BioWF: A naturally‐fused, di‐domain biocatalyst from biotin biosynthesis displays an unexpectedly broad substrate scope

Shona M Richardson, Peter J Harrison, Michael A Herrera, Menglu Wang, Rebecca Verez, Gustavo Perez Ortiz, Dominic James Campopiano

Research output: Contribution to journalArticlepeer-review

Abstract

The carbon backbone of biotin is constructed from the C 7 di-acid pimelate, which is converted to an acyl-CoA thioester by an ATP-dependent, pimeloyl-CoA synthetase (PCAS, encoded by BioW). The acyl-thioester is condensed with ʟ-alanine in a decarboxylative, Claisen-like reaction to form an aminoketone (8-amino-7-oxononanoic acid, AON). This step is catalysed by the pyridoxal 5’-phosphate (PLP)-dependent enzyme (AON synthase, AONS, encoded by BioF). Distinct versions of Bacillus subtilis BioW ( Bs BioW) and E. coli BioF ( Ec BioF) display strict substrate specificity. In contrast, a BioW-BioF fusion from Corynebacterium amycolatum ( Ca BioWF) accepts a wider range of mono- and di-fatty acids. Analysis of the active site of the Bs BioW:pimeloyl-adenylate complex suggested a key role for a Phe (F192) residue in the Ca BioW domain; a F192Y mutant restored the substrate specificity to pimelate. This surprising substrate flexibility also extends to the Ca BioF domain, which accepts ʟ-alanine, ʟ-serine and glycine. Structural models of the Ca BioWF fusion provide insight into how both domains interact with each other and suggest the presence of an intra-domain tunnel. The Ca BioWF fusion catalyses conversion of various fatty acids and amino acids to a range of AON derivatives. Such unexpected, natural broad substrate scope suggests that the Ca BioWF fusion is a versatile biocatalyst that can be used to prepare a number of aminoketone analogues.
Original languageEnglish
JournalChemBioChem
Early online date13 Jun 2022
DOIs
Publication statusE-pub ahead of print - 13 Jun 2022

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