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Identification of protein interaction networks, cofactors and protein structures essential for mercury methylation in anaerobic bacteria


EMSL Project ID
49823

Abstract

Mercury is a pervasive global pollutant. 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. Microbial mercury methylation is an enzyme-catalyzed process carried out by certain anaerobic bacteria and archaea. A two-gene cluster hgcAB is essential for mercury methylation and encodes a cobalamin-dependent protein, HgcA, and a ferredoxin, HgcB, which are predicted to facilitate methyl transfer and cofactor reduction, respectively. Determining the specific roles and interactions of HgcA and HgcB with other cellular components offers insights into the biochemical pathways associated with bacterial mercury methylation common to a broad range of anaerobic bacteria and archaea. The objective of the proposed work is to elucidate fundamental biomolecular mechanisms and interactions of HgcA and HgcB by identifying subcellular components and structures that are critical for Hg methylation in anaerobic bacteria. The proposed work takes advantage of unique EMSL mass spectrometry and spectroscopy capabilities. Covalent crosslinking will be used to reveal protein-protein interactions, which will in combination with structural bioinformatics help delineate the roles of HgcA and HgcB in the context of microbial carbon metabolism. Key cellular components are isolated by affinity purification techniques and identified by high resolution mass spectrometry at EMSL. In addition, nuclear magnetic resonance spectra will be collected to determine molecular structures and protein-ligand interactions. The experimental data obtained in this study will be used to identify how HgcA and HgcB interact with cellular components and pathways required for catalytic turnover leading to mercury methylation in anaerobic bacteria. The results will help reveal key processes that control the fate and transformation of mercury in aquatic ecosystems and will provide essential data to inform metabolic and reactive transport models that can be used to predict mercury cycling from the molecular to the field scale. A comprehensive understanding of the various geochemical and biochemical factors culminating in the production of MeHg will facilitate the development of effective strategies to limit production and bioaccumulation of methylmercury in the environment.

Project Details

Project type
Large-Scale EMSL Research
Start Date
2017-10-01
End Date
2018-09-30
Status
Closed

Team

Principal Investigator

Alexander Johs
Institution
Oak Ridge National Laboratory

Team Members

Swapneeta Date
Institution
Oak Ridge National Laboratory

Stephen Ragsdale
Institution
University of Michigan

Katherine W. Rush
Institution
University of Michigan