文献类型：期刊文献

英文题名：Cloning and biochemical properties of a highly thermostable and enantioselective nitrilase from Alcaligenes sp. ECU0401 and its potential for (R)-(2)-mandelic acid production

作者：Zhang, Zhi-Jun[1]; Xu, Jian-He[1]; He, Yu-Cai[2]; Ouyang, Li-Ming[1]; Liu, You-Yan[3]

机构：[1] Laboratory of Biocatalysis and Bioprocessing, State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road, Shanghai 200237, China; [2] College of Chemistry and Chemical Engineering, Changzhou University, Changzhou 213164, China; [3] College of Chemistry and Chemical Engineering, Guangxi University, Nanning 530004, Guangxi, China

年份：2011

卷号：34

期号：3

起止页码：315

外文期刊名：Bioprocess and Biosystems Engineering

收录：EI(收录号：20112013988101)

语种：英文

外文关键词：Cloning - Escherichia coli - Enantioselectivity - Organic acids - Proteins - Purification

摘要：A nitrilase gene from Alcaligenes sp. ECU0401 was cloned and overexpressed in Escherichia coli BL21 (DE3) in a soluble form. The encoded protein with a His6-tag was purified to nearly homogeneity as revealed by SDS-PAGE with a molecular weight of approximately 38.5 kDa, and the holoenzyme was estimated to be composed of 10 subunits of identical size by size exclusion chromatography. The Vmax and Km parameters were determined to be 27.9 μmol min-1 mg-1 protein and 21.8 mM, respectively, with mandelonitrile as the substrate. The purified enzyme was highly thermostable with a half life of 155 h at 30°C and 94 h at 40°C. Racemic mandelonitrile (50 mM) could be enantioselectively hydrolyzed to (R)-(-)-mandelic acid by the purified nitrilase with an enantiomeric excess of 97%. The extreme stability, high activity and enantioselectivity of this nitrilase provide a solid base for its practical application in the production of (R)-(-)-mandelic acid. ? Springer-Verlag 2010.