FAD binding, cobinamide binding and active site communication in the corrin reductase (CobR).

Andrew D Lawrence,Christopher M Johnson,Stephen E J Rigby,Richard W Pickersgill,Martin J Warren,Michael A Geeves,Alan Scott,Mark J Howard,Samantha L Taylor,Michelle L Rowe

doi:10.1042/bsr20140060

Abstract

Adenosylcobalamin, the coenzyme form of vitamin B12, is one Nature's most complex coenzyme whose de novo biogenesis proceeds along either an anaerobic or aerobic metabolic pathway. The aerobic synthesis involves reduction of the centrally chelated cobalt metal ion of the corrin ring from Co(II) to Co(I) before adenosylation can take place. A corrin reductase (CobR) enzyme has been identified as the likely agent to catalyse this reduction of the metal ion. Herein, we reveal how Brucella melitensis CobR binds its coenzyme FAD (flavin dinucleotide) and we also show that the enzyme can bind a corrin substrate consistent with its role in reduction of the cobalt of the corrin ring. Stopped-flow kinetics and EPR reveal a mechanistic asymmetry in CobR dimer that provides a potential link between the two electron reduction by NADH to the single electron reduction of Co(II) to Co(I).

Highlights

Cobalamin, the biologically active form of vitamin B12, is an exquisitely complex natural product whose synthesis is limited to only some archaea and bacteria
Chemical shift assignments Backbone chemical shift assignments were completed for B. melitensis corrin reductase (CobR) with 94 %, 94 % and 86 % assignments, respectively, for backbone NH, Cα and Cβ resonances
The well-resolved 15N heteronuclear single-quantum coherence (HSQC) are consistent with a symmetrical CobR homodimer

Summary

Introduction

The biologically active form of vitamin B12, is an exquisitely complex natural product whose synthesis is limited to only some archaea and bacteria It functions as a coenzyme or cofactor in a number of important processes, and is associated with isomerization, methylation and dehalogenation reactions [1]. The lower axial ligand is a dimethylbenzimidazole (DMB) moiety that is attached to the corrin ring through an aminopropanol linker, with the upper axial position occupied by either a methyl or adenosyl group covalently linked to the cobalt. It is the properties of this unique cobalt–carbon bond that facilitate the exceptional chemistry associated with cobalamin-dependent enzymes [1]

Methods

Results

Conclusion