低温脂肪酶菌株的诱变筛选及酶学特性研究
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摘要
以本公司菌种保藏室的产脂肪酶黑曲霉菌株AG007作为出发菌株,通过常温常压等离子体(ARTP)诱变筛选得到突变株A45-72,其产脂肪酶的活力较AG007提高了86%。再对A45-72进行亚硝基胍(NTG)复合诱变,得到突变菌株GH-73,其摇瓶发酵酶活达到2 350 U/m L,较出发菌株AG007提高了1.2倍,将GH-73传到第10代,其产酶较稳定。摇瓶中添加2.5%橄榄油作为诱导剂时,产酶能力达到2 875 U/m L。通过酶学特性研究发现,其最适作用温度为30℃,且在0℃左右保留30%的活性,是一种低温脂肪酶。
With the strain for the production of lipase AG007 isolated in the company culture chamber as the original strain,the original strains AG007 were mutated by ARTP,then the new strain A45-72 was screened and their lipase productivity was 86% higher than before. Another new strain Gh-73 was screened after the strain A45-72 was mutated by NTG,the flask fermentation enzyme activity reached 2350 U/m L,1. 2times higher than the qriginal strain AG007.When the GH-73 was propagated to the 10 th generation,their lipase productivity was stable. When2.5% olive oil was added as a inducer,the lipase production ability reached 2 875 U/m L.Based on the studing of properties,it was a cold-adapted lipase,the optimal temperature of the lipase was 30 ℃.Activity of lipase was 30% at 0 ℃.
With the strain for the production of lipase AG007 isolated in the company culture chamber as the original strain,the original strains AG007 were mutated by ARTP,then the new strain A45-72 was screened and their lipase productivity was 86% higher than before. Another new strain Gh-73 was screened after the strain A45-72 was mutated by NTG,the flask fermentation enzyme activity reached 2350 U/m L,1. 2times higher than the qriginal strain AG007.When the GH-73 was propagated to the 10 th generation,their lipase productivity was stable. When2.5% olive oil was added as a inducer,the lipase production ability reached 2 875 U/m L.Based on the studing of properties,it was a cold-adapted lipase,the optimal temperature of the lipase was 30 ℃.Activity of lipase was 30% at 0 ℃.
引文
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[12]范新蕾,肖成建,顾秋亚,等.ARTP诱变选育葡萄糖氧化酶高产菌株及发酵条件优化[J].工业微生物,2015,45(1):15-19.
[13]曾璐,林亲录,王伟.脂肪酶产生菌产酶条件的研究[J].现代食品科技,2007,23(7):15-18,21.
[14]段俊英,韩静淑,柴明,等.解脂假丝酵母变株(19-2)胞内脂酶的研究[J].微生物学通报,1983(3):106-109.
[15]GUPTA R,GUPTA N,RATHI P. Bacterial lipase:An overview of production,purification and biochemical properties[J].Appl Microbiol Biotechnol,2004,64(6):763-781.
[2]DAVIS B G,BOYER B. Biocatalysis and enzymes in organic synthesis[J].Nat Prod Rep,2001,18:618-640.
[3]JAEGER K E,REETZ M T.Microbial lipases form versatile tools for biotechnology[J]. Biotechnol Appl Biochem,1998,16(9):396-403.
[4]CHAHINIAN H,VANOT A,IBRIK A,et al.Production of extracellular lipases by Penicillium cyclopium purification and characterization of a partial acylglycerol lipase[J].Biosci Biotechnol Biotechem,2000,64(2):215-222.
[5]ZHU K,JUTILA A,TUOMINEN E K J,et al.Impact of the tryptophan residues of Humicola lanuginosa lipase on its thermal stabitity[J]. Biochim Biophys Acta,2001,1547(2):329-338.
[6]孙宏丹,孟秀香,贾莉,等.微生物脂肪酶及其相关研究进展[J].大连医科大学学报,2001,23(4):292-295.
[7]FELLER G,THIRY M,ARPIGNY J L,et al.Lipases from psychrotropic antarctic bacteria[J].FEMS Microbiology Letters,1990,66(1/2/3):239-243.
[8]CHOO D W,KURIHARA T,SUZUKI T,et al. A cold-adapted lipase of an Alaskan psychrotroph,Pseudomonas sp. Strain B11-1:Cene cloning and enzyme purification and characterization[J].Applied and environmental microbiology,1998,64(2):486-491.
[9]林学政,边际,何培青.极地微生物低温适应性的分子机制[J].极地研究,2003,15(1):75-82.
[10]ARPIGNY J L,LAMOTTE J,GERDAY C.Molecular adaptation to cold of an Antarctic bacterial lipase[J].Journal of molecular catalysis B:Enzymatic,1997,3:29-35.
[11]郑铁曾,涂提坤,王建华,等.产β-半乳糖苷酶菌株的分离和筛选[J].微生物学通报,1981(6):258-260.
[12]范新蕾,肖成建,顾秋亚,等.ARTP诱变选育葡萄糖氧化酶高产菌株及发酵条件优化[J].工业微生物,2015,45(1):15-19.
[13]曾璐,林亲录,王伟.脂肪酶产生菌产酶条件的研究[J].现代食品科技,2007,23(7):15-18,21.
[14]段俊英,韩静淑,柴明,等.解脂假丝酵母变株(19-2)胞内脂酶的研究[J].微生物学通报,1983(3):106-109.
[15]GUPTA R,GUPTA N,RATHI P. Bacterial lipase:An overview of production,purification and biochemical properties[J].Appl Microbiol Biotechnol,2004,64(6):763-781.