Studies on mercury transporter merT and mercury reductase merA proteins

Abstract

Heavy metal pollution is one of the major pollutants in both developed and developing countries. Heavy metals like Lead, Cadmium, Chromium and Mercury are released into the environment from the mining industries and as industrial effluents. The structures of the mercury regulator (merR) and mercury transporter (merT) are yet to be solved whereas the structures of the other proteins are already solved and available in protein data bank. The first objective of this work is to express and purify the highly toxic, hydrophobic protein mercury transporter (merT) protein in E. coli. Three different groups around the world are currently attempting to solve the structure of merT protein by X-ray, NMR and Electron microscopy. The limitations in these methods are the requirement for high concentration of protein of interest for experimental trials. MerT is a high toxic, hydrophobic protein and expression of them in common E. coli hosts like BL21(DE3), BL21(DE3)pLysS and GJ1158 were not successful. The second objective of this work is to clone and express mercury reductase (merA) enzyme in E. coli for bioremediation of mercury from the environment. Mercury reductase is a oxidoreductase enzyme which vaporizes metallic mercury. The structure of mercury reductase is solved to high resolution and the structure is available in protein data bank. The third objective of this work is to study the protein protein interaction of mercury transporter(merT) and mercury reductase (merA). The mechanism of transport of mercury into the bacterial cell is still debated and area of intense research. Mercury transporter (merT) and mercury reductase (merA) were cloned and expressed as fusion tags with green fluorescent protein and red fluorescent protein and their interactions we observed using 488nm excitation and 590nm emission for red fluorescent protein. We have observed the interaction of the proteins successfully using FRET. We have developed a method to study transient protein-protein interactions between membrane protein and its