文献类型：期刊文献

英文题名：Expression and Characterization of a Novel Lipase from Aspergillus fumigatus with High Specific Activity

作者：Shangguan, Jiao-Jiao[1];Liu, Yu-Qiang[1];Wang, Fu-Jun[2];Zhao, Jian[1];Fan, Li-Qiang[1];Li, Su-Xia[1];Xu, Jian-He[1]

机构：[1]E China Univ Sci & Technol, State Key Lab Bioreactor Engn, Shanghai 200237, Peoples R China;[2]Shanghai Univ Chinese Tradit Med, Inst Chinese Mat Med, Shanghai 201203, Peoples R China

年份：2011

卷号：165

期号：3-4

起止页码：949

外文期刊名：APPLIED BIOCHEMISTRY AND BIOTECHNOLOGY

收录：;EI(收录号：20114714540167);WOS:【SCI-EXPANDED(收录号:WOS:000296640000018)】；

基金：This research was financially supported by China National Special Fund for State Key Laboratory of Bioreactor Engineering (No. 2060204) and Shanghai Leading Academic Discipline Project (No. B505).

语种：英文

外文关键词：Aspergillus fumigatus; Recombinant lipase; Expression; Characterization; Active site

摘要：A novel lipase gene from Aspergillus fumigatus, afl1-1, was cloned and expressed with a molecular mass of 38 kDa in Escherichia coli for the first time. The recombinant lipase had a preference for short carbon chain p-nitrophenyl esters, especially toward C2 p-nitrophenyl ester and exhibited potent hydrolysis activity that had not been observed. The optimum pH and temperature of this new enzyme were 8.5 and 65 A degrees C, respectively. The recombinant lipase (AFL1-1) is an alkaline enzyme which was stable in the pH range 6.0 similar to 8.5 for 16 h (at 4 A degrees C) and at 30 similar to 50 A degrees C for 1 h. It is an intracellular enzyme which was purified approximately 8.47-fold with an overall yield of 86.1% by single-step Ni-NTA affinity purification, with a very high specific activity of approximately 1.00 x 10(3) U mg(-1) on a standard substrate of p-nitrophenyl acetate. The Michaelis-Menten kinetic parameters V (max) and K (m) of the lipase were 1.37 mM mg(-1) min(-1) and 14.0 mM, respectively. Ca2+ and other metal ions could not activate the lipase. According to the homology analysis and site-directed mutagenesis assay, the catalytic triad of the recombinant lipase was identified as Ser-165, Asp-260, and His-290 residues.