Abstract
This study describes the identification and the structural and spectroscopic analysis of a cobalamin-binding protein (termed CobDH) implicated in O-demethylation by the organohalide-respiring bacterium Desulfitobacterium hafniense DCB-2. The 1.5 Å resolution crystal structure of CobDH is presented in the cobalamin-bound state and reveals that the protein is composed of an N-terminal helix-bundle domain and a C-terminal Rossmann-fold domain, with the cobalamin coordinated in the base-off/His-on conformation similar to other cobalamin-binding domains that catalyse methyl-transfer reactions. EPR spectroscopy of CobDH confirms cobalamin binding and reveals the presence of a cob(III)alamin superoxide, indicating binding of oxygen to the fully oxidized cofactor. These data provide the first structural insights into the methyltransferase reactions that occur during O-demethylation by D. hafniense.
Highlights
Lignin breakdown by fungal haloperoxidases in forest soil can give rise to chlorinated phenyl methyl ethers
CobDH was reconstituted during the purification process with exogenous methylcobalamin, as the heterologous E. coli host only synthesizes methylcobalamin when supplied with cobinamide (Lawrence & Roth, 1995)
The structure reveals the characteristic base-off/His-on cobalamin-binding site with the cobalamin coordinated to His102 as part of a conserved sequence motif observed in other cobalamin-dependent methyltransferases and implies that CobDH likewise catalyses the transfer of a methyl group
Summary
Lignin breakdown by fungal haloperoxidases in forest soil can give rise to chlorinated phenyl methyl ethers Such compounds can serve as the terminal electron acceptors for anaerobic organohalide-respiring bacteria, leading to the corresponding dechlorinated phenyl methyl ethers (Bache & Pfennig, 1981; Villemur, 2013). The recent availability of genome sequences from several organohalide-respiring bacteria has revealed, in addition to a surprising wealth of reductive dehalogenase genes (RdhAs), the presence of multiple putative cobalamin-binding proteins (Kim et al, 2012). These are often located in gene clusters that appear to encode multicomponent O-demethylase-like enzyme systems (Studenik et al, 2012)
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
