A three-component monooxygenase from Rhodococcus wratislaviensis may expand industrial applications of bacterial enzymes

Makoto Hibi,Dai Fukuda,Satomi Takahashi,Mitsuyoshi Ueda,Bunzo Mikami,Wataru Aoki,Shunsuke Aburaya,Kimihiko Mizutani,Jun Ogawa,Chihiro Kenchu,Michiki Takeuchi,Masutoshi Nojiri,Yoshihiko Yasohara,Ryotaro Hara

doi:10.1038/s42003-020-01555-3

Abstract

The high-valent iron-oxo species formed in the non-heme diiron enzymes have high oxidative reactivity and catalyze difficult chemical reactions. Although the hydroxylation of inert methyl groups is an industrially promising reaction, utilizing non-heme diiron enzymes as such a biocatalyst has been difficult. Here we show a three-component monooxygenase system for the selective terminal hydroxylation of α-aminoisobutyric acid (Aib) into α-methyl-D-serine. It consists of the hydroxylase component, AibH1H2, and the electron transfer component. Aib hydroxylation is the initial step of Aib catabolism in Rhodococcus wratislaviensis C31-06, which has been fully elucidated through a proteome analysis. The crystal structure analysis revealed that AibH1H2 forms a heterotetramer of two amidohydrolase superfamily proteins, of which AibHm2 is a non-heme diiron protein and functions as a catalytic subunit. The Aib monooxygenase was demonstrated to be a promising biocatalyst that is suitable for bioprocesses in which the inert C–H bond in methyl groups need to be activated.

Highlights

The high-valent iron-oxo species formed in the non-heme diiron enzymes have high oxidative reactivity and catalyze difficult chemical reactions
Similar to previous reports[17,18], four of these isolates produced acetone through Aib degradation. These acetone-forming Aib-degrading strains possess PLP-dependent α,α-dialkylglycine decarboxylase activity because the Aib degradation and acetone production were completely inhibited by aminooxyacetic acid (AOA), a well-known PLP-dependent enzyme inhibitor[19]
Our results indicate that these microorganisms possess an undiscovered Aib catabolic pathway that is independent of the α,α-dialkylglycine decarboxylase reaction

Summary

Introduction

The high-valent iron-oxo species formed in the non-heme diiron enzymes have high oxidative reactivity and catalyze difficult chemical reactions. The hydroxylation of inert methyl groups is an industrially promising reaction, utilizing non-heme diiron enzymes as such a biocatalyst has been difficult. We show a three-component monooxygenase system for the selective terminal hydroxylation of α-aminoisobutyric acid (Aib) into α-methyl-D-serine. It consists of the hydroxylase component, AibH1H2, and the electron transfer component. The non-heme diiron enzymes activate dioxygen at ambient temperature and pressure to catalyze many reactions involved in important biological processes. These processes include: methane hydroxylation in methanotrophs; the degradation of aromatic compounds; the biosynthesis of deoxyribonucleotides, fatty acids, and antibiotics; and the regulation of cell proliferation[11]. The direct and selective functionalization of inert C–H bonds is useful for fuel production and industrial biosynthetic applications by improving stepeconomy and atom-economy[12,13]

Methods

Results

Conclusion