定点突变提高Thermococcus siculi HJ21高温酸性α-淀粉酶的催化活性  

Catalytic Activity Improvement of Acid-stable Alpha-Amylase from Hyperthermophilic Thermococcus siculi HJ21 by Site-directed Mutagenesis

作　　者：姚婷[1,2] 李华钟[1] 房耀维[2] 陆兆新[3] 王淑军[2] 焦豫良[2] 刘姝[2] 

机构地区：[1]江南大学工业生物技术教育部重点实验室,江苏无锡214122 [2]淮海工学院海洋学院,江苏连云港222005 [3]南京农业大学食品科技学院,江苏南京210095

出　　处：《食品科学》2011年第15期148-152,共5页Food Science

基　　金：国家自然科学基金项目(40746030);江苏省科技厅农业科技支撑计划项目(BE2008340);江苏省高校自然科学基础研究项目(08KJB550001)

摘　　要：通过分析超嗜热古菌Thermococcus siculi HJ21高温酸性α-淀粉酶基因,对6个可能提高酶催化活性的氨基酸残基进行定点突变,获得比未突变酶催化活性较高的突变子Y184H、Y121S、A109V、K98R、V125L。将以上5个突变位点集中在一个突变子上得到突变酶TSAM-23,该突变酶与未突变酶相比,酶的催化效率有较大提高,酶的活力和90℃条件下的热稳定性分别提高了3.75倍和2.4%,最适反应pH值为5.0,比未突变酶降低0.5,较之突变前有更好的工业应用价值。Based on the analysis of hyperthermophilic acid-stable α-amylase gene from Thermococcus siculi HJ21（TSA）,site-directed mutagenesis of six amino acid residues could improve the catalytic activity of α-amylase.Five mutants with higher catalytic activity were identified as K98R,A109V,Y121S,V125L and Y184H,respectively.Subsequent recombination of the mutants produced a mutant enzyme named as TSAM-23.Compared with TSA,the catalytic activity of TSAM-23 was enhanced significantly.The catalytic activity and thermal stability of TSAM-23 at 90 ℃were increased by 3.75 times and 2.4%,respectively.TSAM-23 had an optimal pH of 5.0,which revealed a decrease of 0.5 when compared with TSA.

关 键 词：Α-淀粉酶 催化活性 定点突变 

分 类 号：Q939.97[生物学—微生物学] TS201.3[轻工技术与工程—食品科学]