An Aminocaprolactam Racemase from Ochrobactrum anthropi with Promiscuous Amino Acid Ester Racemase Activity

Amina Frese; Sarah V Barrass; Peter W Sutton; Joe P Adams; Gideon Grogan

doi:10.1002/cbic.201800265

An Aminocaprolactam Racemase from Ochrobactrum anthropi with Promiscuous Amino Acid Ester Racemase Activity

Chembiochem. 2018 Jun 13. doi: 10.1002/cbic.201800265. Online ahead of print.

Authors

Amina Frese¹, Sarah V Barrass¹, Peter W Sutton^{2

3}, Joe P Adams², Gideon Grogan¹

Affiliations

¹ York Structural Biology Laboratory, Department of Chemistry, University of York, Heslington, York, YO10 5DD, UK.
² GSK Medicines Research Centre, Gunnels Wood Road, Stevenage, Hertfordshire, SG1 2NY, UK.
³ Present address: Department of Chemical, Biological and Environmental Engineering, Bioprocess Engineering and Applied Biocatalysis Group, Engineering School, Campus de la UAB, 08193, Bellaterra (Cerdanyola del Vallés), Barcelona, Spain.

PMID: 29897155
DOI: 10.1002/cbic.201800265

Abstract

The kinetic resolution of amino acid esters (AAEs) is a useful synthetic strategy for the preparation of single-enantiomer amino acids. The development of an enzymatic dynamic kinetic resolution (DKR) process for AAEs, which would give a theoretical yield of 100 % of the enantiopure product, would require an amino acid ester racemase (AAER); however, no such enzyme has been described. We have identified low AAER activity of 15 U mg^-1 in a homologue of a PLP-dependent α-amino ϵ-caprolactam racemase (ACLR) from Ochrobactrum anthropi. We have determined the structure of this enzyme, OaACLR, to a resolution of 1.87 Å and, by using structure-guided saturation mutagenesis, in combination with a colorimetric screen for AAER activity, we have identified a mutant, L293C, in which the promiscuous AAER activity of this enzyme towards l-phenylalanine methyl ester is improved 3.7-fold.

Keywords: amino acid amides; amino acid esters; amino acids; biocatalysis; pyridoxal phosphate; racemases.