JP7406831B2 - スピラマイシン産生菌、カリマイシン産生菌、作製方法、応用、及び生成物の産生量を増加させる方法 - Google Patents
スピラマイシン産生菌、カリマイシン産生菌、作製方法、応用、及び生成物の産生量を増加させる方法 Download PDFInfo
- Publication number
- JP7406831B2 JP7406831B2 JP2021535984A JP2021535984A JP7406831B2 JP 7406831 B2 JP7406831 B2 JP 7406831B2 JP 2021535984 A JP2021535984 A JP 2021535984A JP 2021535984 A JP2021535984 A JP 2021535984A JP 7406831 B2 JP7406831 B2 JP 7406831B2
- Authority
- JP
- Japan
- Prior art keywords
- spiramycin
- producing
- karimycin
- lrp
- gene
- 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
- 241000894006 Bacteria Species 0.000 title claims description 72
- ACTOXUHEUCPTEW-BWHGAVFKSA-N 2-[(4r,5s,6s,7r,9r,10r,11e,13e,16r)-6-[(2s,3r,4r,5s,6r)-5-[(2s,4r,5s,6s)-4,5-dihydroxy-4,6-dimethyloxan-2-yl]oxy-4-(dimethylamino)-3-hydroxy-6-methyloxan-2-yl]oxy-10-[(2s,5s,6r)-5-(dimethylamino)-6-methyloxan-2-yl]oxy-4-hydroxy-5-methoxy-9,16-dimethyl-2-o Chemical compound O([C@H]1/C=C/C=C/C[C@@H](C)OC(=O)C[C@@H](O)[C@@H]([C@H]([C@@H](CC=O)C[C@H]1C)O[C@H]1[C@@H]([C@H]([C@H](O[C@@H]2O[C@@H](C)[C@H](O)[C@](C)(O)C2)[C@@H](C)O1)N(C)C)O)OC)[C@@H]1CC[C@H](N(C)C)[C@@H](C)O1 ACTOXUHEUCPTEW-BWHGAVFKSA-N 0.000 title claims description 62
- 239000004187 Spiramycin Substances 0.000 title claims description 60
- 229960001294 spiramycin Drugs 0.000 title claims description 60
- 229930191512 spiramycin Natural products 0.000 title claims description 60
- 235000019372 spiramycin Nutrition 0.000 title claims description 60
- 238000004519 manufacturing process Methods 0.000 title claims description 31
- 238000000034 method Methods 0.000 title claims description 23
- 101150000411 lrp gene Proteins 0.000 claims description 67
- 108090000623 proteins and genes Proteins 0.000 claims description 36
- 238000000855 fermentation Methods 0.000 claims description 13
- 230000004151 fermentation Effects 0.000 claims description 13
- 238000004128 high performance liquid chromatography Methods 0.000 claims description 8
- 238000003860 storage Methods 0.000 claims description 8
- 239000012634 fragment Substances 0.000 claims description 7
- 230000000415 inactivating effect Effects 0.000 claims description 6
- 239000007788 liquid Substances 0.000 claims description 5
- 238000012216 screening Methods 0.000 claims description 5
- 238000010367 cloning Methods 0.000 claims description 4
- 210000000349 chromosome Anatomy 0.000 claims description 3
- 230000002779 inactivation Effects 0.000 claims description 3
- 230000036961 partial effect Effects 0.000 claims description 3
- 239000002773 nucleotide Substances 0.000 claims description 2
- 125000003729 nucleotide group Chemical group 0.000 claims description 2
- 125000003275 alpha amino acid group Chemical group 0.000 claims 1
- 238000005259 measurement Methods 0.000 description 16
- 241001217966 Streptomyces spiramyceticus Species 0.000 description 15
- 230000015572 biosynthetic process Effects 0.000 description 15
- 230000014509 gene expression Effects 0.000 description 15
- 230000006696 biosynthetic metabolic pathway Effects 0.000 description 11
- 108700005075 Regulator Genes Proteins 0.000 description 9
- 239000000047 product Substances 0.000 description 8
- 239000000243 solution Substances 0.000 description 8
- 108020004414 DNA Proteins 0.000 description 7
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 description 6
- 238000011529 RT qPCR Methods 0.000 description 6
- 150000001413 amino acids Chemical group 0.000 description 6
- 230000001105 regulatory effect Effects 0.000 description 6
- 108700039691 Genetic Promoter Regions Proteins 0.000 description 5
- QNAYBMKLOCPYGJ-REOHCLBHSA-N L-alanine Chemical compound C[C@H](N)C(O)=O QNAYBMKLOCPYGJ-REOHCLBHSA-N 0.000 description 5
- 235000004279 alanine Nutrition 0.000 description 5
- 235000001014 amino acid Nutrition 0.000 description 5
- 210000004027 cell Anatomy 0.000 description 5
- 230000006698 induction Effects 0.000 description 5
- 230000037353 metabolic pathway Effects 0.000 description 5
- 244000005700 microbiome Species 0.000 description 5
- 238000003786 synthesis reaction Methods 0.000 description 5
- ULGZDMOVFRHVEP-RWJQBGPGSA-N Erythromycin Chemical compound O([C@@H]1[C@@H](C)C(=O)O[C@@H]([C@@]([C@H](O)[C@@H](C)C(=O)[C@H](C)C[C@@](C)(O)[C@H](O[C@H]2[C@@H]([C@H](C[C@@H](C)O2)N(C)C)O)[C@H]1C)(C)O)CC)[C@H]1C[C@@](C)(OC)[C@@H](O)[C@H](C)O1 ULGZDMOVFRHVEP-RWJQBGPGSA-N 0.000 description 4
- 230000001580 bacterial effect Effects 0.000 description 4
- 239000003153 chemical reaction reagent Substances 0.000 description 4
- 238000012986 modification Methods 0.000 description 4
- 230000004048 modification Effects 0.000 description 4
- 239000006228 supernatant Substances 0.000 description 4
- 230000002103 transcriptional effect Effects 0.000 description 4
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 description 3
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 3
- 241000187747 Streptomyces Species 0.000 description 3
- 238000004458 analytical method Methods 0.000 description 3
- 230000003115 biocidal effect Effects 0.000 description 3
- 238000006243 chemical reaction Methods 0.000 description 3
- 238000004587 chromatography analysis Methods 0.000 description 3
- 230000002950 deficient Effects 0.000 description 3
- 230000000694 effects Effects 0.000 description 3
- 238000002337 electrophoretic mobility shift assay Methods 0.000 description 3
- 238000002347 injection Methods 0.000 description 3
- 239000007924 injection Substances 0.000 description 3
- 125000001501 propionyl group Chemical group O=C([*])C([H])([H])C([H])([H])[H] 0.000 description 3
- 235000018102 proteins Nutrition 0.000 description 3
- 102000004169 proteins and genes Human genes 0.000 description 3
- 239000013589 supplement Substances 0.000 description 3
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 description 2
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 2
- JUJWROOIHBZHMG-UHFFFAOYSA-N Pyridine Chemical compound C1=CC=NC=C1 JUJWROOIHBZHMG-UHFFFAOYSA-N 0.000 description 2
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 2
- 150000005693 branched-chain amino acids Chemical class 0.000 description 2
- 239000012159 carrier gas Substances 0.000 description 2
- 238000005119 centrifugation Methods 0.000 description 2
- IJOOHPMOJXWVHK-UHFFFAOYSA-N chlorotrimethylsilane Chemical compound C[Si](C)(C)Cl IJOOHPMOJXWVHK-UHFFFAOYSA-N 0.000 description 2
- 238000012217 deletion Methods 0.000 description 2
- 230000037430 deletion Effects 0.000 description 2
- 238000001962 electrophoresis Methods 0.000 description 2
- 229960003276 erythromycin Drugs 0.000 description 2
- 150000004665 fatty acids Chemical group 0.000 description 2
- 239000007789 gas Substances 0.000 description 2
- 150000002500 ions Chemical class 0.000 description 2
- 150000002596 lactones Chemical group 0.000 description 2
- 230000004060 metabolic process Effects 0.000 description 2
- 239000002547 new drug Substances 0.000 description 2
- 239000013612 plasmid Substances 0.000 description 2
- 239000002244 precipitate Substances 0.000 description 2
- 239000002243 precursor Substances 0.000 description 2
- 238000002360 preparation method Methods 0.000 description 2
- 238000002415 sodium dodecyl sulfate polyacrylamide gel electrophoresis Methods 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- 238000003260 vortexing Methods 0.000 description 2
- KDELTXNPUXUBMU-UHFFFAOYSA-N 2-[2-[bis(carboxymethyl)amino]ethyl-(carboxymethyl)amino]acetic acid boric acid Chemical compound OB(O)O.OB(O)O.OB(O)O.OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KDELTXNPUXUBMU-UHFFFAOYSA-N 0.000 description 1
- VTIKDEXOEJDMJP-UHFFFAOYSA-N Actinorhodine Natural products CC1OC(CC(=O)O)CC2=C1C(=O)c3c(O)c(cc(O)c3C2=O)c4cc(O)c5C(=O)C6=C(C(C)OC(CC(=O)O)C6)C(=O)c5c4O VTIKDEXOEJDMJP-UHFFFAOYSA-N 0.000 description 1
- USFZMSVCRYTOJT-UHFFFAOYSA-N Ammonium acetate Chemical compound N.CC(O)=O USFZMSVCRYTOJT-UHFFFAOYSA-N 0.000 description 1
- 239000005695 Ammonium acetate Substances 0.000 description 1
- 241000203069 Archaea Species 0.000 description 1
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 1
- GIDHDUYAXIVIMN-JJORJHTQSA-N CCC(=O)O[C@@H]1CC(=O)O[C@H](C)C\C=C\C=C\[C@H](O[C@H]2CC[C@@H]([C@@H](C)O2)N(C)C)[C@H](C)C[C@H](CC=O)[C@H](O[C@@H]2O[C@H](C)[C@@H](O[C@H]3C[C@@](C)(O)[C@@H](OC(=O)CC(C)C)[C@H](C)O3)[C@@H]([C@H]2O)N(C)C)[C@H]1OC Chemical compound CCC(=O)O[C@@H]1CC(=O)O[C@H](C)C\C=C\C=C\[C@H](O[C@H]2CC[C@@H]([C@@H](C)O2)N(C)C)[C@H](C)C[C@H](CC=O)[C@H](O[C@@H]2O[C@H](C)[C@@H](O[C@H]3C[C@@](C)(O)[C@@H](OC(=O)CC(C)C)[C@H](C)O3)[C@@H]([C@H]2O)N(C)C)[C@H]1OC GIDHDUYAXIVIMN-JJORJHTQSA-N 0.000 description 1
- PNMQDIDLWTXQLZ-RZELCZIWSA-N CO[C@H]1[C@@H](CC(=O)O[C@H](C)C\C=C\C=C\[C@H](O[C@H]2CC[C@@H]([C@@H](C)O2)N(C)C)[C@H](C)C[C@H](CC=O)[C@@H]1O[C@@H]1O[C@H](C)[C@@H](O[C@H]2C[C@@](C)(O)[C@@H](OC(=O)CC(C)C)[C@H](C)O2)[C@@H]([C@H]1O)N(C)C)OC(C)=O Chemical compound CO[C@H]1[C@@H](CC(=O)O[C@H](C)C\C=C\C=C\[C@H](O[C@H]2CC[C@@H]([C@@H](C)O2)N(C)C)[C@H](C)C[C@H](CC=O)[C@@H]1O[C@@H]1O[C@H](C)[C@@H](O[C@H]2C[C@@](C)(O)[C@@H](OC(=O)CC(C)C)[C@H](C)O2)[C@@H]([C@H]1O)N(C)C)OC(C)=O PNMQDIDLWTXQLZ-RZELCZIWSA-N 0.000 description 1
- GCIQAGXAZPUQNA-YDPAJNKASA-N CO[C@H]1[C@H](O)CC(=O)O[C@H](C)C\C=C\C=C\[C@H](O[C@H]2CC[C@@H]([C@@H](C)O2)N(C)C)[C@H](C)C[C@H](CC=O)[C@@H]1O[C@@H]1O[C@H](C)[C@@H](O[C@H]2C[C@@](C)(O)[C@@H](OC(=O)CC(C)C)[C@H](C)O2)[C@@H]([C@H]1O)N(C)C Chemical compound CO[C@H]1[C@H](O)CC(=O)O[C@H](C)C\C=C\C=C\[C@H](O[C@H]2CC[C@@H]([C@@H](C)O2)N(C)C)[C@H](C)C[C@H](CC=O)[C@@H]1O[C@@H]1O[C@H](C)[C@@H](O[C@H]2C[C@@](C)(O)[C@@H](OC(=O)CC(C)C)[C@H](C)O2)[C@@H]([C@H]1O)N(C)C GCIQAGXAZPUQNA-YDPAJNKASA-N 0.000 description 1
- 229930188120 Carbomycin Natural products 0.000 description 1
- 241000047489 Cohnella thermotolerans Species 0.000 description 1
- 102000007260 Deoxyribonuclease I Human genes 0.000 description 1
- 108010008532 Deoxyribonuclease I Proteins 0.000 description 1
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 241000588724 Escherichia coli Species 0.000 description 1
- 241001198387 Escherichia coli BL21(DE3) Species 0.000 description 1
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 1
- SNDPXSYFESPGGJ-BYPYZUCNSA-N L-2-aminopentanoic acid Chemical compound CCC[C@H](N)C(O)=O SNDPXSYFESPGGJ-BYPYZUCNSA-N 0.000 description 1
- SNDPXSYFESPGGJ-UHFFFAOYSA-N L-norVal-OH Natural products CCCC(N)C(O)=O SNDPXSYFESPGGJ-UHFFFAOYSA-N 0.000 description 1
- 108010025358 Leucine Responsive Regulatory Protein Proteins 0.000 description 1
- XCOBLONWWXQEBS-KPKJPENVSA-N N,O-bis(trimethylsilyl)trifluoroacetamide Chemical compound C[Si](C)(C)O\C(C(F)(F)F)=N\[Si](C)(C)C XCOBLONWWXQEBS-KPKJPENVSA-N 0.000 description 1
- 238000012408 PCR amplification Methods 0.000 description 1
- 238000002123 RNA extraction Methods 0.000 description 1
- 241000187559 Saccharopolyspora erythraea Species 0.000 description 1
- 229910000831 Steel Inorganic materials 0.000 description 1
- 241000958209 Streptomyces flavidovirens Species 0.000 description 1
- 241001600136 Streptomyces thermotolerans Species 0.000 description 1
- 108090000992 Transferases Proteins 0.000 description 1
- FQVHOULQCKDUCY-OGHXVOSASA-N [(2s,3s,4r,6s)-6-[(2r,3s,4r,5r,6s)-6-[[(1s,3r,7r,8s,9s,10r,12r,14e,16s)-7-acetyloxy-8-methoxy-3,12-dimethyl-5,13-dioxo-10-(2-oxoethyl)-4,17-dioxabicyclo[14.1.0]heptadec-14-en-9-yl]oxy]-4-(dimethylamino)-5-hydroxy-2-methyloxan-3-yl]oxy-4-hydroxy-2,4-dimeth Chemical compound O([C@@H]1[C@@H](C)O[C@H]([C@@H]([C@H]1N(C)C)O)O[C@H]1[C@@H](CC=O)C[C@@H](C)C(=O)/C=C/[C@@H]2O[C@H]2C[C@@H](C)OC(=O)C[C@H]([C@@H]1OC)OC(C)=O)[C@H]1C[C@@](C)(O)[C@@H](OC(=O)CC(C)C)[C@H](C)O1 FQVHOULQCKDUCY-OGHXVOSASA-N 0.000 description 1
- 125000002777 acetyl group Chemical group [H]C([H])([H])C(*)=O 0.000 description 1
- VTIKDEXOEJDMJP-WYUUTHIRSA-N actinorhodin Chemical compound C([C@@H](CC(O)=O)O[C@@H]1C)C(C(C2=C(O)C=3)=O)=C1C(=O)C2=C(O)C=3C(C(=C1C2=O)O)=CC(O)=C1C(=O)C1=C2[C@@H](C)O[C@H](CC(O)=O)C1 VTIKDEXOEJDMJP-WYUUTHIRSA-N 0.000 description 1
- 230000003213 activating effect Effects 0.000 description 1
- 239000004480 active ingredient Substances 0.000 description 1
- 230000037354 amino acid metabolism Effects 0.000 description 1
- 229940043376 ammonium acetate Drugs 0.000 description 1
- 235000019257 ammonium acetate Nutrition 0.000 description 1
- 238000000137 annealing Methods 0.000 description 1
- 239000003242 anti bacterial agent Substances 0.000 description 1
- 229940088710 antibiotic agent Drugs 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 125000001797 benzyl group Chemical group [H]C1=C([H])C([H])=C(C([H])=C1[H])C([H])([H])* 0.000 description 1
- 238000003766 bioinformatics method Methods 0.000 description 1
- 229940098773 bovine serum albumin Drugs 0.000 description 1
- 238000010805 cDNA synthesis kit Methods 0.000 description 1
- 229950005779 carbomycin Drugs 0.000 description 1
- 238000003776 cleavage reaction Methods 0.000 description 1
- 239000002299 complementary DNA Substances 0.000 description 1
- 238000010276 construction Methods 0.000 description 1
- 230000007423 decrease Effects 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 230000003111 delayed effect Effects 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 210000003495 flagella Anatomy 0.000 description 1
- 239000012530 fluid Substances 0.000 description 1
- JGBUYEVOKHLFID-UHFFFAOYSA-N gelred Chemical compound [I-].[I-].C=1C(N)=CC=C(C2=CC=C(N)C=C2[N+]=2CCCCCC(=O)NCCCOCCOCCOCCCNC(=O)CCCCC[N+]=3C4=CC(N)=CC=C4C4=CC=C(N)C=C4C=3C=3C=CC=CC=3)C=1C=2C1=CC=CC=C1 JGBUYEVOKHLFID-UHFFFAOYSA-N 0.000 description 1
- 230000002068 genetic effect Effects 0.000 description 1
- 238000010353 genetic engineering Methods 0.000 description 1
- 238000010362 genome editing Methods 0.000 description 1
- 235000011187 glycerol Nutrition 0.000 description 1
- 238000010438 heat treatment Methods 0.000 description 1
- 239000001307 helium Substances 0.000 description 1
- 229910052734 helium Inorganic materials 0.000 description 1
- SWQJXJOGLNCZEY-UHFFFAOYSA-N helium atom Chemical compound [He] SWQJXJOGLNCZEY-UHFFFAOYSA-N 0.000 description 1
- 230000013632 homeostatic process Effects 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- 238000003384 imaging method Methods 0.000 description 1
- 238000003780 insertion Methods 0.000 description 1
- 230000037431 insertion Effects 0.000 description 1
- BPHPUYQFMNQIOC-NXRLNHOXSA-N isopropyl beta-D-thiogalactopyranoside Chemical compound CC(C)S[C@@H]1O[C@H](CO)[C@H](O)[C@H](O)[C@H]1O BPHPUYQFMNQIOC-NXRLNHOXSA-N 0.000 description 1
- 239000007791 liquid phase Substances 0.000 description 1
- 239000003120 macrolide antibiotic agent Substances 0.000 description 1
- 239000002075 main ingredient Substances 0.000 description 1
- 239000003550 marker Substances 0.000 description 1
- 238000000691 measurement method Methods 0.000 description 1
- 238000002844 melting Methods 0.000 description 1
- 230000008018 melting Effects 0.000 description 1
- 125000000956 methoxy group Chemical group [H]C([H])([H])O* 0.000 description 1
- 230000000813 microbial effect Effects 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 230000000877 morphologic effect Effects 0.000 description 1
- 230000035772 mutation Effects 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- 239000012074 organic phase Substances 0.000 description 1
- 239000012071 phase Substances 0.000 description 1
- 230000007180 physiological regulation Effects 0.000 description 1
- 229920002401 polyacrylamide Polymers 0.000 description 1
- 229920000768 polyamine Polymers 0.000 description 1
- 230000001737 promoting effect Effects 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- UMJSCPRVCHMLSP-UHFFFAOYSA-N pyridine Natural products COC1=CC=CN=C1 UMJSCPRVCHMLSP-UHFFFAOYSA-N 0.000 description 1
- 238000004445 quantitative analysis Methods 0.000 description 1
- 230000006798 recombination Effects 0.000 description 1
- 238000005215 recombination Methods 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 238000012552 review Methods 0.000 description 1
- 230000000630 rising effect Effects 0.000 description 1
- 230000007017 scission Effects 0.000 description 1
- 230000024053 secondary metabolic process Effects 0.000 description 1
- 229930000044 secondary metabolite Natural products 0.000 description 1
- 238000012163 sequencing technique Methods 0.000 description 1
- 239000011780 sodium chloride Substances 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 239000002904 solvent Substances 0.000 description 1
- 239000012086 standard solution Substances 0.000 description 1
- 239000010959 steel Substances 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 230000001629 suppression Effects 0.000 description 1
- 210000001519 tissue Anatomy 0.000 description 1
- 238000013518 transcription Methods 0.000 description 1
- 230000035897 transcription Effects 0.000 description 1
- 230000001131 transforming effect Effects 0.000 description 1
- 238000012795 verification Methods 0.000 description 1
- 230000007923 virulence factor Effects 0.000 description 1
- 239000000304 virulence factor Substances 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P19/00—Preparation of compounds containing saccharide radicals
- C12P19/44—Preparation of O-glycosides, e.g. glucosides
- C12P19/60—Preparation of O-glycosides, e.g. glucosides having an oxygen of the saccharide radical directly bound to a non-saccharide heterocyclic ring or a condensed ring system containing a non-saccharide heterocyclic ring, e.g. coumermycin, novobiocin
- C12P19/62—Preparation of O-glycosides, e.g. glucosides having an oxygen of the saccharide radical directly bound to a non-saccharide heterocyclic ring or a condensed ring system containing a non-saccharide heterocyclic ring, e.g. coumermycin, novobiocin the hetero ring having eight or more ring members and only oxygen as ring hetero atoms, e.g. erythromycin, spiramycin, nystatin
-
- 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
- C12N15/76—Vectors or expression systems specially adapted for prokaryotic hosts other than E. coli, e.g. Lactobacillus, Micromonospora for Actinomyces; for Streptomyces
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/195—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from bacteria
- C07K14/36—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from bacteria from Actinomyces; from Streptomyces (G)
-
- 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
- C12N2800/00—Nucleic acids vectors
- C12N2800/10—Plasmid DNA
- C12N2800/101—Plasmid DNA for bacteria
-
- 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
- C12R2001/465—Streptomyces
Landscapes
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Genetics & Genomics (AREA)
- Engineering & Computer Science (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)
- Biophysics (AREA)
- Molecular Biology (AREA)
- General Chemical & Material Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Medicinal Chemistry (AREA)
- Physics & Mathematics (AREA)
- Plant Pathology (AREA)
- Gastroenterology & Hepatology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Tropical Medicine & Parasitology (AREA)
- Virology (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Description
イソバレリルスピラマイシンIII:R=COCH2CH3
R′=COCH2CH(CH3)2
イソバレリルスピラマイシンII :R=COCH3
R′=COCH2CH(CH3)2
イソバレリルスピラマイシンI :R=H
R′=COCH2CH(CH3)2
95℃ 10s
65℃ 60s
97℃ 1s
Claims (10)
- 菌株内のLrp遺伝子が不活化しており、前記菌株の保管番号がCGMCC No.16056であることを特徴とするスピラマイシン産生菌。
- 菌株内のLrp遺伝子が不活化しており、前記菌株の保管番号がCGMCC No.16055であることを特徴とするカリマイシン産生菌。
- 前記Lrpタンパク質のアミノ酸配列はSeq.2で示されることを特徴とする、請求項1に記載のスピラマイシン産生菌、又は請求項2に記載のカリマイシン産生菌。
- 前記Lrp遺伝子のヌクレオチド配列はSeq.1で示されることを特徴とする、請求項3に記載のスピラマイシン産生菌、又は請求項3に記載のカリマイシン産生菌。
- 前記Lrp遺伝子の不活性化が全部のノックアウトにより行われることを特徴とする、請求項1に記載のスピラマイシン産生菌、又は請求項2に記載のカリマイシン産生菌。
- スピラマイシン産生菌及び/又はカリマイシン産生菌の作製方法であって、
(1)Lrp遺伝子を不活化するための組換えベクターを作製して、スピラマイシン産生菌及び/又はカリマイシン産生菌に導入し、
(2)Lrp遺伝子が不活化した組換え菌をスクリーニングにより取得すること
を含み、
スピラマイシン産生菌又はカリマイシン産生菌のゲノムを鋳型とし、全部又は部分的にLrp遺伝子をノックアウトすることを原則に、適切な部位にプライマーを設計して、左右のアームにおける遺伝子断片をPCR増幅し、ノックアウトベクターにクローニングすることで組換えベクターを作製し、これをスピラマイシン産生菌及び/又はカリマイシン産生菌に導入して、Lrp遺伝子が不活化した組換え菌を耐性及び継代スクリーニングにより取得することを特徴とする、作製方法。 - スピラマイシン及び/又はカリマイシンの成分IIIの産生量を増加させる方法であって、
(1)スピラマイシン産生菌又はカリマイシン産生菌の染色体上のLrp遺伝子を改変してLrp遺伝子を不活化させ、
(2)ステップ(1)で改変したスピラマイシン産生菌又はカリマイシン産生菌を用いて発酵生産を行うこと
を含む、方法。 - スピラマイシン産生菌又はカリマイシン産生菌におけるLrp遺伝子を不活化したあと、スピラマイシン生合成経路関連遺伝子、アラニン生合成経路関連遺伝子、及びアシルCoA代謝経路関連遺伝子の発現を向上させることで、スピラマイシン及び/又はカリマイシンの成分IIIの産生量を増加させることを特徴とする、請求項7記載の方法。
- 改変したスピラマイシン産生菌又はカリマイシン産生菌を種培養、発酵培養したあとに発酵液を抽出し、取得した発酵生成物をHPLCで測定分析することを特徴とする請求項8に記載の方法。
- Lrp遺伝子の不活化は、部分的なノックアウト又は全部のノックアウトにより行われることを特徴とする、請求項7又は8に記載の方法。
Applications Claiming Priority (3)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN201811577592.8 | 2018-12-20 | ||
CN201811577592.8A CN111349595A (zh) | 2018-12-20 | 2018-12-20 | 一种螺旋霉素产生菌、可利霉素产生菌、构建方法、应用及提高产物产量的方法 |
PCT/CN2019/124724 WO2020125531A1 (zh) | 2018-12-20 | 2019-12-12 | 一种螺旋霉素产生菌、可利霉素产生菌、构建方法、应用及提高产物产量的方法 |
Publications (2)
Publication Number | Publication Date |
---|---|
JP2022524688A JP2022524688A (ja) | 2022-05-10 |
JP7406831B2 true JP7406831B2 (ja) | 2023-12-28 |
Family
ID=71102067
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
JP2021535984A Active JP7406831B2 (ja) | 2018-12-20 | 2019-12-12 | スピラマイシン産生菌、カリマイシン産生菌、作製方法、応用、及び生成物の産生量を増加させる方法 |
Country Status (8)
Country | Link |
---|---|
US (1) | US20230159971A1 (ja) |
EP (1) | EP3940056A4 (ja) |
JP (1) | JP7406831B2 (ja) |
KR (1) | KR20210108411A (ja) |
CN (1) | CN111349595A (ja) |
AU (1) | AU2019408296A1 (ja) |
CA (1) | CA3124477A1 (ja) |
WO (1) | WO2020125531A1 (ja) |
Citations (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JP2006501863A (ja) | 2002-10-08 | 2006-01-19 | アベンティス・ファーマ・ソシエテ・アノニム | スピラマイシンの生合成に関与するポリペプチド、これらポリペプチドをコードするヌクレオチド配列、および、その使用 |
JP2013532981A (ja) | 2010-07-23 | 2013-08-22 | シェンヤン・トンリアン・グループ・カンパニー・リミテッド | 一株イソバレリルスピラマイシンi成分遺伝子工程菌wsj−ia |
Family Cites Families (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
EP1905833B1 (fr) * | 2002-10-08 | 2015-08-12 | Aventis Pharma S.A. | Polypeptides impliqués dans la biosynthèse des spiramycines, séquences nucléotidiques codant ces polypeptides et leurs applications |
CN1169947C (zh) * | 2002-11-19 | 2004-10-06 | 中国医学科学院医药生物技术研究所 | 必特螺旋霉素的基因工程菌株螺旋霉素链霉菌wsj-195 |
CN101054553A (zh) * | 2007-04-09 | 2007-10-17 | 中国医学科学院医药生物技术研究所 | 异戊酰螺旋霉素i基因工程菌株的构建 |
CN107868789B (zh) * | 2015-12-31 | 2020-11-17 | 沈阳福洋医药科技有限公司 | 可利霉素生物合成基因簇 |
-
2018
- 2018-12-20 CN CN201811577592.8A patent/CN111349595A/zh active Pending
-
2019
- 2019-12-12 AU AU2019408296A patent/AU2019408296A1/en not_active Abandoned
- 2019-12-12 JP JP2021535984A patent/JP7406831B2/ja active Active
- 2019-12-12 US US17/416,134 patent/US20230159971A1/en not_active Abandoned
- 2019-12-12 KR KR1020217022463A patent/KR20210108411A/ko not_active Application Discontinuation
- 2019-12-12 CA CA3124477A patent/CA3124477A1/en active Pending
- 2019-12-12 EP EP19899645.6A patent/EP3940056A4/en not_active Withdrawn
- 2019-12-12 WO PCT/CN2019/124724 patent/WO2020125531A1/zh unknown
Patent Citations (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JP2006501863A (ja) | 2002-10-08 | 2006-01-19 | アベンティス・ファーマ・ソシエテ・アノニム | スピラマイシンの生合成に関与するポリペプチド、これらポリペプチドをコードするヌクレオチド配列、および、その使用 |
JP2013532981A (ja) | 2010-07-23 | 2013-08-22 | シェンヤン・トンリアン・グループ・カンパニー・リミテッド | 一株イソバレリルスピラマイシンi成分遺伝子工程菌wsj−ia |
Non-Patent Citations (2)
Title |
---|
LIU, Jing, et al.,Applied Microbiology and Biotechnology,2017年,Vol. 101,pp. 5773-5783 |
LIU, Jing, et al.,Metabolic Engineering,2017年,Vol. 39,pp. 29-37 |
Also Published As
Publication number | Publication date |
---|---|
EP3940056A4 (en) | 2022-08-03 |
EP3940056A1 (en) | 2022-01-19 |
JP2022524688A (ja) | 2022-05-10 |
KR20210108411A (ko) | 2021-09-02 |
US20230159971A1 (en) | 2023-05-25 |
AU2019408296A1 (en) | 2021-08-05 |
CN111349595A (zh) | 2020-06-30 |
WO2020125531A1 (zh) | 2020-06-25 |
CA3124477A1 (en) | 2020-06-25 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
Liang et al. | Biosensor-assisted transcriptional regulator engineering for Methylobacterium extorquens AM1 to improve mevalonate synthesis by increasing the acetyl-CoA supply | |
Liu et al. | Structure and biosynthesis of fumosorinone, a new protein tyrosine phosphatase 1B inhibitor firstly isolated from the entomogenous fungus Isaria fumosorosea | |
TW200835792A (en) | Process for the biological production of n-butanol with high yield | |
Shao et al. | Manipulating natural product biosynthetic pathways via DNA assembler | |
CN111235169A (zh) | 一种GTP环化水解酶I基因folE及应用 | |
CN108463555A (zh) | 生产苯甲醛的方法 | |
CN110494567A (zh) | 生产rna的方法 | |
JP2023507891A (ja) | リパーゼ改変株 | |
BR112014020852B1 (pt) | Método para a produção de um hidrocarboneto | |
Lee et al. | A sigma-Like Factor Responsible for Carotenoid Biosynthesis in Streptomyces griseus | |
JP2023500781A (ja) | カンナビノイド類の生成のための操作された微生物 | |
CN110760453B (zh) | 一种高产乙酸苯乙酯的基因工程酵母菌株及其构建方法和生产乙酸苯乙酯的方法 | |
Friedmann et al. | Properties of succinyl-coenzyme A: L-malate coenzyme A transferase and its role in the autotrophic 3-hydroxypropionate cycle of Chloroflexus aurantiacus | |
CN111117942B (zh) | 一种产林可霉素的基因工程菌及其构建方法和应用 | |
JP7406831B2 (ja) | スピラマイシン産生菌、カリマイシン産生菌、作製方法、応用、及び生成物の産生量を増加させる方法 | |
CN109321618B (zh) | 一种通过糖多孢红霉菌sace_5717基因提高红霉素产量的方法 | |
CN114262695B (zh) | 一种生产cbga前体的酿酒酵母工程菌及其构建方法和应用 | |
Segura et al. | Isolation and characterization of Azotobacter vinelandii mutants impaired in alkylresorcinol synthesis: alkylresorcinols are not essential for cyst desiccation resistance | |
CN112375725B (zh) | 一种生产维生素b6的代谢工程菌株及其构建方法与应用 | |
Chen et al. | HetI-Like Phosphopantetheinyl Transferase Posttranslationally Modifies Acyl Carrier Proteins in Xanthomonas spp. | |
CN112410353B (zh) | 一种fkbS基因、含其的基因工程菌及其制备方法和用途 | |
CN109593740B (zh) | 一种糖基转移酶及其应用 | |
CN115873775B (zh) | 一种基于调控蛋白BldD翻译后修饰提升放线菌次级代谢能力的方法 | |
Jin et al. | Probing the molecular mechanisms for pristinamycin yield enhancement in Streptomyces pristinaespiralis | |
Sinkler et al. | Metabolic engineering of E. coli: influence of gene deletions and heterologous genes on physiological traits |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
A521 | Request for written amendment filed |
Free format text: JAPANESE INTERMEDIATE CODE: A523 Effective date: 20210823 |
|
A621 | Written request for application examination |
Free format text: JAPANESE INTERMEDIATE CODE: A621 Effective date: 20211206 |
|
A131 | Notification of reasons for refusal |
Free format text: JAPANESE INTERMEDIATE CODE: A131 Effective date: 20221115 |
|
A601 | Written request for extension of time |
Free format text: JAPANESE INTERMEDIATE CODE: A601 Effective date: 20230215 |
|
A601 | Written request for extension of time |
Free format text: JAPANESE INTERMEDIATE CODE: A601 Effective date: 20230414 |
|
A521 | Request for written amendment filed |
Free format text: JAPANESE INTERMEDIATE CODE: A523 Effective date: 20230424 |
|
A521 | Request for written amendment filed |
Free format text: JAPANESE INTERMEDIATE CODE: A821 Effective date: 20230424 |
|
A131 | Notification of reasons for refusal |
Free format text: JAPANESE INTERMEDIATE CODE: A131 Effective date: 20230704 |
|
A521 | Request for written amendment filed |
Free format text: JAPANESE INTERMEDIATE CODE: A523 Effective date: 20230914 |
|
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: 20231121 |
|
A61 | First payment of annual fees (during grant procedure) |
Free format text: JAPANESE INTERMEDIATE CODE: A61 Effective date: 20231211 |
|
R150 | Certificate of patent or registration of utility model |
Ref document number: 7406831 Country of ref document: JP Free format text: JAPANESE INTERMEDIATE CODE: R150 |