CN117431197A - 具有增强的l-精氨酸或l-瓜氨酸生产力的棒状杆菌属微生物及其相关方法 - Google Patents
具有增强的l-精氨酸或l-瓜氨酸生产力的棒状杆菌属微生物及其相关方法 Download PDFInfo
- Publication number
- CN117431197A CN117431197A CN202211274517.0A CN202211274517A CN117431197A CN 117431197 A CN117431197 A CN 117431197A CN 202211274517 A CN202211274517 A CN 202211274517A CN 117431197 A CN117431197 A CN 117431197A
- Authority
- CN
- China
- Prior art keywords
- corynebacterium
- arginine
- citrulline
- strain
- productivity
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- RHGKLRLOHDJJDR-BYPYZUCNSA-N L-citrulline Chemical compound NC(=O)NCCC[C@H]([NH3+])C([O-])=O RHGKLRLOHDJJDR-BYPYZUCNSA-N 0.000 title claims abstract description 94
- 229960002173 citrulline Drugs 0.000 title claims abstract description 51
- ODKSFYDXXFIFQN-BYPYZUCNSA-N L-arginine Chemical compound OC(=O)[C@@H](N)CCCN=C(N)N ODKSFYDXXFIFQN-BYPYZUCNSA-N 0.000 title claims abstract description 49
- 229930064664 L-arginine Natural products 0.000 title claims abstract description 49
- 235000014852 L-arginine Nutrition 0.000 title claims abstract description 49
- 241000186216 Corynebacterium Species 0.000 title claims abstract description 36
- 238000000034 method Methods 0.000 title abstract description 18
- 244000005700 microbiome Species 0.000 title abstract description 17
- 101710165738 Acetylornithine aminotransferase Proteins 0.000 claims abstract description 23
- 230000000694 effects Effects 0.000 claims abstract description 16
- 108090000623 proteins and genes Proteins 0.000 claims description 37
- 150000001413 amino acids Chemical group 0.000 claims description 29
- 241000186226 Corynebacterium glutamicum Species 0.000 claims description 26
- 235000001014 amino acid Nutrition 0.000 claims description 14
- 238000006467 substitution reaction Methods 0.000 claims description 14
- 230000035772 mutation Effects 0.000 claims description 12
- AYFVYJQAPQTCCC-UHFFFAOYSA-N Threonine Natural products CC(O)C(N)C(O)=O AYFVYJQAPQTCCC-UHFFFAOYSA-N 0.000 claims description 8
- 239000004473 Threonine Substances 0.000 claims description 8
- 235000004279 alanine Nutrition 0.000 claims description 8
- 238000004519 manufacturing process Methods 0.000 claims description 7
- QNAYBMKLOCPYGJ-REOHCLBHSA-N L-alanine Chemical compound C[C@H](N)C(O)=O QNAYBMKLOCPYGJ-REOHCLBHSA-N 0.000 claims description 5
- 238000012258 culturing Methods 0.000 claims description 4
- 125000000341 threoninyl group Chemical class [H]OC([H])(C([H])([H])[H])C([H])(N([H])[H])C(*)=O 0.000 claims description 2
- 230000006696 biosynthetic metabolic pathway Effects 0.000 abstract description 5
- 239000002609 medium Substances 0.000 description 28
- 239000013598 vector Substances 0.000 description 27
- MWUXSHHQAYIFBG-UHFFFAOYSA-N Nitric oxide Chemical compound O=[N] MWUXSHHQAYIFBG-UHFFFAOYSA-N 0.000 description 20
- 210000004027 cell Anatomy 0.000 description 20
- 235000018102 proteins Nutrition 0.000 description 12
- 102000004169 proteins and genes Human genes 0.000 description 12
- 239000004475 Arginine Substances 0.000 description 11
- ODKSFYDXXFIFQN-UHFFFAOYSA-N arginine Natural products OC(=O)C(N)CCCNC(N)=N ODKSFYDXXFIFQN-UHFFFAOYSA-N 0.000 description 11
- 235000009697 arginine Nutrition 0.000 description 11
- RHGKLRLOHDJJDR-UHFFFAOYSA-N Ndelta-carbamoyl-DL-ornithine Natural products OC(=O)C(N)CCCNC(N)=O RHGKLRLOHDJJDR-UHFFFAOYSA-N 0.000 description 8
- 229940024606 amino acid Drugs 0.000 description 8
- 235000013477 citrulline Nutrition 0.000 description 8
- 239000002773 nucleotide Substances 0.000 description 8
- 125000003729 nucleotide group Chemical group 0.000 description 8
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 6
- 102000004190 Enzymes Human genes 0.000 description 6
- 108090000790 Enzymes Proteins 0.000 description 6
- 101150050866 argD gene Proteins 0.000 description 6
- 239000000126 substance Substances 0.000 description 6
- 241000588724 Escherichia coli Species 0.000 description 5
- 229930027917 kanamycin Natural products 0.000 description 5
- SBUJHOSQTJFQJX-NOAMYHISSA-N kanamycin Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CN)O[C@@H]1O[C@H]1[C@H](O)[C@@H](O[C@@H]2[C@@H]([C@@H](N)[C@H](O)[C@@H](CO)O2)O)[C@H](N)C[C@@H]1N SBUJHOSQTJFQJX-NOAMYHISSA-N 0.000 description 5
- 229960000318 kanamycin Drugs 0.000 description 5
- 229930182823 kanamycin A Natural products 0.000 description 5
- 239000000203 mixture Substances 0.000 description 5
- 230000006798 recombination Effects 0.000 description 5
- 238000005215 recombination Methods 0.000 description 5
- 230000001105 regulatory effect Effects 0.000 description 5
- QGZKDVFQNNGYKY-UHFFFAOYSA-N Ammonia Chemical compound N QGZKDVFQNNGYKY-UHFFFAOYSA-N 0.000 description 4
- 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
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 4
- 239000012153 distilled water Substances 0.000 description 4
- 238000011156 evaluation Methods 0.000 description 4
- 238000000855 fermentation Methods 0.000 description 4
- 230000004151 fermentation Effects 0.000 description 4
- 239000012634 fragment Substances 0.000 description 4
- 239000008103 glucose Substances 0.000 description 4
- 239000001963 growth medium Substances 0.000 description 4
- 238000004128 high performance liquid chromatography Methods 0.000 description 4
- 239000003550 marker Substances 0.000 description 4
- 229920001184 polypeptide Polymers 0.000 description 4
- 238000004321 preservation Methods 0.000 description 4
- 102000004196 processed proteins & peptides Human genes 0.000 description 4
- 108090000765 processed proteins & peptides Proteins 0.000 description 4
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 4
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 3
- 108020004414 DNA Proteins 0.000 description 3
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 3
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 3
- ODKSFYDXXFIFQN-BYPYZUCNSA-P L-argininium(2+) Chemical compound NC(=[NH2+])NCCC[C@H]([NH3+])C(O)=O ODKSFYDXXFIFQN-BYPYZUCNSA-P 0.000 description 3
- JRLGPAXAGHMNOL-LURJTMIESA-N N(2)-acetyl-L-ornithine Chemical compound CC(=O)N[C@H](C([O-])=O)CCC[NH3+] JRLGPAXAGHMNOL-LURJTMIESA-N 0.000 description 3
- RFMMMVDNIPUKGG-YFKPBYRVSA-N N-acetyl-L-glutamic acid Chemical compound CC(=O)N[C@H](C(O)=O)CCC(O)=O RFMMMVDNIPUKGG-YFKPBYRVSA-N 0.000 description 3
- KWYUFKZDYYNOTN-UHFFFAOYSA-M Potassium hydroxide Chemical compound [OH-].[K+] KWYUFKZDYYNOTN-UHFFFAOYSA-M 0.000 description 3
- 101100217185 Pseudomonas aeruginosa (strain ATCC 15692 / DSM 22644 / CIP 104116 / JCM 14847 / LMG 12228 / 1C / PRS 101 / PAO1) aruC gene Proteins 0.000 description 3
- 101100022072 Sulfolobus acidocaldarius (strain ATCC 33909 / DSM 639 / JCM 8929 / NBRC 15157 / NCIMB 11770) lysJ gene Proteins 0.000 description 3
- 125000003295 alanine group Chemical group N[C@@H](C)C(=O)* 0.000 description 3
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 3
- 230000015572 biosynthetic process Effects 0.000 description 3
- 229940041514 candida albicans extract Drugs 0.000 description 3
- 238000010367 cloning Methods 0.000 description 3
- 239000003814 drug Substances 0.000 description 3
- 239000003797 essential amino acid Substances 0.000 description 3
- 230000004048 modification Effects 0.000 description 3
- 238000012986 modification Methods 0.000 description 3
- 229910052757 nitrogen Inorganic materials 0.000 description 3
- 239000001301 oxygen Substances 0.000 description 3
- 229910052760 oxygen Inorganic materials 0.000 description 3
- 239000013612 plasmid Substances 0.000 description 3
- 229920001223 polyethylene glycol Polymers 0.000 description 3
- 150000003839 salts Chemical class 0.000 description 3
- 150000003588 threonines Chemical class 0.000 description 3
- 238000013518 transcription Methods 0.000 description 3
- 230000035897 transcription Effects 0.000 description 3
- 239000012138 yeast extract Substances 0.000 description 3
- FWMNVWWHGCHHJJ-SKKKGAJSSA-N 4-amino-1-[(2r)-6-amino-2-[[(2r)-2-[[(2r)-2-[[(2r)-2-amino-3-phenylpropanoyl]amino]-3-phenylpropanoyl]amino]-4-methylpentanoyl]amino]hexanoyl]piperidine-4-carboxylic acid Chemical compound C([C@H](C(=O)N[C@H](CC(C)C)C(=O)N[C@H](CCCCN)C(=O)N1CCC(N)(CC1)C(O)=O)NC(=O)[C@H](N)CC=1C=CC=CC=1)C1=CC=CC=C1 FWMNVWWHGCHHJJ-SKKKGAJSSA-N 0.000 description 2
- 229920001817 Agar Polymers 0.000 description 2
- NLXLAEXVIDQMFP-UHFFFAOYSA-N Ammonia chloride Chemical compound [NH4+].[Cl-] NLXLAEXVIDQMFP-UHFFFAOYSA-N 0.000 description 2
- UXVMQQNJUSDDNG-UHFFFAOYSA-L Calcium chloride Chemical compound [Cl-].[Cl-].[Ca+2] UXVMQQNJUSDDNG-UHFFFAOYSA-L 0.000 description 2
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 2
- 108010078791 Carrier Proteins Proteins 0.000 description 2
- 241000186145 Corynebacterium ammoniagenes Species 0.000 description 2
- 241001014386 Corynebacterium canis Species 0.000 description 2
- 241001134763 Corynebacterium flavescens Species 0.000 description 2
- 241001485655 Corynebacterium glutamicum ATCC 13032 Species 0.000 description 2
- 241000158523 Corynebacterium striatum Species 0.000 description 2
- AHLPHDHHMVZTML-BYPYZUCNSA-N L-Ornithine Chemical compound NCCC[C@H](N)C(O)=O AHLPHDHHMVZTML-BYPYZUCNSA-N 0.000 description 2
- WHUUTDBJXJRKMK-VKHMYHEASA-N L-glutamic acid Chemical compound OC(=O)[C@@H](N)CCC(O)=O WHUUTDBJXJRKMK-VKHMYHEASA-N 0.000 description 2
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 description 2
- 102000004132 Ornithine aminotransferases Human genes 0.000 description 2
- 108090000691 Ornithine aminotransferases Proteins 0.000 description 2
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 description 2
- 239000002202 Polyethylene glycol Substances 0.000 description 2
- 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 2
- 229930006000 Sucrose Natural products 0.000 description 2
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 description 2
- 102000040945 Transcription factor Human genes 0.000 description 2
- 108091023040 Transcription factor Proteins 0.000 description 2
- 108700019146 Transgenes Proteins 0.000 description 2
- XSQUKJJJFZCRTK-UHFFFAOYSA-N Urea Chemical compound NC(N)=O XSQUKJJJFZCRTK-UHFFFAOYSA-N 0.000 description 2
- 239000000654 additive Substances 0.000 description 2
- 230000000996 additive effect Effects 0.000 description 2
- 239000008272 agar Substances 0.000 description 2
- 229910021529 ammonia Inorganic materials 0.000 description 2
- 230000003115 biocidal effect Effects 0.000 description 2
- 239000001110 calcium chloride Substances 0.000 description 2
- 229910001628 calcium chloride Inorganic materials 0.000 description 2
- 229910052799 carbon Inorganic materials 0.000 description 2
- 238000006243 chemical reaction Methods 0.000 description 2
- 239000013599 cloning vector Substances 0.000 description 2
- 238000010276 construction Methods 0.000 description 2
- 239000013601 cosmid vector Substances 0.000 description 2
- 238000012136 culture method Methods 0.000 description 2
- 238000012217 deletion Methods 0.000 description 2
- 230000037430 deletion Effects 0.000 description 2
- 235000014113 dietary fatty acids Nutrition 0.000 description 2
- 229940079593 drug Drugs 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 235000020776 essential amino acid Nutrition 0.000 description 2
- 239000013604 expression vector Substances 0.000 description 2
- 239000000284 extract Substances 0.000 description 2
- 239000000194 fatty acid Substances 0.000 description 2
- 229930195729 fatty acid Natural products 0.000 description 2
- 150000004665 fatty acids Chemical class 0.000 description 2
- 235000013305 food Nutrition 0.000 description 2
- 230000006870 function Effects 0.000 description 2
- IPCSVZSSVZVIGE-UHFFFAOYSA-N hexadecanoic acid Chemical compound CCCCCCCCCCCCCCCC(O)=O IPCSVZSSVZVIGE-UHFFFAOYSA-N 0.000 description 2
- 238000001727 in vivo Methods 0.000 description 2
- 239000012499 inoculation medium Substances 0.000 description 2
- 239000007788 liquid Substances 0.000 description 2
- 230000007246 mechanism Effects 0.000 description 2
- 229910052698 phosphorus Inorganic materials 0.000 description 2
- 239000013600 plasmid vector Substances 0.000 description 2
- 108091033319 polynucleotide Proteins 0.000 description 2
- 239000002157 polynucleotide Substances 0.000 description 2
- 102000040430 polynucleotide Human genes 0.000 description 2
- 238000001556 precipitation Methods 0.000 description 2
- 230000008569 process Effects 0.000 description 2
- 230000010076 replication Effects 0.000 description 2
- 238000012163 sequencing technique Methods 0.000 description 2
- 238000002741 site-directed mutagenesis Methods 0.000 description 2
- 239000011780 sodium chloride Substances 0.000 description 2
- 239000007787 solid Substances 0.000 description 2
- UCSJYZPVAKXKNQ-HZYVHMACSA-N streptomycin Chemical compound CN[C@H]1[C@H](O)[C@@H](O)[C@H](CO)O[C@H]1O[C@@H]1[C@](C=O)(O)[C@H](C)O[C@H]1O[C@@H]1[C@@H](NC(N)=N)[C@H](O)[C@@H](NC(N)=N)[C@H](O)[C@H]1O UCSJYZPVAKXKNQ-HZYVHMACSA-N 0.000 description 2
- 239000005720 sucrose Substances 0.000 description 2
- 230000009466 transformation Effects 0.000 description 2
- 238000013519 translation Methods 0.000 description 2
- 230000024883 vasodilation Effects 0.000 description 2
- 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 1
- VZSRBBMJRBPUNF-UHFFFAOYSA-N 2-(2,3-dihydro-1H-inden-2-ylamino)-N-[3-oxo-3-(2,4,6,7-tetrahydrotriazolo[4,5-c]pyridin-5-yl)propyl]pyrimidine-5-carboxamide Chemical compound C1C(CC2=CC=CC=C12)NC1=NC=C(C=N1)C(=O)NCCC(N1CC2=C(CC1)NN=N2)=O VZSRBBMJRBPUNF-UHFFFAOYSA-N 0.000 description 1
- PAWQVTBBRAZDMG-UHFFFAOYSA-N 2-(3-bromo-2-fluorophenyl)acetic acid Chemical compound OC(=O)CC1=CC=CC(Br)=C1F PAWQVTBBRAZDMG-UHFFFAOYSA-N 0.000 description 1
- 241000251468 Actinopterygii 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
- ATRRKUHOCOJYRX-UHFFFAOYSA-N Ammonium bicarbonate Chemical compound [NH4+].OC([O-])=O ATRRKUHOCOJYRX-UHFFFAOYSA-N 0.000 description 1
- VHUUQVKOLVNVRT-UHFFFAOYSA-N Ammonium hydroxide Chemical compound [NH4+].[OH-] VHUUQVKOLVNVRT-UHFFFAOYSA-N 0.000 description 1
- 239000004254 Ammonium phosphate Substances 0.000 description 1
- KDZOASGQNOPSCU-WDSKDSINSA-N Argininosuccinic acid Chemical compound OC(=O)[C@@H](N)CCC\N=C(/N)N[C@H](C(O)=O)CC(O)=O KDZOASGQNOPSCU-WDSKDSINSA-N 0.000 description 1
- 239000002028 Biomass Substances 0.000 description 1
- 102000014914 Carrier Proteins Human genes 0.000 description 1
- 241000272194 Ciconiiformes Species 0.000 description 1
- 241000186248 Corynebacterium callunae Species 0.000 description 1
- 241000644075 Corynebacterium caspium Species 0.000 description 1
- 241001605246 Corynebacterium crudilactis Species 0.000 description 1
- 241000446654 Corynebacterium deserti Species 0.000 description 1
- 241000272936 Corynebacterium doosanense Species 0.000 description 1
- 241001644925 Corynebacterium efficiens Species 0.000 description 1
- 241000521406 Corynebacterium freiburgense Species 0.000 description 1
- 241000291063 Corynebacterium halotolerans Species 0.000 description 1
- 241000015585 Corynebacterium humireducens Species 0.000 description 1
- 241000024402 Corynebacterium imitans Species 0.000 description 1
- 241000095130 Corynebacterium lubricantis Species 0.000 description 1
- 241000778959 Corynebacterium marinum Species 0.000 description 1
- 241000128247 Corynebacterium pollutisoli Species 0.000 description 1
- 241000186246 Corynebacterium renale Species 0.000 description 1
- 241000334675 Corynebacterium singulare Species 0.000 description 1
- 241000186249 Corynebacterium sp. Species 0.000 description 1
- 241001098119 Corynebacterium spheniscorum Species 0.000 description 1
- 241000186308 Corynebacterium stationis Species 0.000 description 1
- 241000960580 Corynebacterium testudinoris Species 0.000 description 1
- 241000737368 Corynebacterium uterequi Species 0.000 description 1
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N DMSO Substances CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 1
- 238000001712 DNA sequencing Methods 0.000 description 1
- 108010014303 DNA-directed DNA polymerase Proteins 0.000 description 1
- 102000016928 DNA-directed DNA polymerase Human genes 0.000 description 1
- 229920002307 Dextran Polymers 0.000 description 1
- 241000196324 Embryophyta Species 0.000 description 1
- 229930091371 Fructose Natural products 0.000 description 1
- 239000005715 Fructose Substances 0.000 description 1
- RFSUNEUAIZKAJO-ARQDHWQXSA-N Fructose Chemical compound OC[C@H]1O[C@](O)(CO)[C@@H](O)[C@@H]1O RFSUNEUAIZKAJO-ARQDHWQXSA-N 0.000 description 1
- 206010020772 Hypertension Diseases 0.000 description 1
- DGAQECJNVWCQMB-PUAWFVPOSA-M Ilexoside XXIX Chemical compound C[C@@H]1CC[C@@]2(CC[C@@]3(C(=CC[C@H]4[C@]3(CC[C@@H]5[C@@]4(CC[C@@H](C5(C)C)OS(=O)(=O)[O-])C)C)[C@@H]2[C@]1(C)O)C)C(=O)O[C@H]6[C@@H]([C@H]([C@@H]([C@H](O6)CO)O)O)O.[Na+] DGAQECJNVWCQMB-PUAWFVPOSA-M 0.000 description 1
- CKLJMWTZIZZHCS-REOHCLBHSA-N L-aspartic acid Chemical compound OC(=O)[C@@H](N)CC(O)=O CKLJMWTZIZZHCS-REOHCLBHSA-N 0.000 description 1
- 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 1
- OYHQOLUKZRVURQ-HZJYTTRNSA-N Linoleic acid Chemical compound CCCCC\C=C/C\C=C/CCCCCCCC(O)=O OYHQOLUKZRVURQ-HZJYTTRNSA-N 0.000 description 1
- WHXSMMKQMYFTQS-UHFFFAOYSA-N Lithium Chemical compound [Li] WHXSMMKQMYFTQS-UHFFFAOYSA-N 0.000 description 1
- 239000006142 Luria-Bertani Agar Substances 0.000 description 1
- 208000007466 Male Infertility Diseases 0.000 description 1
- 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 1
- 206010028980 Neoplasm Diseases 0.000 description 1
- 108091028043 Nucleic acid sequence Proteins 0.000 description 1
- 108091034117 Oligonucleotide Proteins 0.000 description 1
- AHLPHDHHMVZTML-UHFFFAOYSA-N Orn-delta-NH2 Natural products NCCCC(N)C(O)=O AHLPHDHHMVZTML-UHFFFAOYSA-N 0.000 description 1
- 235000021314 Palmitic acid Nutrition 0.000 description 1
- 239000001888 Peptone Substances 0.000 description 1
- 108010080698 Peptones Proteins 0.000 description 1
- 108010002747 Pfu DNA polymerase Proteins 0.000 description 1
- OAICVXFJPJFONN-UHFFFAOYSA-N Phosphorus Chemical compound [P] OAICVXFJPJFONN-UHFFFAOYSA-N 0.000 description 1
- 244000007853 Sarothamnus scoparius Species 0.000 description 1
- 235000019764 Soybean Meal Nutrition 0.000 description 1
- 229920002472 Starch Polymers 0.000 description 1
- 235000021355 Stearic acid Nutrition 0.000 description 1
- 235000019486 Sunflower oil Nutrition 0.000 description 1
- 108010006785 Taq Polymerase Proteins 0.000 description 1
- 241000270708 Testudinidae Species 0.000 description 1
- 240000008042 Zea mays Species 0.000 description 1
- 235000005824 Zea mays ssp. parviglumis Nutrition 0.000 description 1
- 235000002017 Zea mays subsp mays Nutrition 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 238000007792 addition Methods 0.000 description 1
- -1 al Species 0.000 description 1
- 150000001298 alcohols Chemical class 0.000 description 1
- 229910000147 aluminium phosphate Inorganic materials 0.000 description 1
- 235000005550 amino acid supplement Nutrition 0.000 description 1
- 239000001099 ammonium carbonate Substances 0.000 description 1
- 235000012501 ammonium carbonate Nutrition 0.000 description 1
- 235000019270 ammonium chloride Nutrition 0.000 description 1
- 239000000908 ammonium hydroxide Substances 0.000 description 1
- 229910000148 ammonium phosphate Inorganic materials 0.000 description 1
- 235000019289 ammonium phosphates Nutrition 0.000 description 1
- BFNBIHQBYMNNAN-UHFFFAOYSA-N ammonium sulfate Chemical compound N.N.OS(O)(=O)=O BFNBIHQBYMNNAN-UHFFFAOYSA-N 0.000 description 1
- 229910052921 ammonium sulfate Inorganic materials 0.000 description 1
- 238000012870 ammonium sulfate precipitation Methods 0.000 description 1
- 235000011130 ammonium sulphate Nutrition 0.000 description 1
- 238000000137 annealing Methods 0.000 description 1
- 239000002518 antifoaming agent Substances 0.000 description 1
- 235000003704 aspartic acid Nutrition 0.000 description 1
- 230000037429 base substitution Effects 0.000 description 1
- 235000015278 beef Nutrition 0.000 description 1
- 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 1
- 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 1
- GUBGYTABKSRVRQ-QUYVBRFLSA-N beta-maltose Chemical compound OC[C@H]1O[C@H](O[C@H]2[C@H](O)[C@@H](O)[C@H](O)O[C@@H]2CO)[C@H](O)[C@@H](O)[C@@H]1O GUBGYTABKSRVRQ-QUYVBRFLSA-N 0.000 description 1
- 235000013361 beverage Nutrition 0.000 description 1
- 230000017531 blood circulation Effects 0.000 description 1
- 230000036772 blood pressure Effects 0.000 description 1
- 230000003925 brain function Effects 0.000 description 1
- 239000000872 buffer Substances 0.000 description 1
- 239000001506 calcium phosphate Substances 0.000 description 1
- 229910000389 calcium phosphate Inorganic materials 0.000 description 1
- 235000011010 calcium phosphates Nutrition 0.000 description 1
- 201000011510 cancer Diseases 0.000 description 1
- 239000004202 carbamide Substances 0.000 description 1
- 150000001720 carbohydrates Chemical class 0.000 description 1
- 235000014633 carbohydrates Nutrition 0.000 description 1
- 239000004359 castor oil Substances 0.000 description 1
- 235000019438 castor oil Nutrition 0.000 description 1
- 125000002091 cationic group Chemical group 0.000 description 1
- 239000001913 cellulose Substances 0.000 description 1
- 229920002678 cellulose Polymers 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 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 1
- 229960005091 chloramphenicol Drugs 0.000 description 1
- 238000004587 chromatography analysis Methods 0.000 description 1
- 239000013611 chromosomal DNA Substances 0.000 description 1
- 230000002759 chromosomal effect Effects 0.000 description 1
- 210000000349 chromosome Anatomy 0.000 description 1
- 239000003240 coconut oil Substances 0.000 description 1
- 235000019864 coconut oil Nutrition 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 235000005822 corn Nutrition 0.000 description 1
- 238000002425 crystallisation Methods 0.000 description 1
- 230000008025 crystallization Effects 0.000 description 1
- SUYVUBYJARFZHO-RRKCRQDMSA-N dATP Chemical compound C1=NC=2C(N)=NC=NC=2N1[C@H]1C[C@H](O)[C@@H](COP(O)(=O)OP(O)(=O)OP(O)(O)=O)O1 SUYVUBYJARFZHO-RRKCRQDMSA-N 0.000 description 1
- SUYVUBYJARFZHO-UHFFFAOYSA-N dATP Natural products C1=NC=2C(N)=NC=NC=2N1C1CC(O)C(COP(O)(=O)OP(O)(=O)OP(O)(O)=O)O1 SUYVUBYJARFZHO-UHFFFAOYSA-N 0.000 description 1
- RGWHQCVHVJXOKC-SHYZEUOFSA-J dCTP(4-) Chemical compound O=C1N=C(N)C=CN1[C@@H]1O[C@H](COP([O-])(=O)OP([O-])(=O)OP([O-])([O-])=O)[C@@H](O)C1 RGWHQCVHVJXOKC-SHYZEUOFSA-J 0.000 description 1
- HAAZLUGHYHWQIW-KVQBGUIXSA-N dGTP Chemical compound C1=NC=2C(=O)NC(N)=NC=2N1[C@H]1C[C@H](O)[C@@H](COP(O)(=O)OP(O)(=O)OP(O)(O)=O)O1 HAAZLUGHYHWQIW-KVQBGUIXSA-N 0.000 description 1
- NHVNXKFIZYSCEB-XLPZGREQSA-N dTTP Chemical compound O=C1NC(=O)C(C)=CN1[C@@H]1O[C@H](COP(O)(=O)OP(O)(=O)OP(O)(O)=O)[C@@H](O)C1 NHVNXKFIZYSCEB-XLPZGREQSA-N 0.000 description 1
- 230000003247 decreasing effect Effects 0.000 description 1
- 238000004925 denaturation Methods 0.000 description 1
- 230000036425 denaturation Effects 0.000 description 1
- MNNHAPBLZZVQHP-UHFFFAOYSA-N diammonium hydrogen phosphate Chemical compound [NH4+].[NH4+].OP([O-])([O-])=O MNNHAPBLZZVQHP-UHFFFAOYSA-N 0.000 description 1
- ZPWVASYFFYYZEW-UHFFFAOYSA-L dipotassium hydrogen phosphate Chemical compound [K+].[K+].OP([O-])([O-])=O ZPWVASYFFYYZEW-UHFFFAOYSA-L 0.000 description 1
- 238000004090 dissolution Methods 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 238000001962 electrophoresis Methods 0.000 description 1
- 238000004520 electroporation Methods 0.000 description 1
- 230000002708 enhancing effect Effects 0.000 description 1
- 230000002255 enzymatic effect Effects 0.000 description 1
- 238000001704 evaporation Methods 0.000 description 1
- 230000008020 evaporation Effects 0.000 description 1
- 230000007717 exclusion Effects 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 238000001914 filtration Methods 0.000 description 1
- 238000005187 foaming Methods 0.000 description 1
- 239000007789 gas Substances 0.000 description 1
- 230000002068 genetic effect Effects 0.000 description 1
- 238000010353 genetic engineering Methods 0.000 description 1
- 229960002989 glutamic acid Drugs 0.000 description 1
- ZEMPKEQAKRGZGQ-XOQCFJPHSA-N glycerol triricinoleate Natural products CCCCCC[C@@H](O)CC=CCCCCCCCC(=O)OC[C@@H](COC(=O)CCCCCCCC=CC[C@@H](O)CCCCCC)OC(=O)CCCCCCCC=CC[C@H](O)CCCCCC ZEMPKEQAKRGZGQ-XOQCFJPHSA-N 0.000 description 1
- 230000012010 growth Effects 0.000 description 1
- 239000003630 growth substance Substances 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 238000002744 homologous recombination Methods 0.000 description 1
- 230000006801 homologous recombination Effects 0.000 description 1
- 230000015784 hyperosmotic salinity response Effects 0.000 description 1
- 230000001631 hypertensive effect Effects 0.000 description 1
- 230000036039 immunity Effects 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
- 238000000338 in vitro Methods 0.000 description 1
- 238000011534 incubation Methods 0.000 description 1
- 208000014674 injury Diseases 0.000 description 1
- 150000002484 inorganic compounds Chemical class 0.000 description 1
- 229910010272 inorganic material Inorganic materials 0.000 description 1
- 238000003780 insertion Methods 0.000 description 1
- 230000037431 insertion Effects 0.000 description 1
- 239000000543 intermediate Substances 0.000 description 1
- 238000005342 ion exchange Methods 0.000 description 1
- 238000004255 ion exchange chromatography Methods 0.000 description 1
- 229910000358 iron sulfate Inorganic materials 0.000 description 1
- BAUYGSIQEAFULO-UHFFFAOYSA-L iron(2+) sulfate (anhydrous) Chemical compound [Fe+2].[O-]S([O-])(=O)=O BAUYGSIQEAFULO-UHFFFAOYSA-L 0.000 description 1
- 210000003734 kidney Anatomy 0.000 description 1
- 239000008101 lactose Substances 0.000 description 1
- 235000020778 linoleic acid Nutrition 0.000 description 1
- OYHQOLUKZRVURQ-IXWMQOLASA-N linoleic acid Natural products CCCCC\C=C/C\C=C\CCCCCCCC(O)=O OYHQOLUKZRVURQ-IXWMQOLASA-N 0.000 description 1
- 239000002502 liposome Substances 0.000 description 1
- 229910052744 lithium Inorganic materials 0.000 description 1
- 230000003908 liver function Effects 0.000 description 1
- 229910052943 magnesium sulfate Inorganic materials 0.000 description 1
- 235000019341 magnesium sulphate Nutrition 0.000 description 1
- 235000013372 meat Nutrition 0.000 description 1
- 108020004999 messenger RNA Proteins 0.000 description 1
- 230000004060 metabolic process Effects 0.000 description 1
- 229910052751 metal Inorganic materials 0.000 description 1
- 239000002184 metal Substances 0.000 description 1
- 238000009629 microbiological culture Methods 0.000 description 1
- 238000000520 microinjection Methods 0.000 description 1
- 108091005573 modified proteins Proteins 0.000 description 1
- 102000035118 modified proteins Human genes 0.000 description 1
- 229910000402 monopotassium phosphate Inorganic materials 0.000 description 1
- 235000019796 monopotassium phosphate Nutrition 0.000 description 1
- 210000000214 mouth Anatomy 0.000 description 1
- WQEPLUUGTLDZJY-UHFFFAOYSA-N n-Pentadecanoic acid Natural products CCCCCCCCCCCCCCC(O)=O WQEPLUUGTLDZJY-UHFFFAOYSA-N 0.000 description 1
- 150000007523 nucleic acids Chemical group 0.000 description 1
- QIQXTHQIDYTFRH-UHFFFAOYSA-N octadecanoic acid Chemical compound CCCCCCCCCCCCCCCCCC(O)=O QIQXTHQIDYTFRH-UHFFFAOYSA-N 0.000 description 1
- OQCDKBAXFALNLD-UHFFFAOYSA-N octadecanoic acid Natural products CCCCCCCC(C)CCCCCCCCC(O)=O OQCDKBAXFALNLD-UHFFFAOYSA-N 0.000 description 1
- 235000014593 oils and fats Nutrition 0.000 description 1
- 150000007524 organic acids Chemical class 0.000 description 1
- 235000005985 organic acids Nutrition 0.000 description 1
- 229960003104 ornithine Drugs 0.000 description 1
- 235000019319 peptone Nutrition 0.000 description 1
- PJNZPQUBCPKICU-UHFFFAOYSA-N phosphoric acid;potassium Chemical compound [K].OP(O)(O)=O PJNZPQUBCPKICU-UHFFFAOYSA-N 0.000 description 1
- 239000011574 phosphorus Substances 0.000 description 1
- 229920001522 polyglycol ester Polymers 0.000 description 1
- 238000006116 polymerization reaction Methods 0.000 description 1
- 239000002243 precursor Substances 0.000 description 1
- 238000002360 preparation method Methods 0.000 description 1
- 239000000047 product Substances 0.000 description 1
- 108091008146 restriction endonucleases Proteins 0.000 description 1
- 230000002000 scavenging effect Effects 0.000 description 1
- 238000001338 self-assembly Methods 0.000 description 1
- 230000035939 shock Effects 0.000 description 1
- 239000011734 sodium Substances 0.000 description 1
- 229910052708 sodium Inorganic materials 0.000 description 1
- 239000004455 soybean meal Substances 0.000 description 1
- 239000003549 soybean oil Substances 0.000 description 1
- 235000012424 soybean oil Nutrition 0.000 description 1
- 238000005507 spraying Methods 0.000 description 1
- 239000008107 starch Substances 0.000 description 1
- 235000019698 starch Nutrition 0.000 description 1
- 239000007858 starting material Substances 0.000 description 1
- 239000008117 stearic acid Substances 0.000 description 1
- 229960005322 streptomycin Drugs 0.000 description 1
- 235000000346 sugar Nutrition 0.000 description 1
- 150000008163 sugars Chemical class 0.000 description 1
- 239000002600 sunflower oil Substances 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 230000005062 synaptic transmission Effects 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 230000002194 synthesizing effect Effects 0.000 description 1
- 229940124597 therapeutic agent Drugs 0.000 description 1
- 239000011573 trace mineral Substances 0.000 description 1
- 235000013619 trace mineral Nutrition 0.000 description 1
- 230000008733 trauma Effects 0.000 description 1
- QORWJWZARLRLPR-UHFFFAOYSA-H tricalcium bis(phosphate) Chemical compound [Ca+2].[Ca+2].[Ca+2].[O-]P([O-])([O-])=O.[O-]P([O-])([O-])=O QORWJWZARLRLPR-UHFFFAOYSA-H 0.000 description 1
- 239000001226 triphosphate Substances 0.000 description 1
- 235000011178 triphosphate Nutrition 0.000 description 1
- UNXRWKVEANCORM-UHFFFAOYSA-N triphosphoric acid Chemical compound OP(O)(=O)OP(O)(=O)OP(O)(O)=O UNXRWKVEANCORM-UHFFFAOYSA-N 0.000 description 1
- 239000012137 tryptone Substances 0.000 description 1
- 230000004143 urea cycle Effects 0.000 description 1
- 239000013603 viral vector Substances 0.000 description 1
- 235000013343 vitamin Nutrition 0.000 description 1
- 239000011782 vitamin Substances 0.000 description 1
- 229940088594 vitamin Drugs 0.000 description 1
- 229930003231 vitamin Natural products 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
- 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
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/10—Transferases (2.)
- C12N9/1096—Transferases (2.) transferring nitrogenous groups (2.6)
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
- C12N15/74—Vectors or expression systems specially adapted for prokaryotic hosts other than E. coli, e.g. Lactobacillus, Micromonospora
- C12N15/77—Vectors or expression systems specially adapted for prokaryotic hosts other than E. coli, e.g. Lactobacillus, Micromonospora for Corynebacterium; for Brevibacterium
-
- 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
- C12P13/00—Preparation of nitrogen-containing organic compounds
- C12P13/04—Alpha- or beta- amino acids
- C12P13/10—Citrulline; Arginine; Ornithine
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Y—ENZYMES
- C12Y206/00—Transferases transferring nitrogenous groups (2.6)
- C12Y206/01—Transaminases (2.6.1)
- C12Y206/01011—Acetylornithine transaminase (2.6.1.11)
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Y—ENZYMES
- C12Y206/00—Transferases transferring nitrogenous groups (2.6)
- C12Y206/01—Transaminases (2.6.1)
- C12Y206/01013—Ornithine aminotransferase (2.6.1.13)
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
- C12R2001/15—Corynebacterium
Landscapes
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Organic Chemistry (AREA)
- Engineering & Computer Science (AREA)
- Genetics & Genomics (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Biotechnology (AREA)
- General Engineering & Computer Science (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- Microbiology (AREA)
- Biomedical Technology (AREA)
- Medicinal Chemistry (AREA)
- Molecular Biology (AREA)
- General Chemical & Material Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Tropical Medicine & Parasitology (AREA)
- Virology (AREA)
- Biophysics (AREA)
- Plant Pathology (AREA)
- Physics & Mathematics (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Enzymes And Modification Thereof (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
具有增强的L‑精氨酸或L‑瓜氨酸生产力的棒状杆菌属微生物及其相关方法。本公开内容涉及具有提高的L‑精氨酸或L‑瓜氨酸生产力的棒状杆菌属(Corynebacterium sp.)突变体菌株,以及使用其产生L‑精氨酸或L‑瓜氨酸的方法。所述棒状杆菌属突变体菌株具有参与L‑精氨酸生物合成途径的乙酰鸟氨酸氨基转移酶的增强的活性,并且因此能够具有以与亲本菌株相比提高的产率来产生L‑精氨酸或L‑瓜氨酸的生产力。
Description
技术领域
本公开内容涉及具有提高的L-精氨酸或L-瓜氨酸生产力的棒状杆菌属(Corynebacterium sp.)微生物以及使用其产生L-精氨酸或L-瓜氨酸的方法。
背景技术
已知精氨酸以游离状态包含于植物等中。精氨酸是一种非必需氨基酸,但在成长中的儿童中以及在例如压力、创伤和癌症的特定情况下,它被认为是必须要提供的半必需氨基酸,并被广泛用作用于氨基酸补充剂、药物、食品等的组分。对于药物,精氨酸用于肝功能促进剂、脑功能促进剂、男性不育治疗剂、氨基酸综合制剂等中;以及对于食品,精氨酸用作鱼饼(fish cake)添加剂、健康饮料添加剂和用于高血压患者的盐替代品。
瓜氨酸是一种非必需氨基酸,并且已知瓜氨酸具有有用的作用,例如促进氨代谢、通过血管舒张改善血流、降低血压、神经传递、增强免疫力以及清除活性氧物质。在肾中,瓜氨酸被代谢为精氨酸,由此产生一氧化氮(NO)。即,瓜氨酸在体内不是构成蛋白质的氨基酸,而是尿素循环的中间体之一,并且由精氨酸与被称为具有血管舒张作用的物质的NO一起产生。另外,瓜氨酸与天冬氨酸缩合并再生为精氨酸。
这样的精氨酸和瓜氨酸的产生可使用天然存在的野生型菌株或从野生型菌株修饰以具有提高的精氨酸或瓜氨酸生产力的突变体菌株来进行。最近,为了提高精氨酸或瓜氨酸的产生效率,基因重组技术已经被用于例如大肠杆菌(Escherichia coli)和棒状杆菌(Corynebacterium)的微生物。在微生物中的L-精氨酸的生物合成中,L-谷氨酸被用作起始物质,并被依次转化成N-乙酰谷氨酸、N-乙酰谷氨酰基-P、N-乙酰谷氨酸5-半醛、N-乙酰鸟氨酸、L-鸟氨酸、L-瓜氨酸和精氨琥珀酸,从而合成L-精氨酸。在这种逐步合成过程中涉及多种蛋白质,例如酶、转录因子和转运蛋白。因此,可通过基因重组技术通过调节参与L-精氨酸或L-瓜氨酸生物合成的多种蛋白质的活性来提高L-精氨酸或L-瓜氨酸生产力,并且为了开发具有优异的L-精氨酸或L-瓜氨酸生产力的重组菌株或突变体菌株,需要进行许多研究。
[现有技术文献]
[专利文献]
韩国专利No.10-2269637
发明内容
本公开内容的一个目的是提供具有提高的L-精氨酸或L-瓜氨酸生产力的棒状杆菌属突变体菌株。
本公开内容的另一个目的是提供使用所述突变体菌株产生L-精氨酸或L-瓜氨酸的方法。
本公开内容的一个方面提供了通过具有增强的乙酰鸟氨酸氨基转移酶活性而具有提高的L-精氨酸或L-瓜氨酸生产力的棒状杆菌属突变体菌株。
本公开内容中使用的“乙酰鸟氨酸氨基转移酶”用于催化L-精氨酸生物合成途径中使N-乙酰谷氨酸5-半醛转化成N-乙酰鸟氨酸的反应。N-乙酰鸟氨酸通过多种酶依次转化成其他物质,并且作为结果,合成为L-精氨酸或L-瓜氨酸。
根据本公开内容的一个实施方案,乙酰鸟氨酸氨基转移酶可来源于棒状杆菌属微生物。
更具体地,棒状杆菌属微生物可以是谷氨酸棒状杆菌(Corynebacteriumglutamicum)、Corynebacterium crudilactis、沙漠棒状杆菌(Corynebacteriumdeserti)、帚石南棒状杆菌(Corynebacterium callunae)、Corynebacteriumsuranareeae、Corynebacterium lubricantis、Corynebacterium doosanense、有效棒状杆菌(Corynebacterium efficiens)、Corynebacterium uterequi、停滞棒状杆菌(Corynebacterium stationis)、Corynebacterium pacaense、单一棒状杆菌(Corynebacterium singulare)、Corynebacterium humireducens、海洋棒状杆菌(Corynebacterium marinum)、耐盐棒状杆菌(Corynebacterium halotolerans)、企鹅棒状杆菌(Corynebacterium spheniscorum)、Corynebacterium freiburgense、纹状棒状杆菌(Corynebacterium striatum)、犬棒状杆菌(Corynebacterium canis)、产氨棒状杆菌(Corynebacterium ammoniagenes)、牛肾盂炎棒状杆菌(Corynebacterium renale)、Corynebacterium pollutisoli、汉氏棒状杆菌(Corynebacterium imitans)、黑海棒状杆菌(Corynebacterium caspium)、龟口腔棒状杆菌(Corynebacterium testudinoris)、Corynebacaterium pseudopelargi或微黄棒状杆菌(Corynebacterium flavescens),但不限于此。
本文使用的术语“增强的活性”意指编码目的蛋白质例如酶、转录因子和转运蛋白的基因的表达已经被新引入或已经提高,使得该蛋白质的表达水平与在野生型菌株或修饰之前的菌株中的那些相比有所提高。术语“增强的活性”还包括:通过编码基因的核苷酸序列中的一个或更多个核苷酸的替换、插入、缺失或其组合,与由微生物最初具有的蛋白质活性相比,蛋白质本身的活性已提高的情况;由于编码酶的基因的表达或翻译提高,细胞中的酶的整体活性高于野生型菌株或修饰之前的菌株的那些的情况;及其组合。
根据本公开内容的一个实施方案,乙酰鸟氨酸氨基转移酶活性的增强可通过在编码乙酰鸟氨酸氨基转移酶的基因中进行定点突变来实现。
根据本公开内容的一个实施方案,编码乙酰鸟氨酸氨基转移酶的基因可由SEQ IDNO:1的核苷酸序列表示,或者可由SEQ ID NO:2的氨基酸序列组成。
根据本公开内容的一个实施方案,定点突变可以是SEQ ID NO:1的核苷酸序列中的一个或更多个核苷酸的替换。
根据本公开内容的一个实施方案,定点突变可以是SEQ ID NO:2的氨基酸序列中的一个或更多个氨基酸的替换。
更具体地,定点突变可以是在SEQ ID NO:2的氨基酸序列中位置50至200或100至150处的一个、两个、三个、四个或五个氨基酸的连续或非连续替换。
根据本公开内容的一个实施方案,定点突变可以是在SEQ ID NO:2的氨基酸序列中氨基酸位置131处的苏氨酸对丙氨酸的替换。
本文中使用的术语“提高的生产力”意指L-精氨酸或L-瓜氨酸生产力与亲本菌株中的L-精氨酸或L-瓜氨酸生产力相比有所提高。本文中使用的术语“亲本菌株”是指野生型菌株或待突变的突变体菌株,并且包括待直接突变或待用重组载体等转化的菌株。在本公开内容中,亲本菌株可以是野生型棒状杆菌属微生物或从野生型微生物突变而来的微生物或菌株。
如上所述,根据本公开内容的具有提高的L-精氨酸或L-瓜氨酸生产力的棒状杆菌属突变体菌株包含编码乙酰鸟氨酸氨基转移酶的基因的突变的核苷酸序列或氨基酸序列,并且因此与亲本菌株相比表现出提高的L-精氨酸或L-瓜氨酸生产力。特别地,棒状杆菌属突变体菌株的L-精氨酸或L-瓜氨酸生产力比亲本菌株的L-精氨酸或L-瓜氨酸生产力高至少5%,特别地5%至80%、10%至70%、20%至60%或30%至50%。
根据本公开内容的一个实施方案,棒状杆菌属突变体菌株可以是谷氨酸棒状杆菌。
根据本公开内容的一个实施方案的棒状杆菌属突变体菌株可通过重组载体获得,所述重组载体包含具有亲本菌株中的乙酰鸟氨酸氨基转移酶序列的一部分被替换的变体。
本文中使用的术语“部分”不是意指核苷酸序列、多核苷酸序列或氨基酸序列的全部,而是可以是1至300,优选1至100,更优选1至50,但不限于此。
本文中使用的术语“变体”是指由于在氨基酸序列的N端、C端和/或在氨基酸序列中的一个或更多个氨基酸的保守替换、缺失、修饰或添加而不同于突变之前编码蛋白质的特定基因的氨基酸序列但保留该蛋白质的功能或特性的多肽。本文中使用的术语“保守替换”意指将一个氨基酸用具有类似结构和/或化学性质的另一氨基酸替换。保守替换可对所得蛋白质或多肽的活性具有极小影响或没有影响。另外,变体包括其中一个或更多个部分例如N端前导序列或跨膜结构域已被去除的那些。此外,变体包括其中一部分已从成熟蛋白质的N和/或C端去除的那些。变体的能力与突变之前的多肽的能力相比可提高、不变或降低。在本公开内容中,术语“变体”可与术语例如突变体、修饰、变体多肽、经修饰的蛋白质、突变等互换使用。
根据本公开内容的一个实施方案,乙酰鸟氨酸氨基转移酶变体可具有SEQ ID NO:4的氨基酸序列。
本文中使用的术语“载体”意指能够在合适的宿主细胞中表达目的蛋白质的表达载体,并且是指包含与转基因可操作地连接以使得转基因得以表达的必需调节元件的基因构建体。本文中使用的术语“可操作地连接”意指待表达的基因以允许该基因表达的方式与其调节序列功能性地连接。“调节元件”包括用于影响转录的启动子、用于调节转录的任选操纵子序列、编码合适的mRNA核糖体结合位点的序列以及控制转录和翻译的终止的序列。载体的实例包括但不限于质粒载体、黏粒载体、噬菌体载体、病毒载体等。可使用的噬菌体载体或黏粒载体的实例包括pWE15、M13、λEMBL3、λEMBL4、λFIXII、λDASHII、λZAPII、λgt10、λgt11、Charon4A和Charon21A,并且可使用的质粒载体的实例包括pDZ载体,以及基于pBR、基于pUC、基于pBluescriptll、基于pGEM、基于pTZ、基于pCL和基于pET的载体。可使用的载体没有特别的限制,并且可使用任何已知的表达载体而没有限制。
在本公开内容中使用的“重组载体”被转化到合适的宿主细胞中之后,其可任选地独立于宿主基因组进行复制和发挥功能,或者在一些情况下可整合至基因组本身中。在这种情况下,“合适的宿主细胞”是其中载体可复制的宿主细胞,并且可包含复制起点,其是复制开始的特定核苷酸序列。
可使用根据宿主细胞选择的合适的载体引入技术进行转化,并且可在宿主细胞中表达目的基因。例如,载体引入可通过电穿孔、热激、磷酸钙(CaPO4)沉淀、氯化钙(CaCl2)沉淀、显微注射、聚乙二醇(polyethylene glycol,PEG)法、DEAE-葡聚糖法、阳离子脂质体法、乙酸锂-DMSO法或其组合来进行。转化的基因可包括任何基因,无论该基因是插入至宿主细胞的染色体中还是位于染色体外,只要该基因可在宿主细胞中表达即可。
宿主细胞包括在体内或体外用本公开内容的重组载体或多核苷酸转染、转化或感染的细胞。包含本公开内容的重组载体的宿主细胞是重组宿主细胞、重组细胞或重组微生物。
另外,根据本公开内容的重组载体可包含选择标志物。选择标志物用于选择用载体转化的转化体(宿主细胞)。由于只有表达选择标志物的细胞才可在用选择标志物处理的培养基中存活,所以可选择经转化的细胞。选择标志物的代表性实例包括但不限于卡那霉素、链霉素、氯霉素等。
插入至根据本公开内容的用于转化的重组载体中的基因可通过同源重组交叉整合至宿主细胞(例如棒状杆菌属微生物)中。
根据本公开内容的一个实施方案,宿主细胞可以是棒状杆菌属菌株,例如谷氨酸棒状杆菌菌株。
本公开内容的另一个方面提供了用于产生L-精氨酸或L-瓜氨酸的方法,其包括以下步骤:在培养基中培养谷氨酸棒状杆菌突变体菌株;以及从所培养的突变体菌株中或从培养所述突变体菌株的培养基中回收L-精氨酸或L-瓜氨酸。
可使用本领域已知的合适培养基和培养条件进行培养,并且本领域任何技术人员可容易地调整和使用培养基和培养条件。特别地,培养基可以是液体培养基,但不限于此。培养方法的一些实例包括但不限于分批培养、连续培养、补料分批培养或其组合。
根据本公开内容的一个实施方案,培养基应当以适当的方式满足特定菌株的需求,并且可由本领域技术人员进行适当的修改。针对用于棒状杆菌属微生物或菌株的培养基,可参考已知文献(但不限于此)(Manual of Methods for General Bacteriology,American Society for Bacteriology,Washington D.C.,USA,1981)。
根据本公开内容的一个实施方案,培养基可包含多种碳源、氮源和微量元素组分。可使用的碳源的一些实例包括:糖类和碳水化合物,例如葡萄糖、蔗糖、乳糖、果糖、麦芽糖、淀粉和纤维素;油和脂肪,例如大豆油、葵花籽油、蓖麻油和椰子油;脂肪酸,例如棕榈酸、硬脂酸和亚油酸;醇,例如甘油和乙醇;以及有机酸,例如乙酸。这些物质可单独使用或作为混合物使用,但不限于此。可使用的氮源的一些实例包括蛋白胨、酵母提取物、肉提取物、麦芽提取物、玉米浆、大豆粉、尿素,或者无机化合物例如硫酸铵、氯化铵、磷酸铵、碳酸铵和硝酸铵。氮源也可单独使用或作为混合物使用,但不限于此。可使用的磷源的一些实例包括但不限于磷酸二氢钾或磷酸氢二钾、或者相应的含钠盐。另外,培养基可包含但不限于生长所需的金属盐,例如硫酸镁或硫酸铁。另外,培养基可包含必需的生长物质,例如氨基酸和维生素。此外,可在培养基中使用合适的前体。可在培养期间将培养基或单独的组分通过合适的方法分批或以连续方式添加至培养基中,但不限于此。
根据本公开内容的一个实施方案,培养基的pH可通过在培养期间以适当方式向微生物培养基中添加化合物例如氢氧化铵、氢氧化钾、氨、磷酸和硫酸来调节。另外,在培养期间,可使用消泡剂例如脂肪酸聚乙二醇酯来抑制发泡。另外,为了将培养基保持在有氧条件下,可将氧气或含氧气体(例如,空气)注射至培养基中。培养基的温度通常可以是20℃至45℃,例如25℃至40℃。可继续培养直至产生所期望量的可用物质。例如,培养时间可以是10小时至160小时。
根据本公开内容的一个实施方案,在从所培养的突变体菌株中和从培养所述突变体菌株的培养基中回收L-精氨酸或L-瓜氨酸的步骤中,所产生的L-精氨酸或L-瓜氨酸可根据培养方法使用本领域已知的合适方法来从培养基中收集或回收。可使用的用于回收L-精氨酸或L-瓜氨酸的方法的实例包括但不限于离心、过滤、萃取、喷洒、干燥、蒸发、沉淀、结晶、电泳、分级溶解(例如,硫酸铵沉淀)、色谱(例如离子交换、亲和力、疏水性和尺寸排阻)等。
根据本公开内容的一个实施方案,回收L-精氨酸或L-瓜氨酸的步骤可通过将培养基在低速下离心以去除生物质并通过离子交换色谱分离所获得的上清液来进行。
根据本公开内容的一个实施方案,回收L-精氨酸或L-瓜氨酸的步骤可包括纯化L-精氨酸或L-瓜氨酸的过程。
根据本公开内容的棒状杆菌属突变体菌株具有参与L-精氨酸生物合成途径的乙酰鸟氨酸氨基转移酶的增强的活性,并且因此能够以与亲本菌株相比提高的产率来产生L-精氨酸或L-瓜氨酸。
附图说明
图1示出了根据本公开内容的一个实施方案的包含编码乙酰鸟氨酸氨基转移酶之基因的pk19msb+argD(A131T)载体的结构,所述乙酰鸟氨酸氨基转移酶具有在氨基酸位置131处的苏氨酸对丙氨酸的替换。
具体实施方式
下文中,将更详细地描述本公开内容。然而,提供这些描述仅用于说明性目的以帮助理解本公开内容,并且本公开内容的范围不受这些说明性描述的限制。
实施例1.谷氨酸棒状杆菌突变体菌株的构建
为了构建具有增强的乙酰鸟氨酸氨基转移酶活性的谷氨酸棒状杆菌突变体菌株,使用了谷氨酸棒状杆菌14GR(KCCM13219P)菌株(其是产生L-精氨酸的菌株)和大肠杆菌DH5a(HIT Competent cellsTM,目录号RH618)。
将谷氨酸棒状杆菌14GR菌株在30℃的温度下在包含以下的ARG-肉汤培养基(pH7.2)中进行培养:1L的蒸馏水中10.5g的98%葡萄糖、1g的牛肉提取物、4g的酵母提取物、2g的多聚蛋白胨(polypeptone)、2g的NaCl和40g的(NH4)2SO4。
将大肠杆菌DH5a在37℃的温度下在包含以下的LB培养基上进行培养:1L的蒸馏水中10.0g的胰蛋白胨、10.0g的NaCl和5.0g的酵母提取物。
使用的抗生素卡那霉素是Sigma的产品。
DNA测序由Macrogen,Inc.进行。
1-1.重组载体
在乙酰鸟氨酸氨基转移酶中诱导突变以增强菌株中的生物合成途径。在本实施例中使用的方法中,对编码乙酰鸟氨酸氨基转移酶的argD基因进行定点诱变以提高该基因的表达。将由argD基因编码的乙酰鸟氨酸氨基转移酶的氨基酸位置131处的丙氨酸替换为苏氨酸,并将相对于谷氨酸棒状杆菌基因组上的argD基因之中心的左臂部分(529bp)和右臂部分(525bp)通过PCR进行扩增并通过重叠PCR连接在一起,然后克隆至pk19msb载体中。将所得质粒命名为pk19msb+argD(A131T)(参见图1)。为了构建所述质粒,使用下表1中示出的引物来扩增每个基因片段。
[表1]
使用上述引物,在以下条件下进行PCR。使用Thermocycler(TP600,TAKARA BIOInc.,Japan)在包含以下的反应溶液中进行PCR:100μM的每种脱氧核苷酸三磷酸(dATP、dCTP、dGTP、dTTP)、1pM寡核苷酸、10ng的谷氨酸棒状杆菌ATCC 13032的作为模板的染色体DNA和1单位的pfu-X DNA聚合酶混合物(Solgent)。PCR进行25至30个循环,每个循环由(i)在94℃下变性30秒,(ii)在58℃下退火30秒,以及(iii)在72℃下延伸1至2分钟(聚合时间为每kb 2分钟)组成。
将如上所述制备的基因片段通过自组装克隆克隆至pk19msb载体中。将所述载体转化至大肠杆菌DH5a中,然后将其平板接种在包含50μg/ml的卡那霉素的LB-琼脂板上,并在37℃下培养24小时。分离最终形成的菌落并检查插入物是否准确地存在于载体中。接下来,分离载体并将其用于谷氨酸棒状杆菌菌株的重组。
作为上述方法中普遍执行的过程,通过PCR自谷氨酸棒状杆菌ATCC13032的基因组DNA来扩增相应基因,并根据策略通过自组装克隆方法将其插入至pk19msb载体中,并在大肠杆菌DH5a中选择所得质粒。对于染色体碱基替换,将基因片段单独扩增并通过重叠PCR连接在一起,从而制备所期望的DNA片段。对于遗传操作,使用Ex Taq聚合酶(Takara)和Pfu聚合酶(Solgent)作为PCR聚合酶,并使用购自NEB的多种限制酶和DNA修饰酶,以及根据制造商提供的缓冲液和方案来使用它们。
1-2.谷氨酸棒状杆菌突变体菌株AD1
使用克隆载体构建突变体菌株AD1。以1μg/μl或更高的终浓度制备克隆载体并将其电穿孔至谷氨酸棒状杆菌14GR菌株中(参见Tauch et al.,FEMS Microbiology letters123(1994)343-347)以诱导第一重组。在这种情况下,将经电穿孔的菌株平板接种在包含50μg/μl的卡那霉素的琼脂培养基上,分离菌落,并通过PCR和测序来检查载体是否合适地插入在基因组上的所期望位置处。将所分离的各菌株再次接种在液体培养基中以诱导第二重组,培养过夜或更长时间,并随后平板接种在包含10%蔗糖的琼脂培养基上,并分离菌落。检查最终分离的菌落对卡那霉素是否具有抗性,并随后通过测序来检查突变是否被引入至没有抗生素抗性的菌株的乙酰鸟氨酸氨基转移酶中(参见Schafer et al.,Gene 145(1994)69-73)。作为结果,构建了能够产生L-精氨酸的谷氨酸棒状杆菌突变体菌株AD1,其具有在乙酰鸟氨酸氨基转移酶的氨基酸序列(SEQ ID NO:2)中位置131处的苏氨酸对丙氨酸的替换。
示例性实施例1.谷氨酸棒状杆菌突变体菌株的L-精氨酸生产力的评价
将实施例1中构建的谷氨酸棒状杆菌突变体菌株AD1的L-精氨酸生产力与亲本菌株的L-精氨酸生产力进行比较来进行评价。
使各菌株贴附在烧瓶固体接种培养基上并在30℃下培养24小时。将所培养的各菌落接种至10-ml烧瓶滴度培养基中,并在32℃下以200rpm培养30小时。此处使用的培养基的组成在下表2中示出。培养完成之后,将各培养物用蒸馏水稀释100倍并通过0.45-μm过滤器过滤,并随后使用配备有柱(DionexIonPacTM CS12A)和UV检测器(195mm)的高效液相色谱(high-performance liquid chromatography,HPLC)(AgilentTechnologies1260Infinity,Agilent Technologies)分析所产生的L-精氨酸的量,并且结果在下表3中示出。在表3中,L-精氨酸(%)表示由各菌株产生的精氨酸的量(百分比),并且发酵产率(Yp/s)(%)表示产生的L-精氨酸的量/所消耗葡萄糖。
[表2]
[表3]
菌株 | OD610 | L-精氨酸(%) | 发酵产率(%) |
亲本菌株 | 18.0 | 1.63 | 16.31 |
AD1 | 29.0 | 2.42 | 24.25 |
如上表3中所示,确定了由于在乙酰鸟氨酸氨基转移酶的氨基酸序列中氨基酸位置131处的苏氨酸对丙氨酸的替换,谷氨酸棒状杆菌突变体菌株AD1与亲本菌株相比具有显著提高的L-精氨酸生产力。
实施例2.谷氨酸棒状杆菌突变体菌株的构建
以与实施例1中相同的方式构建具有在乙酰鸟氨酸氨基转移酶的氨基酸序列(SEQID NO:2)中位置131处的苏氨酸对丙氨酸的替换并能够产生L-瓜氨酸的谷氨酸棒状杆菌突变体菌株CD1,不同之处在于将实施例1-1的pk19msb+argD(A131T)克隆载体引入至谷氨酸棒状杆菌15GD(KCCM13220P)中来代替引入至谷氨酸棒状杆菌14GR中。
示例性实施例2.谷氨酸棒状杆菌突变体菌株的L-瓜氨酸生产力的评价
将实施例2中构建的谷氨酸棒状杆菌突变体菌株CD1的L-瓜氨酸生产力与亲本菌株的L-瓜氨酸生产力进行比较来进行评价。
使各菌株贴附在烧瓶固体接种培养基上并在30℃下培养24小时。将所培养的各菌落接种至10-ml烧瓶滴度培养基中,并在32℃下以200rpm培养30小时。此处使用的培养基的组成在下表4中示出。培养完成之后,将各培养物用蒸馏水稀释100倍并通过0.45-μm过滤器过滤,并随后使用配备有柱(DionexIonPacTM CS12A)和UV检测器(195mm)的高效液相色谱(HPLC)(Agilent Technologies 1260Infinity,Agilent Technologies)分析所产生的L-瓜氨酸的量,并且结果在下表5中示出。在表5中,L-瓜氨酸(%)表示由各菌株产生的瓜氨酸的量(百分比),并且发酵产率(Yp/s)(%)表示产生的L-瓜氨酸的量/所消耗葡萄糖。
[表4]
[表5]
菌株 | OD610 | L-瓜氨酸(%) | 发酵产率(%) |
亲本菌株 | 9.0 | 1.08 | 10.84 |
CD1 | 14.0 | 1.53 | 15.33 |
如上表5中所示,确定了由于在乙酰鸟氨酸氨基转移酶的氨基酸序列中氨基酸位置131处的苏氨酸对丙氨酸的替换,谷氨酸棒状杆菌突变体菌株CD1与亲本菌株相比具有显著提高的L-瓜氨酸生产力。
这些结果表明,通过凭借在参与L-精氨酸生物合成途径的乙酰鸟氨酸氨基转移酶的核酸序列或氨基酸序列中的定点突变而增强酶活性提高了L-精氨酸和L-瓜氨酸生产力。
迄今为止,已参照一些实施方案描述了本公开内容。本公开内容所属领域的普通技术人员将理解,在不脱离本公开内容的实质特征的情况下,本公开内容可以以经修改形式实施。因此,应从说明性的角度而不是从限制性的角度考虑所公开的实施方案。本公开内容的范围由权利要求书而不是以上描述来限定,并且与之等同的范围内的所有差异均应被解释为包括在本公开内容中。
[登记号]
保藏机构:韩国微生物保藏中心(Korean Culture Center of Microorganisms,KCCM)
登记号:KCCM13219P
保藏日期:2022年6月29日
保藏机构:韩国微生物保藏中心(KCCM)
登记号:KCCM13220P
保藏日期:2022年6月29日
Claims (5)
1.棒状杆菌属(Corynebacterium sp.)突变体菌株,其通过具有增强的乙酰鸟氨酸氨基转移酶活性而具有提高的L-精氨酸或L-瓜氨酸生产力。
2.权利要求1所述的棒状杆菌属突变体菌株,其中所述乙酰鸟氨酸氨基转移酶活性的增强是通过编码所述乙酰鸟氨酸氨基转移酶的基因中的定点突变而实现的。
3.权利要求2所述的棒状杆菌属突变体菌株,其中所述定点突变是在SEQ ID NO:2的氨基酸序列中氨基酸位置131处的苏氨酸对丙氨酸的替换。
4.权利要求1所述的棒状杆菌属突变体菌株,其是谷氨酸棒状杆菌(Corynebacteriumglutamicum)。
5.用于产生L-精氨酸或L-瓜氨酸的方法,其包括以下步骤:
在培养基中培养权利要求1所述的棒状杆菌属突变体菌株;以及
从所培养的突变体菌株中或从培养所述突变体菌株的培养基中回收L-精氨酸或L-瓜氨酸。
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
KR10-2022-0090641 | 2022-07-21 | ||
KR1020220090641A KR20240013960A (ko) | 2022-07-21 | 2022-07-21 | L-아르기닌 또는 l-시트룰린 생산능이 향상된 코리네박테리움 속 미생물 및 이를 이용한 l-아르기닌 또는 l-시트룰린의 생산 방법 |
Publications (1)
Publication Number | Publication Date |
---|---|
CN117431197A true CN117431197A (zh) | 2024-01-23 |
Family
ID=84329438
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CN202211274517.0A Pending CN117431197A (zh) | 2022-07-21 | 2022-10-18 | 具有增强的l-精氨酸或l-瓜氨酸生产力的棒状杆菌属微生物及其相关方法 |
Country Status (6)
Country | Link |
---|---|
US (1) | US20240026283A1 (zh) |
EP (1) | EP4310179A1 (zh) |
JP (1) | JP7475408B2 (zh) |
KR (1) | KR20240013960A (zh) |
CN (1) | CN117431197A (zh) |
WO (1) | WO2024019215A1 (zh) |
Family Cites Families (8)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
KR100854234B1 (ko) * | 2006-07-13 | 2008-08-25 | 씨제이제일제당 (주) | 역가가 향상된 유전자 argF 염기서열 및 그를 포함하는형질전환 균주를 이용한 L―알지닌의 생산방법 |
KR100830289B1 (ko) * | 2007-01-18 | 2008-05-16 | 씨제이제일제당 (주) | L-아르기닌 생산 변이주 및 이의 제조방법 |
KR101348461B1 (ko) * | 2010-12-08 | 2014-01-08 | 씨제이제일제당 (주) | 퓨트레신을 생산하는 미생물 및 이를 이용하여 퓨트레신을 생산하는 방법 |
JP6623690B2 (ja) | 2015-10-30 | 2019-12-25 | 味の素株式会社 | グルタミン酸系l−アミノ酸の製造法 |
KR101937569B1 (ko) | 2017-06-14 | 2019-01-11 | 씨제이제일제당 (주) | 신규 폴리펩타이드 및 이를 이용한 오르니틴계 산물 생산방법 |
KR102269637B1 (ko) | 2018-12-26 | 2021-06-28 | 대상 주식회사 | L-시트룰린 또는 l-아르기닌 생산능이 향상된 변이 균주 및 이를 이용한 l-시트룰린 또는 l-아르기닌의 제조 방법 |
CN113736719B (zh) | 2020-05-29 | 2023-04-21 | 陕西鸿道生物分析科学技术研究院有限公司 | 一种谷氨酸棒杆菌基因工程菌及其在亚精胺生产中的应用 |
CN115803442A (zh) | 2020-07-09 | 2023-03-14 | 赢创运营有限公司 | 发酵制备胍基乙酸的方法 |
-
2022
- 2022-07-21 KR KR1020220090641A patent/KR20240013960A/ko unknown
- 2022-08-17 WO PCT/KR2022/012253 patent/WO2024019215A1/ko unknown
- 2022-10-04 US US17/937,833 patent/US20240026283A1/en active Pending
- 2022-10-06 EP EP22200149.7A patent/EP4310179A1/en active Pending
- 2022-10-07 JP JP2022162571A patent/JP7475408B2/ja active Active
- 2022-10-18 CN CN202211274517.0A patent/CN117431197A/zh active Pending
Also Published As
Publication number | Publication date |
---|---|
KR20240013960A (ko) | 2024-01-31 |
JP2024014656A (ja) | 2024-02-01 |
EP4310179A1 (en) | 2024-01-24 |
US20240026283A1 (en) | 2024-01-25 |
JP7475408B2 (ja) | 2024-04-26 |
WO2024019215A1 (ko) | 2024-01-25 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
JP6998466B2 (ja) | クエン酸シンターゼの活性が弱化された変異型ポリペプチド及びそれを用いたl-アミノ酸生産方法 | |
BR112019011994B1 (pt) | Variante de proteína que tem uma atividade de exportação de inosina-5-monofosfato, polinucleotídeo, vetor, micro-organismo do gênero corynebacterium que produz inosina-5- monofosfato, método para preparar inosina-5'-monofosfato, método para aumentar a exportação de inosina-5-monofosfato e uso da proteína | |
BR112019014101B1 (pt) | Variante de proteína que tem uma atividade de exportação de inosina-5'-monofosfato, polinucleotídeo, vetor, micro-organismo do gênero corynebacterium que produz inosina-5'- monofosfato, método para preparar inosina-5'-monofosfato e uso da variante de proteína | |
ES2955417T3 (es) | Variante fosforribosil pirofosfato amidotransferasa y método para producir nucleótido de purina usando la misma | |
EP4056675A1 (en) | Mutant of corynebacterium glutamicum with enhanced l-lysine productivity and method for preparing l-lysine using the same | |
KR102589135B1 (ko) | 3-메틸-2-옥소뷰타노에이트 하이드록시 메틸트랜스퍼라아제의 활성이 강화된 미생물, 및 이의 용도 | |
JP2024503049A (ja) | GlxR蛋白質変異体またはこれを利用したスレオニン生産方法 | |
US20240026283A1 (en) | Microorganism of corynebacterium genus having enhanced l-arginine or l-citrulline productivity and a method for producing l-arginine or l-citrulline using the same | |
CN114134062A (zh) | L-赖氨酸生产能力得到提高的谷氨酸棒状杆菌突变株及利用其的l-赖氨酸的生产方法 | |
CN116171327A (zh) | 具有增强的l-瓜氨酸生产能力的谷氨酸棒状杆菌变体,以及用于使用其产生l-瓜氨酸的方法 | |
CN117431196A (zh) | 具有增强的l-精氨酸或l-瓜氨酸生产力的棒状杆菌属微生物及其相关方法 | |
EP4332229A1 (en) | Corynebacterium glutamicum variant having improved l-lysine production ability, and method for producing l-lysine using same | |
KR102434925B1 (ko) | 3-메틸-2-옥소뷰타노에이트 하이드록시 메틸트랜스퍼라아제의 활성이 강화된 미생물, 및 이의 용도 | |
CN115044490B (zh) | L-赖氨酸生产能力得到提高的谷氨酸棒状杆菌突变株及利用其的l-赖氨酸的生产方法 | |
EP4083197A1 (en) | Corynebacterium glutamicum mutant strain having enhanced l-lysine productivity and method of producing l-lysine using the same | |
US20220275412A1 (en) | Microorganism for producing l-amino acid having increased cytochrome c activity, and l-amino acid production method using same | |
CN116997655A (zh) | L-瓜氨酸生产能力得到提高的谷氨酸棒状杆菌变异株及利用其的l-瓜氨酸的生产方法 | |
JP2024515390A (ja) | L-リシン生産能が向上したコリネバクテリウム・グルタミカム変異株及びそれを用いたl-リシン生産方法 | |
KR20230084993A (ko) | L-라이신 생산능이 향상된 코리네박테리움 글루타미쿰 변이주 및 이를 이용한 l-라이신의 생산 방법 | |
JP2024513452A (ja) | 変異型SpoTタンパク質及びそれを用いたL-アミノ酸を生産する方法 | |
CN115261246A (zh) | L-赖氨酸生产能力得到提高的谷氨酸棒状杆菌突变株及利用其的l-赖氨酸的生产方法 |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
PB01 | Publication | ||
PB01 | Publication | ||
SE01 | Entry into force of request for substantive examination | ||
SE01 | Entry into force of request for substantive examination |