Escherichia coli H(+)-ATPase: role of the delta subunit in binding Fl to the Fo sector.
The roles of the Escherichia coli H(+)-ATPase (FoFl) delta subunit (177 amino acid residues) was studied by analyzing mutants. The membranes of nonsense (Gln-23----end, Gln-29----end, Gln-74----end) and missense (Gly-150----Asp) mutants had very low ATPase activities, indicating that the delta subunit is essential for the binding of the Fl portion to Fo. The Gln-176----end mutant had essentially the same membrane-bound activity as the wild type, whereas in the Val-174----end mutant most of the ATPase activity was in the cytoplasm. Thus Val-174 (and possibly Leu-175 also) was essential for maintaining the structure of the subunit, whereas the two carboxyl terminal residues Gln-176 and Ser-177 were dispensable. Substitutions were introduced at various residues (Thr-11, Glu-26, Asp-30, Glu-42, Glu-82, Arg-85, Asp-144, Arg-154, Asp-161, Ser-163), including apparently conserved hydrophilic ones. The resulting mutants had essentially the same phenotypes as the wild type, indicating that these residues do not have any significant functional role(s). Analysis of mutations (Gly-150----Asp, Pro, or Ala) indicated that Gly-150 itself was not essential, but that the mutations might affect the structure of the subunit. These results suggest that the overall structure of the delta subunit is necessary, but that individual residues may not have strict functional roles.
Jounouchi
M
M
Department of Organic Chemistry and Biochemistry, Osaka University, Japan.
Takeyama
M
M
Chaiprasert
P
P
Noumi
T
T
Moriyama
Y
Y
Maeda
M
M
Futai
M
M
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003062:Codon / Q000235:genetics; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D007700:Kinetics; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009838:Oligodeoxyribonucleotides; D006180:Proton-Translocating ATPases / Q000235:genetics / Q000378:metabolism*; D015183:Restriction Mapping; D012689:Sequence Homology, Nucleic Acid
D003062:Codon; D046911:Macromolecular Substances; D009838:Oligodeoxyribonucleotides; D006180:Proton-Translocating ATPases