Nuclear magnetic resonance spectroscopy structure determination of the cobalamin binding protein HgcA required for bacterial mercury methylation
EMSL Project ID
48184
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 and functional characterization of 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. Nuclear magnetic resonance (NMR) spectroscopy can be applied to determine the molecular structure of the N-terminal cobalamin-binding domain of the protein HgcA from Desulfovibrio desulfuricans ND132. The results will enable experimental verification of the proposed thiolate-cobalt coordination and provide experimental data to verify our hypothesis that this configuration promotes the transfer of a methyl carbanion to a Hg(II) substrate. EMSL provides NMR capabilities that are well suited for obtaining 2D and 3D spectra from the HgcA cobalamin binding domain as well as the necessary support to perform structure calculations and refinement. The structural data obtained from this study can also be used to identify how HgcA interacts with 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
Exploratory Research
Start Date
2013-12-02
End Date
2014-09-30
Status
Closed
Released Data Link
Team
Principal Investigator
Team Members