NMR structure determination of the cobalamin binding protein HgcA and the ferredoxin-like protein HgcB required for bacterial mercury methylation
EMSL Project ID
48393
Abstract
Mercury is a pervasive global pollutant. Its methylated form, methylmercury, poses a substantial threat to human health. Methylation of mercury (Hg) by anaerobic bacteria is the primary source of toxic methylmercury, which bioaccumulates up trophic levels and affects humans primarily through consumption of fish and other seafood. The recent discovery of two genes, hgcA and hgcB, required for Hg methylation, enables structural characterization of the protein to elucidate the methylation mechanism, common to a broad range of anaerobic bacteria and archaea. Homology modeling of the cytosolic HgcA cobalamin-binding domain suggests a Rossman-like fold with a direct coordination of a cysteine thiolate to the Co center of its cobalamin cofactor in the "base-off" configuration. This unique thiolate-cobalt coordination in HgcA, which has never been observed previously in a biological context, is predicted to facilitate methyl transfer to a Hg(II) substrate during the formation of MeHg. The associated 2[4Fe-4S] ferredoxin HgcB presumably reduces the cobalamin cofactor of HgcA to the Co(I) state. Nuclear magnetic resonance (NMR) spectroscopy can be applied to determine the molecular structure of HgcA and HgcB from the model organism Desulfovibrio desulfuricans ND132. The results will enable experimental verification of the proposed thiolate-cobalt coordination in HgcA and provide experimental data to verify our hypothesis that this configuration promotes the transfer of a methyl carbanion to a Hg(II) substrate. The structure of HgcB will help elucidate the functional role of the two iron-sulfur clusters and EMSL provides NMR capabilities for obtaining 2D and 3D NMR spectra from the HgcA cobalamin binding domain and its electron transfer partner HgcB and essential support to perform structure calculations and refinement. The structural data obtained from this study can also be used to identify how HgcA and HgcB interact with each other and help with identifying other functional domains required for catalytic turnover. The results are expected to significantly improve our understanding of the underlying biomolecular mechanism leading to the formation of toxic methylmercury in the environment.
Project Details
Project type
Large-Scale EMSL Research
Start Date
2014-10-01
End Date
2016-09-30
Status
Closed
Released Data Link
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