Conserved Glu-181 and Arg-182 residues of Escherichia coli H(+)-ATPase (ATP synthase) beta subunit are essential for catalysis: properties of 33 mutants between beta Glu-161 and beta Lys-201 residues.
Twenty-two mutants between beta Glu-161 and beta Lys-201 of Escherichia coli H(+)-ATPase beta subunit could grow by oxidative phosphorylation, but 11 other such mutants, beta Glu-181-->Gln, Asp, Asn, Thr, Ser, Ala, or Lys and beta Arg-182-->Lys, Ala, Glu, or Gln, could not. The beta Asp-181, beta Lys-182, and other defective mutants had 1.4, 1, and < 0.1%, respectively, of the wild-type membrane ATPase activity. Partially purified F1-ATPases from all mutants at positions 181 and 182, except for the beta Asp-181 and beta Lys-182 mutants, showed very low unisite catalysis. Purified F1-ATPases of the beta Gln-181 and beta Ala-181 mutants showed no multisite (or steady state) catalysis and slow unisite catalysis (< or = 1% of that of the wild type): their defects could be attributed to decreased catalytic rates (low k+2 and k-2). Changes of the k+2 and k-2 values in the beta Asp-181 enzyme, which showed detectable multi- and unisite catalysis, were less marked (27 and 21%, respectively, of wild-type rates). The beta Gln-182 enzyme showed defective catalysis (< or = 0.1% of the multi- and approximately 1% of the unisite catalyses of the wild type), whereas the beta Lys-182 enzyme showed 1 and 85% of the wild-type multisite and unisite catalytic rates, respectively. beta Lys-182 had wild-type values of k+2 and k-2, but beta Gln-182 had k+2 about 10-fold lower than that of wild type.(ABSTRACT TRUNCATED AT 250 WORDS)
Park
M Y
MY
Department of Organic Chemistry and Biochemistry, Osaka University, Ibaraki.
Omote
H
H
Maeda
M
M
Futai
M
M
D000227:Adenine Nucleotides; D001120:Arginine / Q000737:chemistry*; D001483:Base Sequence; D001665:Binding Sites; D018698:Glutamic Acid / Q000737:chemistry*; D007700:Kinetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D006180:Proton-Translocating ATPases / Q000737:chemistry*
D000227:Adenine Nucleotides; D018698:Glutamic Acid; D001120:Arginine; D006180:Proton-Translocating ATPases