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Abstract: Aminotransferases (ATs) are crucial enzymes for prokaryote and eukaryote metabolism, and have attracted industrial attention for large scale synthesis of amino acids and chiral amines. A high throughput screening procedure for branched chain aminotransferases (BCATs) was developed to identify optimal expression on a large number of expression clones. Escherichia coli BCAT, encoded by the ilvE gene, was expressed in E. coli and Pichia pastoris. This simple colorimetric assay procedure allowed the identification of optimal clones for ilvE expression and enabled the testing of its activity in cell lysates or on whole cell catalysis. [Copyright &y& Elsevier]

Abstract: Rhodococcus equi A4, Rhodococcus erythropolis NCIMB 11540 and Rhodococcus sp. R312 were investigated towards their ability to produce β-amino amides and acids from β-amino nitriles. The microorganisms show comparable trends: five-membered alicyclic 2-amino nitriles were transformed significantly faster than the six-membered compounds and the products of trans-2-amino nitriles (amides and acids) were formed considerably faster than the cis-counterparts (amides). The trans-five membered nitriles gave the amides (1b, 5b) in excellent enantiomeric excess (94–99%), the biotransformation of trans-six membered substrates resulted in the formation of the acid (3c, 7c) in excellent ee (87–99%). The ee''s of the cis-compounds were throughout lower. Fifteen new substances were synthesized and characterized in the course of this work. [Copyright &y& Elsevier]

Microbial cytochrome P450 enzymes (CYPs) are able to mimic the metabolism of human CYPs. One challenge is to identify the respective drug metabolites and to compare substrate specificities to those of the human enzymes. In this study, a class VIII self-sufficient CYP from Aspergillus fumigatus (CYP505X) and variants of this enzyme were heterologously expressed in E. coli. The substrate scope of the variants was determined using active pharmaceutical ingredients (APIs) and (hetero)cyclic compounds. Capsaicin – the active compound in chili peppers – was oxidized most efficiently (4.36 μM/min) in a whole cell mediated biotransformation. The products were isolated, purified and their structures elucidated by 1D and 2D NMR. The two major metabolites showed modifications on the lipophilic side chain. Specifically, capsaicin was hydroxylated at position 8 to give (E) -8-hydroxy- N -(4-hydroxy-3-methoxybenzyl)-8-methylnon-6-enamide and epoxidized at the double bond to give N -(4-hydroxy-3-methoxybenzyl)-5-(3-isopropyloxiran-2-yl)-pentanamide. Graphical abstract Image 1 [ABSTRACT FROM AUTHOR]