X‐ray Structure of a Hg Complex of Mercuric Reductase (MerA) and Quantum Mechanical/Molecular Mechanical Study of Hg Transfer between the C‐Terminal and Buried Catalytic Site Cysteine Pairs

Mercuric reductase, MerA, is a key enzyme in bacterial mercury resistance. This homodimeric enzyme captures and reduces toxic Hg to Hg, which is relatively unreactive and can exit the cell passively. Prior to reduction, the Hg is transferred from a pair of cysteines (C558′ and C559′ using Tn501 numbering) at the C-terminus of one monomer to another pair of cysteines (C136 and C141) in the catalytic site of the other monomer. Here, we present the X-ray structure of the C-terminal Hg complex of the C136A/C141A double mutant of the Tn501 MerA catalytic core and explore the molecular mechanism of this Hg transfer with quantum mechanical/molecular mechanical (QM/MM) calculations. The transfer is found to be nearly thermoneutral and to pass through a stable tricoordinated intermediate that is marginally less stable than the two end states. For the overall process, Hg is always paired with at least two thiolates and thus is present at both the C-terminal and catalytic binding sites as a neutral complex. Prior to Hg transfer, C141 is negatively charged. As Hg is transferred into the catalytic site, a proton is transferred from C136 to C559′ while C558′ becomes negatively charged, resulting in the net transfer of a negative charge over a distance of ∼7.5 Å. Thus, the transport of this soft divalent cation is made energetically feasible by pairing a competition between multiple Cys thiols and/or thiolates for Hg with a competition between the Hg and protons for the thiolates. Metal ions play important functional roles in biological systems but can also be significant environmental pollutants. A detailed understanding of the mechanisms of speciation and transfer of heavy metals in biological and environmental systems is thus of both fundamental and practical interest. Among those chemical mechanisms involving heavy metals, of particular biological importance are ion transfers in proteins. Some microorganisms are able to overcome high concentrations of toxic heavy metals and can directly biotransform contaminants to innocuous or immobile forms. A key example is mercury resistance in bacteria conferred by the mer operon, which encodes a suite of proteins that carry out the transport and reduction of Hg to transform this toxic ion into less toxic, elemental Hg0. Mer loci have been discovered in many different species, underscoring the ubiquitous nature of this mode of mercury detoxification among bacterial communities. One of the key enzymes in the mer system is mercuric reductase, MerA, which catalyzes the reduction of Hg to Hg within the bacterial cytoplasm. The active form of MerA is a homodimer in which the two active sites within the catalytic Received: May 19, 2014 Revised: October 17, 2014 Published: October 24, 2014 Article pubs.acs.org/biochemistry © 2014 American Chemical Society 7211 dx.doi.org/10.1021/bi500608u | Biochemistry 2014, 53, 7211−7222 This is an open access article published under an ACS AuthorChoice License, which permits copying and redistribution of the article or any adaptations for non-commercial purposes.