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Kinetic resolution of α‐methylbenzylamine with o‐transaminase screened from soil microorganisms: Application of a biphasic system to overcome product inhibition
Author(s) -
Shin JongShik,
Kim ByungGee
Publication year - 1997
Publication title -
biotechnology and bioengineering
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.136
H-Index - 189
eISSN - 1097-0290
pISSN - 0006-3592
DOI - 10.1002/(sici)1097-0290(19970720)55:2<348::aid-bit12>3.0.co;2-d
Subject(s) - acetophenone , transaminase , chemistry , enantiomeric excess , cyclohexanone , chromatography , nuclear chemistry , organic chemistry , enantioselective synthesis , catalysis , enzyme
Two microorganisms showing high o‐transaminase activity ( Klebsiella pneumoniae JS2F and Bacillus thuringiensis JS64) were screened by the enrichment method using ( S )‐α‐methylbenzylamine (α‐MBA) as a sole nitrogen source. Optimal carbon and nitrogen sources for enzyme induction and the properties of o‐transaminases were investigated. o‐Transaminase from B. thuringiensis JS64 was highly enantioselective ( E = 75.3) for ( S )‐enantiomer of α‐MBA and showed remarkable stability. However, o‐transaminase showed severe product inhibition by acetophenone. An aqueous/organic two‐phase system was introduced to overcome this problem. Through solvent screening, cyclohexanone and ethyl acetate were selected as the best organic phases. The acetophenone‐extracting capacity of the solvent and the biocompatibility of the solvent to the cell were important determinants in the reaction rate at high concentrations of α‐MBA. The reaction rate of o‐transamination was strongly influenced by the volume ratio of organic phase to aqueous phase as well as agitation speed in the biphasic mixture. Using the optimal volume ratio ( V org: V aq = 1:4) in the biphasic system with cyclohexanone, the reaction rate of o‐transaminase under vigorous mixing conditions increased ninefold compared with that in the monophasic aqueous system. At the same optimal conditions, using whole cells, 500 m M α‐MBA could be resolved successfully to above 95% enantiomeric excess of ( R )‐α‐MBA with ca. 51% conversion. © 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 55 : 348–358, 1997.