Saccharomy cescerevisiae氨基转移酶基因ARO8克隆及对3甲硫基丙醇合成的影响  

Cloning of the aminotransferase gene ARO8 and its influence on the production of methionol in Saccharomyces cerevisiae

作　　者：刘丽[1] 尹胜[1,2] 梁婧如 王成涛[1,2] 赵吉兴 

机构地区：[1]北京工商大学北京市食品添加剂工程技术研究中心,北京100048 [2]北京市食品风味化学重点实验室,北京100048 [3]山东中惠食品有限公司,山东惠民251706

出　　处：《食品工业科技》2014年第9期132-135,139,共5页Science and Technology of Food Industry

基　　金：国家自然科学基金项目(31071593);教育部科学技术研究重点项目(211101);北京市教委科技面上项目(KM201110011001)

摘　　要：酿酒酵母可通过艾希利(Ehrlich)途径将L-蛋氨酸转化为3-甲硫基丙醇等食品风味成分,本文研究了氨基转移酶及其基因在Ehrlich途径及3-甲硫基丙醇合成代谢的调控作用。从Saccharomyces cerevisiae S288C克隆氨基转移酶ARO8基因,并基于酿酒酵母-大肠杆菌穿梭型表达载体pYES-pgk,构建重组质粒表达载体pYES-pgk-ARO8,PEG/LiAc转化法将其导入S.cerevisiae S288C。结果表明,克隆的ARO8基因全长1503bp,与NCBI GenBank中酿酒酵母芳香族氨基转移酶Ⅰ编码基因ARO8的序列相似度为100%;构建的ARO8基因过表达工程菌S.cerevisiae AR8,其氨基转移酶活力为187U/mL,较野生型S288C菌株的酶活性提高1.63倍;工程菌AR8的摇瓶发酵3-甲硫基丙醇产量为0.76g/L,较野生型S288C提高28.8%。表明氨基转移酶Aro8p及其ARO8基因在S.cerevisiae 3-甲硫基丙醇合成代谢中发挥重要作用,增强其ARO8基因表达,有助于提高3-甲硫基丙醇的产量。Methionol is an important volatile sulphur flavor compound.It can be produced in Saccharomyces cerevisiae using L-methionine by the Ehrlich pathway.To verify aminotransferase＇ effect, gene AR08 encoding aminotransferase was cloned from S.cerevisiae S288C, and the gene ARO8 was inserted into the S.cerevisiae- E.coli shuttle constitutive expression vector pYES-pgk, generating a recombinant vector pYES-pgk-AROS.The vector pYES-pgk-ARO8 was then transformed into S.cerevisiae S288C.The results indicated that the encoding gene shared 100% homology with the aromatic aminotransferase I gene ARC8 reported in NCBI GenBank.A recombinant strain S.cerevisiae AR8 was constructed and screened successfully.The aminotransferase activity of S. cerevisiae AR8 was 187 U/m L, which was increased by 2.8.8 % compared to what in S. cerevisiae S288 C. Methionol yield of AR8 in shaking flask reached 0.76g/L and it was increased by 28.8% .These results showed that aminotransferase Aro8p and gene ARO8 of S.cerevisiae played an important role in methionol anabolism,and the gene AR08 over-expression was help to methionol yield.

关 键 词：酿酒酵母 3-甲硫基丙醇氨基 转移酶基因 AR08基因 表达 

分 类 号：TS201.3[轻工技术与工程—食品科学]