摘要
为了研究亚油酸异构酶(LAI)的功能并构建基因工程菌株,克隆了植物乳杆菌lp15-2-1的lai基因并对其编码序列进行分析比对,随后将其融合酵母信号肽序列和α-凝集素锚定序列,构建酵母表面展示载体;使用电击法转化酿酒酵母K601,并在尿嘧啶缺失型培养基上筛选出转化子.氨基酸序列比对分析表明,LAI与肌球蛋白交叉反应抗原蛋白家族同源性极高,N端含有一个半保守的黄素腺嘌呤二核苷酸结合基序.对酵母工程菌株的研究表明,LAI成功地在酵母细胞表面展示,酶活在诱导培养48 h时达到最大,为40.5 U/mL.气相色谱检测发现,酵母工程菌株能够合成单一的c9t,11-共轭亚油酸异构体.
In order to investigate the function of linoleic acid isomerase （LAI） and construct the recombinant yeast strains, first, the lai gene was cloned from Lactobacillus plantarum lp15-2-1, with its coding sequence being also analyzed. Then, lai was fused to the sequences of both the signal peptide of yeast and the anchor region of α-agglutinin, thus constructing the yeast cell-surface display vector. Finally, the vector was transformed into Saccharomyces cerevisiae K601 by means of the electroporation, and the transformants were screened by using the synthetic dropout medium without uracil. Amino acid sequence analysis shows that LAI is of a high homology to the myosin-cross-reactive antigen proteins as well as of a semi-conserved flavin adenine dinucleotide binding motif near the N-terminus. The activity results of the recombinant yeast strains demonstrate that LAI from Lactobacillus plantarum is successfully displayed on the cell-surface of yeast K601, and that the activity of LAI reaches the maximum 40. 5 U/mL in 48 h. Moreover, GC results indicate that the recombinant yeast strains are capable of producing a single c9, tl 1- CLA isomer.
出处
《华南理工大学学报（自然科学版）》
EI
CAS
CSCD
北大核心
2011年第9期140-146,共7页
Journal of South China University of Technology(Natural Science Edition)
基金
国家"863"计划重点项目(2007AA100402)
关键词
植物乳杆菌
酿酒酵母
亚油酸异构酶
表面展示
黄素腺嘌呤二核苷酸
Lactobacillus plantarum
Saccharomyces cerevisiae
linoleic acid isomerase
surface display
flavin adenine dinucleotide
