Complementation between nucleotide binding domains in an anion-translocating ATPase.
The catalytic component of the oxyanion-translocating ATPase of the plasmid-encoded ars operon of Escherichia coli is a homodimer of the ArsA protein. This enzyme is an oxyanion-stimulated ATPase with two consensus nucleotide binding sequences in each subunit, one in the N-terminal (A1) half and one in the C-terminal (A2) half of the ArsA protein. The two halves of both the arsA gene and the ArsA protein exhibit similar nucleotide and amino acid sequences, respectively. The two halves of the arsA gene were subcloned into compatible plasmids. Neither alone was sufficient to confer resistance, but cells in which the arsA1 and arsA2 half genes were coexpressed were resistant to arsenicals. Genetic complementation was also observed in cells bearing plasmids with point mutations in the two halves of the arsA gene and between cells with plasmids carrying combinations of the arsA1 or arsA2 subclones and point mutations. In every case, complementation was observed only when one plasmid contained a wild-type arsA1 sequence and the other contained a wild-type arsA2 sequence. These results demonstrate that both sites are required for resistance but that the two nucleotide binding domains need not reside in a single polypeptide. We propose a model in which the ArsA dimer has two catalytic units, each composed of an A1 domain from one monomer and an A2 domain from the other monomer.
Kaur
P
P
Department of Biochemistry, Wayne State University School of Medicine, Detroit, Michigan 48201.
Rosen
B P
BP
D000251:Adenosine Triphosphatases / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D000595:Amino Acid Sequence; D001151:Arsenic / Q000494:pharmacology; D053501:Arsenite Transporting ATPases; D018053:Arsenites; D001665:Binding Sites; D004352:Drug Resistance, Microbial / Q000235:genetics; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics* / Q000254:growth & development; D005798:Genes, Bacterial; D005816:Genetic Complementation Test; D016623:Ion Pumps; D007700:Kinetics; D046911:Macromolecular Substances; D008961:Models, Structural; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes; D009876:Operon; D017670:Sodium Compounds
D018053:Arsenites; D016623:Ion Pumps; D046911:Macromolecular Substances; D009097:Multienzyme Complexes; D017670:Sodium Compounds; C017947:sodium arsenite; D000251:Adenosine Triphosphatases; D053501:Arsenite Transporting ATPases; D001151:Arsenic