%0 Journal Article
%T Directed evolution of thermophilic esterase from the archaeon Aeropyrum pernix K1
嗜热酯酶APE1547催化活性的定向进化研究
%A WANG Qiu-yan
%A YANG Guang-yu
%A LIU Yan-li
%A WANG Yan-ping
%A FENG Yan
%A
王秋岩
%A 杨广宇
%A 刘艳莉
%A 王艳平
%A 冯雁
%J 微生物学报
%D 2006
%I
%X Thermophilic esterase(APE1547) from Aeropyrum pernix K1 was subjected to error-prone PCR(epPCR) to enhance activity.For the screening of mutants,an efficient and reliable assay suitable for high throughput screening was developed based on the enzyme thermostability.Two successive rounds of random mutagenesis by epPCR resulted in a four amino acid substitution variant M020 with significantly increased activity(six-fold under the screening condition.Further assay for the purified enzymes showed that the mutant possess 1.5-fold higher specific activity and nearly 4-fold higher expressed level than the wild-type.The mutant has an optimal activity at pH 8.5,corresponding to an alkaline shift of 0.5 pH unit compared to the wild type.The structure analysis suggests that R526S may contribute to the enhanced activity and the shift of pK-1.
%K Directed evolution
%K Thermophilic enzyme
%K Esterase
定向进化
%K 嗜热酶
%K 酯酶活力
%K 解离常数
%U http://www.alljournals.cn/get_abstract_url.aspx?pcid=90BA3D13E7F3BC869AC96FB3DA594E3FE34FBF7B8BC0E591&jid=A3C6BA55AB623B90FA9104CFFC826F3C&aid=AC387423AFF79AC3&yid=37904DC365DD7266&vid=D997634CFE9B6321&iid=0B39A22176CE99FB&sid=0F7768518993EDDE&eid=30897FA31CA3354D&journal_id=0001-6209&journal_name=微生物学报&referenced_num=1&reference_num=14