Substrate spectrum of ω-transaminase from Chromobacterium violaceum DSM30191 and its potential for biocatalysis

A new omega-transaminase (omega-TA) from Chromobacterium violaceum DSM30191 has been recruited from the pool of fully sequenced genomes, using the V. fluvialis JS17 omega-TA sequence [Shin JS, Kim BG. Biosci Biotechnol Biochem 2001;65:1782-8; Shin JS, Yun H, Jang JW, Park I, Kim BG. Appl Microbiol Biotechnol 2003;61:463-71] in a BLASTP search. The protein sequence of the C. violaceum enzyme exhibits 38% sequence identity to the V fluvialis enzyme. The gene was functionally expressed in Escherichia coli BL21 by using the pET-expression system, making up 63% of the protein in the clarified cell extract (1.2U/mg). As with the V. fluvialis TA, the C violaceum enzyme shows (S)-enantioselective TA activity and converts (S)-alpha-methylbenzylamine and pyruvate almost completely to acetophenone and L-Ala. Glyoxylate proved to be the best acceptor and aliphatic and aromatic aldehydes were both readily converted. Importantly we show here for the first time that an w-TA can be used for the amination of aromatic 2-keto-1,3-diols. An LC-ESI-MS method was developed to quantitatively detect aromatic aminodiol formation. Using clarified cell extract, we were able to convert 20 mM of racemic 1,3-dihydroxy-1-phenylpropan-2-one to 2-amino-1-phenylpropane-1,3-diol with 60% conversion in 24 h. (C) 2007 Elsevier Inc. All rights reserved.