Role of conserved histidine residues in metalloactivation of the ArsA ATPase

The ArsA ATPase is the catalytic subunit of a pump that is responsible for resistance to arsenicals and antimonials in Escherichia coli. Arsenite or a ntimonite allosterically activates the ArsA ATPase activity. ArsA homologue s from eubacteria, archaea and eukarya have a signature sequence (DTAPTGHT) that includes a conserved histidine. The ArsA ATPase has two such conserve d motifs, one in the NH2-terminal (A1) half and the other in the COOH-termi nal (A2) half of the protein. These sequences have been proposed to be sign al transduction domains that transmit the information of metal occupancy at the allosteric to the catalytic site to activate ATP hydrolysis. The role of the conserved residues His148 and His453, which reside in the A1 and A2 signal transduction domains respectively, was investigated by mutagenesis t o create H148A, H453A or H148A/H453A ArsAs. Each altered protein exhibited a decrease in the V-max of metalloid-activated ATP hydrolysis, in the order wild type ArsA > H148A > H453A > H148A/H453A. These results suggest that t he histidine residues play a role in transmission of the signal between the catalytic and allosteric sites.