Effects of phosphate and hydrophobic molecules on two mutations in the beta-strand sector of the H(+)-ATPase from the yeast plasma membrane.
At concentrations from 10 to 100 mM, inorganic phosphate and sulfate stimulate the activity of the H(+)-ATPase purified from the wild type Schizosaccharomyces pombe plasma membranes. Compared to the wild type ATPase, the stimulation by phosphate is more pronounced in the mutant pma1-1 (Gly-268----Asp) and is much reduced in the mutant pma1-2 (Lys-250----Thr) enzymes. In contrast, the inhibition by trifluoperazine is less pronounced in the pma1-1 mutant than in the wild type or pma1-2 mutant. The mutant pma1-2 ATPase activity is markedly stimulated by 10-20% dimethyl sulfoxide, which has a limited effect on the wild type and pma1-1 enzymes. These data indicate that the protein domain located in the beta-strand sector, including Lys-250 and Gly-268, is located in the active site and that its hydrophobic character influences the interactions of the yeast H(+)-ATPase with inorganic phosphate, as well as with the hydrophobic inhibitor trifluoperazine or the hydrophobic solvent dimethyl sulfoxide.
Goffeau
A
A
Unité de Biochimie Physiologique, Université de Louvain, Louvain-la-Neuve, Belgium.
de Meis
L
L
D002462:Cell Membrane / Q000201:enzymology; D004121:Dimethyl Sulfoxide / Q000494:pharmacology*; D005998:Glycine; D008239:Lysine; D009154:Mutation; D010710:Phosphates / Q000494:pharmacology*; D006180:Proton-Translocating ATPases / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D004718:Saccharomycetales / Q000201:enzymology*; D012568:Schizosaccharomyces / Q000201:enzymology* / Q000235:genetics; D014268:Trifluoperazine / Q000494:pharmacology*
D010710:Phosphates; D014268:Trifluoperazine; D006180:Proton-Translocating ATPases; D008239:Lysine; D005998:Glycine; D004121:Dimethyl Sulfoxide