耐甲醇脂肪酶产生菌的筛选鉴定及其酶学性质研究  

Screening of Methanol-tolerant Lipase-producing Strains and Study of Properties of Lipase

作　　者：王斌[1] 宗浩[1] 张志敏[2] 王琰[2] 万一[2] 

机构地区：[1]四川师范大学生命科学院,四川成都610068 [2]陕西省微生物研究所,陕西西安710043

出　　处：《安徽农业科学》2011年第20期12001-12004,共4页Journal of Anhui Agricultural Sciences

基　　金：陕西省科学院青年基金项目(2009k-27)

摘　　要：[目的]筛选耐甲醇脂肪酶产生菌,并对其酶学性质进行研究。[方法]采用培养基中添加不同浓度甲醇（0、5%、10%、15%、20%）的方法筛选甲醇耐受性脂肪酶产生菌,形态学和ITS序列分析确定菌株的种属关系,硫酸铵分级沉淀和DEAE-Sepharose FF纯化脂肪酶,并对纯化的酶进行耐甲醇试验和酶学性质研究。[结果]筛选到183株脂肪酶产生菌,菌株SXL-107对浓度20%甲醇具有较好耐受性,脂肪酶活力达104 U/ml;ITS同源性分析确定该菌为Galactomyces geotrichum;经过纯化的SXL-107脂肪酶在浓度10%甲醇溶液中作用48 h后,剩余酶活为96.15%;该酶最适温度为40℃,pH为7.0,50℃温浴60 min仍有80%的酶活力,在pH 4.0~9.0间稳定,微量的Mg2＋、K＋和Zn2＋对该酶有激活作用,Mn2＋和CO2＋有抑制作用,分子量约为66 kDa。[结论]筛选到一株耐甲醇能力较强的脂肪酶产生菌,为该酶在生物柴油中的应用奠定基础。[Objective] The methanol-tolerant lipase-producing strains was screened and the properties of lipase was studied.[Method] Methanol-tolerant lipase-producing strains were screened with the method to add different concentrations of methanol（0.5%,10%,15%,20%） to the medium.SXL-107 was identified as Galactomyces geotrichum by its morphological and ITS sequences.The lipase was depurated by ammonium sulfate fractionation and DEAE-Sepharose FF.Properties of lipase and methanol-tolerance of purified enzyme were studied.[Result] 183 lipase-producing strains were isolated from rich oil soil.The strain SXL-107 had a better tolerance to 20% methanol.The enzyme activity of SXL-107 achieved 104 U/ml.It was identified as Galactomyces geotrichum by its morphological and ITS sequences.When SXL-107 lipase reacted in 10% methanol for 48 h,the remaining activity was 96.15%.The optimal pH was 7.0 and the temperature of the lipase was 40 ℃.The activity of the lipase reached 80% at 50 ℃ for 60 min,and was stable from pH 4.0 to 9.0.The enzyme activity was stimulated by Mg2＋,K＋and Zn2＋,but inhibited by Mn2＋ and CO2＋.The molecular weight of the purified SXL-107 lipase was about 66 kDa.[Conclusion] A Methanol-tolerant lipase-producing strain was screened and it provided the foundation for bio-diesel production.

关 键 词：脂肪酶 菌种筛选 鉴定 耐甲醇 酶学性质 

分 类 号：S188.4[农业科学—农业基础科学]