2007013695A1 (ja) * | 2005-07-29 | 2009-02-12 | 株式会社日本触媒 | 細菌にグリセリン資化能を付与する方法 |
DE102005047596A1 (de) * | 2005-10-05 | 2007-04-12 | Degussa Ag | Verfahren zur fermentativen Herstellung von L-Aminosäuren unter Verwendung coryneformer Bakterien |
KR100780324B1 (ko) * | 2006-07-28 | 2007-11-29 | 한국과학기술원 | 신규 순수 숙신산 생성 변이 미생물 및 이를 이용한 숙신산제조방법 |
JP5218400B2 (ja) * | 2007-04-17 | 2013-06-26 | 味の素株式会社 | カルボキシル基を有する酸性物質の製造法 |
BRPI0815409B1 (pt) * | 2007-08-17 | 2023-01-24 | Basf Se | Processos para a produção fermentativa de um composto, e para a produção de um composto, e, uso de uma cepa bacteriana |
WO2009048202A1 (en) * | 2007-10-09 | 2009-04-16 | Korea Advanced Institute Of Science And Technology | Method for preparing succinic acid using glycerol as carbon source |
US8691552B2 (en) * | 2008-10-28 | 2014-04-08 | William Marsh Rice University | Microaerobic cultures for converting glycerol to chemicals |
KR101093199B1 (ko) * | 2009-02-12 | 2011-12-12 | 한국과학기술원 | 글리세롤 대사능력 및 숙신산 생산능력이 향상된 재조합 미생물 및 이를 이용한 숙신산의 제조방법 |
WO2010092155A1 (en) * | 2009-02-16 | 2010-08-19 | Basf Se | Novel microbial succinic acid producers and purification of succinic acid |
ES2686301T3 (es) * | 2010-12-13 | 2018-10-17 | Myriant Corporation | Método de producción de ácido succínico y otros productos químicos utilizando una materia prima que contiene sacarosa |
KR20160116352A (ko) * | 2011-07-22 | 2016-10-07 | 미리안트 코포레이션 | 유기산으로의 글리세롤의 발효 |
-
2010
- 2010-08-30 KR KR1020100084327A patent/KR101221557B1/ko active IP Right Grant
-
2011
- 2011-08-30 US US13/819,339 patent/US8691516B2/en active Active
- 2011-08-30 CN CN201510616196.1A patent/CN105154381A/zh active Pending
- 2011-08-30 ES ES11822105.0T patent/ES2602780T3/es active Active
- 2011-08-30 RU RU2013109055/10A patent/RU2537003C2/ru active
- 2011-08-30 MY MYPI2013000662A patent/MY183660A/en unknown
- 2011-08-30 EP EP15188336.0A patent/EP2993225B1/en active Active
- 2011-08-30 BR BR112013004652-0A patent/BR112013004652B1/pt active IP Right Grant
- 2011-08-30 CN CN201180051937.6A patent/CN103249832B/zh active Active
- 2011-08-30 NZ NZ607585A patent/NZ607585A/en unknown
- 2011-08-30 MX MX2013002394A patent/MX344549B/es active IP Right Grant
- 2011-08-30 EP EP11822105.0A patent/EP2612905B1/en active Active
- 2011-08-30 CA CA2809942A patent/CA2809942C/en active Active
- 2011-08-30 JP JP2013527009A patent/JP5805768B2/ja active Active
- 2011-08-30 ES ES15188336.0T patent/ES2654512T3/es active Active
- 2011-08-30 DK DK11822105.0T patent/DK2612905T3/en active
- 2011-08-30 WO PCT/KR2011/006382 patent/WO2012030130A2/ko active Application Filing
- 2011-08-30 AU AU2011296791A patent/AU2011296791B2/en active Active
Also Published As
Publication number | Publication date |
---|---|
AU2011296791B2 (en) | 2015-05-21 |
MX2013002394A (es) | 2013-12-16 |
MY183660A (en) | 2021-03-05 |
EP2993225A2 (en) | 2016-03-09 |
EP2612905B1 (en) | 2016-10-05 |
AU2011296791A1 (en) | 2013-04-11 |
RU2013109055A (ru) | 2014-09-10 |
US20130217087A1 (en) | 2013-08-22 |
WO2012030130A2 (ko) | 2012-03-08 |
MX344549B (es) | 2016-12-20 |
BR112013004652A2 (pt) | 2016-07-05 |
RU2537003C2 (ru) | 2014-12-27 |
KR20120020614A (ko) | 2012-03-08 |
DK2612905T3 (en) | 2016-12-12 |
CA2809942A1 (en) | 2012-03-08 |
US8691516B2 (en) | 2014-04-08 |
EP2993225B1 (en) | 2017-11-29 |
KR101221557B1 (ko) | 2013-01-14 |
CN103249832B (zh) | 2016-09-21 |
EP2612905A2 (en) | 2013-07-10 |
CN103249832A (zh) | 2013-08-14 |
EP2993225A3 (en) | 2016-04-20 |
JP2013539368A (ja) | 2013-10-24 |
CN105154381A (zh) | 2015-12-16 |
ES2602780T3 (es) | 2017-02-22 |
ES2654512T3 (es) | 2018-02-14 |
BR112013004652B1 (pt) | 2021-05-18 |
NZ607585A (en) | 2014-08-29 |
WO2012030130A3 (ko) | 2012-06-28 |
EP2612905A4 (en) | 2014-12-17 |
CA2809942C (en) | 2016-04-12 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
JP5805768B2 (ja) | スクロースとグリセロールとを同時に利用する新規コハク酸生成変異微生物及びこれを利用したコハク酸製造方法 | |
Zhang et al. | Production of L-alanine by metabolically engineered Escherichia coli | |
KR100780324B1 (ko) | 신규 순수 숙신산 생성 변이 미생물 및 이를 이용한 숙신산제조방법 | |
JP4672671B2 (ja) | 新規ルーメンバクテリア変異菌株及びそれを用いたコハク酸の製造方法 | |
JP6191061B2 (ja) | コハク酸を生産するための組換え大腸菌、及び組み換え大腸菌の使用 | |
WO2006020663A2 (en) | Aerobic succinate production in bacteria | |
US8877466B2 (en) | Bacterial cells having a glyoxylate shunt for the manufacture of succinic acid | |
WO2011163128A1 (en) | Engineered bacteria produce succinate from sucrose | |
CN102154339A (zh) | 一种产丁二酸大肠杆菌基因工程菌株的构建方法 | |
KR20090006713A (ko) | 순수 숙신산 생성 변이균주 및 이를 이용한 숙신산제조방법 | |
KR102129379B1 (ko) | 고활성의 말산 탈수소효소가 도입된 숙신산 생성용 변이 미생물 및 이를 이용한 숙신산 제조방법 | |
KR100630819B1 (ko) | 신규 루멘 박테리아 변이균주 및 이를 이용한 숙신산의제조방법 | |
Tajima et al. | Study of the role of anaerobic metabolism in succinate production by Enterobacter aerogenes | |
KR100556099B1 (ko) | 루멘 박테리아 변이균주 및 이를 이용한 숙신산의 제조방법 | |
KR20050051149A (ko) | 루멘 박테리아 변이균주 및 이를 이용한 숙신산의 제조방법 |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
A977 | Report on retrieval |
Free format text: JAPANESE INTERMEDIATE CODE: A971007 Effective date: 20140730 |
|
A131 | Notification of reasons for refusal |
Free format text: JAPANESE INTERMEDIATE CODE: A131 Effective date: 20140805 |
|
A521 | Request for written amendment filed |
Free format text: JAPANESE INTERMEDIATE CODE: A523 Effective date: 20141105 |
|
A521 | Request for written amendment filed |
Free format text: JAPANESE INTERMEDIATE CODE: A821 Effective date: 20141105 |
|
A02 | Decision of refusal |
Free format text: JAPANESE INTERMEDIATE CODE: A02 Effective date: 20150331 |
|
A521 | Request for written amendment filed |
Free format text: JAPANESE INTERMEDIATE CODE: A523 Effective date: 20150729 |
|
A911 | Transfer to examiner for re-examination before appeal (zenchi) |
Free format text: JAPANESE INTERMEDIATE CODE: A911 Effective date: 20150805 |
|
TRDD | Decision of grant or rejection written | ||
A01 | Written decision to grant a patent or to grant a registration (utility model) |
Free format text: JAPANESE INTERMEDIATE CODE: A01 Effective date: 20150901 |
|
A61 | First payment of annual fees (during grant procedure) |
Free format text: JAPANESE INTERMEDIATE CODE: A61 Effective date: 20150902 |
|
R150 | Certificate of patent or registration of utility model |
Ref document number: 5805768 Country of ref document: JP Free format text: JAPANESE INTERMEDIATE CODE: R150 |
|
R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |