title: Nitrilase-catalyzed conversion of (R,S)-mandelonitrile by immobilized recombinant Escherichia coli cells harboring nitrilase.
abstract: (R)-(-)-Mandelic acid (R-MA) is widely used both as a versatile intermediate for pharmaceuticals and a resolving agent in chiral resolution processes. In the current study, to improve the stability of operation, recombinant Escherichia coli cells expressing nitrilase from Alcaligenes faecalis were immobilized for the enantioselective hydrolysis of (R,S)-mandelonitrile to R-MA. Different immobilization methods including entrapment matrices, entrapment matrices cross-linked by cross-linking and polymerization agents, and direct cross-linking cells using glutaraldehyde (GA) or bionic silicon were investigated. To facilitate industrial solid-liquid separation, the direct cross-linking recombinant E. coli cells using diatomite/GA/polyethyleneimine with 135.95% relative activity compared with free cells was chosen using water as the reaction medium. The operational stability of the immobilized cells was obviously superior to that of free cells, without significant activity loss after 28 cycles of batch reaction and the successive production of R-MA could reach 1.88 M. Moreover, the immobilized cells showed good storage stability with about 52% relative activity after storing for 30 days at 4 °C. Therefore, the immobilized biocatalyst is very promising for upscale production of optically pure R-MA with high performance and low cost.
authors.affiliation: Institute of Bioengineering, Zhejiang University of Technology, Hangzhou, Zhejiang, People's Republic of China.
authors.forename: Xin-Hong
authors.initials: XH
authors.lastname: Zhang
authors.affiliation: Institute of Bioengineering, Zhejiang University of Technology, Hangzhou, Zhejiang, People's Republic of China.
authors.forename: Zhi-Qiang
authors.initials: ZQ
authors.lastname: Liu
authors.affiliation: Institute of Bioengineering, Zhejiang University of Technology, Hangzhou, Zhejiang, People's Republic of China.
authors.forename: Ya-Ping
authors.initials: YP
authors.lastname: Xue
authors.affiliation: Zhejiang Laiyi Biotechnology Co, Shengzhou, Zhejiang, People's Republic of China.
authors.forename: Ming
authors.initials: M
authors.lastname: Xu
authors.affiliation: Institute of Bioengineering, Zhejiang University of Technology, Hangzhou, Zhejiang, People's Republic of China.
authors.forename: Yu-Guo
authors.initials: YG
authors.lastname: Zheng
mesh_terms: D000097:Acetonitriles / Q000378:metabolism*; D000619:Aminohydrolases / Q000096:biosynthesis / Q000378:metabolism*; D061249:Batch Cell Culture Techniques; D055162:Biocatalysis; D001711:Biotransformation; D018914:Cells, Immobilized / Q000378:metabolism; D004926:Escherichia coli / Q000166:cytology* / Q000235:genetics / Q000378:metabolism*; D005285:Fermentation; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism*; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D000097:Acetonitriles; D011994:Recombinant Proteins; C045516:mandelonitrile; D000619:Aminohydrolases; C021867:nitrilase
keywords: (R)-(−)-mandelic acid; cross-linking; enantioselective hydrolysis; entrapment matrices; immobilized cells; nitrilase