达托霉素产生菌前体物耐受选育及其流加补料发酵
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摘要
目的通过对达托霉素产生菌进行诱变选育,以及前体物补料发酵方式提高达托霉素的产量。方法采用常压室温等离子体(atmospheric and room temperature plasma,ARTP)技术对玫瑰孢链霉菌进行诱变,以癸酸铵和甘氨酸耐受作为选择压力进行菌株筛选,在摇瓶和100L发酵罐上进行癸酸铵流加补料试验确定最佳的发酵工艺。结果经诱变选育获得1株突变株FIM-D1568摇瓶效价达到380mg/L,发酵单位较出发菌株提高了35.7%;通过优化100L发酵罐流加补料癸酸铵溶液工艺,使达托霉素发酵效价达到2276mg/L。结论以ARTP为诱变源,甘氨酸及癸酸铵耐受性为选择性压力,可以快速筛选获得达托霉素高产菌;高产突变菌株在流加补料发酵工艺上优良性状得以发挥,发酵效价大幅提高。
Objective Resistance screening on mutation breeding and fed-batch fermentation were studied to improve production of daptomycin. Methods Streptomyces roseosporus, a daptomycin-producing strain was bred by mutation of atmospheric and room temperature plasma(ARTP), with ammonium decanoate and glycine as selection factors. Adding precursor ammonium decanoate for fermentation of daptomycin were researched in a 100 L fermentor. Results A mutant strain named FIM-D1568 was obtained and the yield of daptomycin reached 380 mg/L which was 35.7% higher than that of the original one. Additionally, the yield of daptomycin reached 2276 mg/L in a 100 L fermentor by the optimization of feeding ammonium decanoate. Conclusion ARTP in combination with ammonium decanoate and glycine resistance screening can effectively improve the daptomycin production of strains.
Objective Resistance screening on mutation breeding and fed-batch fermentation were studied to improve production of daptomycin. Methods Streptomyces roseosporus, a daptomycin-producing strain was bred by mutation of atmospheric and room temperature plasma(ARTP), with ammonium decanoate and glycine as selection factors. Adding precursor ammonium decanoate for fermentation of daptomycin were researched in a 100 L fermentor. Results A mutant strain named FIM-D1568 was obtained and the yield of daptomycin reached 380 mg/L which was 35.7% higher than that of the original one. Additionally, the yield of daptomycin reached 2276 mg/L in a 100 L fermentor by the optimization of feeding ammonium decanoate. Conclusion ARTP in combination with ammonium decanoate and glycine resistance screening can effectively improve the daptomycin production of strains.
引文
[1]Debono M,Barnhart M,Carrell C B,et a1.A21978C,a complex of new acidic peptide antibiotics:Isolation,chemistry,and mass spectral structure elucidation[J].J Antibiot,1987,40(6):761-777.
[2]Vance G,Fowler,Jr M D,et a1.Daptomycin versus standard therapy for bacteremia and endocarditis caused by Staphylococcus aureus[J].N Engl J Med,2006,355(7):653-665.
[3]Barry A L,Fuchs P C,Brown S D.In vitro activities of daptomycin against 2,789 clinical isolates from 11 North American medical centers[J].Antimicrob Agents Chemother.2001,45(6):1919-1922.
[4]Steenbergen J N,Alder J,Thorne G M,et a1.Daptomycin:A lipopeptide antibiotic for the treatment of serious Grampositive infections[J].J Antimicrob Chemother,2005,55(3):283-288.
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[10]徐波,王明蓉,夏永,等.应用基因组重排育种新方法筛选替考拉宁高产菌[J].中国抗生素杂志,2006,31(4):237-243.
[11]胡海艳,黄魁英,杨冠东,等.推理选育与抗性筛选在罗咪脂高产菌的育种研究[J].中国抗生素杂志,2016,41(5):340-343.
[12]邢新会.食品微生物育种新技术—常压室温等离子体微生物基因组快速突变技术的开发难点和育种优势[J].乳业科学与技术,2013,36(1):2-3.
[13]Li G,Li H P,Wang L Y,et a1.Genetic effects of radiofrequency,atmospheric-pressure glow discharges with helium[J].Appl Phys Lett,2008,92(22):221504-1-221504-3.
[14]Zhang X,Zhang X F,Li H P,et a1.Atmospheric room temperature plasma(ARTP)as a new powerful mutagenesis tool[J].Appl Microbiol Biotechnol,2014,98(12):5387-5396.
[15]高芳霞,俞岩青,王昆蓉,等.常压室温等离子体及紫外复合诱变选育达托霉素高产菌株[J].中国抗生素杂志,2016,41(6):425-428.
[16]向荣华,詹晓北,朱莉,等.常压室温等离子体诱变绿色产色链霉菌及阿维拉霉素高产菌选育[J].中国抗生素杂志,2015,40(10):732-737.
[17]周剑,谢颖,黄建峰,等.RRLC测定发酵液中达托霉素的含量[J].福建分析测试,2012,21(1):46-49.
[18]吴国峰,李国全,马永.工业发酵分析[M].北京:化学工业出版社,2006:11-12.
[19]Huber F M,Berry D M,Pieper R L,et a1.The synthesis of A21978 by Streptomyces roseosporus cultivated under carbon limitation and fed fatty acids[J].Biotechnol Lett,1990,12(11):789-792.
[20]吴旻,王文轶,王泽根,等.达托霉素发酵过程中前体癸酸的添加策略研究[J].药物生物技术,2012,19(2):0142-0145.
[21]郭朝江,张兰,王成,等.癸酸缓释颗粒对玫瑰孢链霉菌发酵产达托霉素的影响[J].中国医药工业杂志,2013,44(3):238-241.
[22]吴远杰,王向阳,陈少欣.达托霉素的菌种选育及补料发酵工艺[J].中国医药工业杂志,2013,44(9):864-867.
[23]郭朝江,王相金,王化民.玫瑰孢链霉菌补料分批发酵生产达托霉素的动力学模型[J].中国抗生素杂志,2014,39(4):241-244.
[24]郑卫,程元荣,周剑,等.一种癸酸盐发酵达托霉素的方法:CN,101880703[P]B.20120808.
[25]周剑,黄建峰,张引,等.低能离子注入诱变达托霉素高产菌株及其发酵的研究(英文)[J].中国抗生素杂志,2012,37(8):587-592.
[2]Vance G,Fowler,Jr M D,et a1.Daptomycin versus standard therapy for bacteremia and endocarditis caused by Staphylococcus aureus[J].N Engl J Med,2006,355(7):653-665.
[3]Barry A L,Fuchs P C,Brown S D.In vitro activities of daptomycin against 2,789 clinical isolates from 11 North American medical centers[J].Antimicrob Agents Chemother.2001,45(6):1919-1922.
[4]Steenbergen J N,Alder J,Thorne G M,et a1.Daptomycin:A lipopeptide antibiotic for the treatment of serious Grampositive infections[J].J Antimicrob Chemother,2005,55(3):283-288.
[5]Debono M,Abbott B J,Molloy R M,et a1.Enzymatic and chemical modifications of lipopeptide antibiotic A21978C:the synthesis and evaluation of daptomycin(LY146032)[J].J Antibiot,1988,41(8):1093-1105.
[6]Hamill R L,Hoehn M M.A-21978 antibiotics and process for their production:US,4208403[P].1980-06-17.
[7]Miao V,Co?ffet-Legal M F,Brian P,et a1.Daptomycin biosynthesis in Streptomyces roseosporus:Cloning and analysis of the gene cluster and revision of peptide stereochemistry[J].Microbiology,2005,151(5):1507-1523.
[8]Huber F M,Pieper R L,Tietz A J.The formation of daptomycin by supplying decanoic acid to Streptomyces roseosporus cultures producing the antibiotic complex A21978C[J].J Biotechnol,1988,7(4):283-292.
[9]Bertetti G,Malcangi A,Muraca R,et a1.Process for the production of daptomycin:US,8313922[P].2012-11-20.
[10]徐波,王明蓉,夏永,等.应用基因组重排育种新方法筛选替考拉宁高产菌[J].中国抗生素杂志,2006,31(4):237-243.
[11]胡海艳,黄魁英,杨冠东,等.推理选育与抗性筛选在罗咪脂高产菌的育种研究[J].中国抗生素杂志,2016,41(5):340-343.
[12]邢新会.食品微生物育种新技术—常压室温等离子体微生物基因组快速突变技术的开发难点和育种优势[J].乳业科学与技术,2013,36(1):2-3.
[13]Li G,Li H P,Wang L Y,et a1.Genetic effects of radiofrequency,atmospheric-pressure glow discharges with helium[J].Appl Phys Lett,2008,92(22):221504-1-221504-3.
[14]Zhang X,Zhang X F,Li H P,et a1.Atmospheric room temperature plasma(ARTP)as a new powerful mutagenesis tool[J].Appl Microbiol Biotechnol,2014,98(12):5387-5396.
[15]高芳霞,俞岩青,王昆蓉,等.常压室温等离子体及紫外复合诱变选育达托霉素高产菌株[J].中国抗生素杂志,2016,41(6):425-428.
[16]向荣华,詹晓北,朱莉,等.常压室温等离子体诱变绿色产色链霉菌及阿维拉霉素高产菌选育[J].中国抗生素杂志,2015,40(10):732-737.
[17]周剑,谢颖,黄建峰,等.RRLC测定发酵液中达托霉素的含量[J].福建分析测试,2012,21(1):46-49.
[18]吴国峰,李国全,马永.工业发酵分析[M].北京:化学工业出版社,2006:11-12.
[19]Huber F M,Berry D M,Pieper R L,et a1.The synthesis of A21978 by Streptomyces roseosporus cultivated under carbon limitation and fed fatty acids[J].Biotechnol Lett,1990,12(11):789-792.
[20]吴旻,王文轶,王泽根,等.达托霉素发酵过程中前体癸酸的添加策略研究[J].药物生物技术,2012,19(2):0142-0145.
[21]郭朝江,张兰,王成,等.癸酸缓释颗粒对玫瑰孢链霉菌发酵产达托霉素的影响[J].中国医药工业杂志,2013,44(3):238-241.
[22]吴远杰,王向阳,陈少欣.达托霉素的菌种选育及补料发酵工艺[J].中国医药工业杂志,2013,44(9):864-867.
[23]郭朝江,王相金,王化民.玫瑰孢链霉菌补料分批发酵生产达托霉素的动力学模型[J].中国抗生素杂志,2014,39(4):241-244.
[24]郑卫,程元荣,周剑,等.一种癸酸盐发酵达托霉素的方法:CN,101880703[P]B.20120808.
[25]周剑,黄建峰,张引,等.低能离子注入诱变达托霉素高产菌株及其发酵的研究(英文)[J].中国抗生素杂志,2012,37(8):587-592.