Mercury resistance in the archaeon Sulfolobus solfataricus

Biological resistance to mercury has been widely studied in bacteria. Although archaeal genome sequencing indicates the presence of orthologous mercury resistance genes, there are virtually no studies regarding mercury resistance in archaea. This study focuses on the mechanism of mercury resistance in archaeon Sulfolobus solfataricus. Specifically, this study was conducted to (a) provide in vivo confirmation of the mercury resistance genes in the S. solfataricus mer operon, (b) investigate the regulatory role of MerR in mercury resistant transcription of the mer genes, (c) characterize the mer operon promoters, (d) and characterize the merA gene product mercuric reductase (MerA) and the merH gene product, a putative metallochaperone (MerH). Gene disruption identified merR as the gene which encodes the repressor of the mer operon MerR and merA as the gene which encodes mercuric reductase (MerA). Further genetic manipulations along with northern blot analysis and electromobility shift assays suggest that MerR prevents mercury from inhibiting transcription of the mer operon by binding to an inverted repeat just upstream of the merH promoter. Physiological and RTPCR analysis of the merRp-TATA, merHp-TATA and merAp-TATA mutants provide, for the first time, in vivo confirmation of the TATA box in S. solfataricus. Using markerless exchange, a hexahistidine tag was fused to the N-terminal end of the MerA protein in vivo. This allowed for subsequent Ni2+-nitrilotriacetic acid purification of native MerA and preliminary MerA assays. The results presented in this study contribute to improved understanding of archaeal mercury resistance. The methods developed throughout the course of this work expand the growing set of tools available for archaeal research efforts