The yeast plasma membrane proton pumping ATPase is a viable antifungal target. I. Effects of the cysteine-modifying reagent omeprazole.
The yeast plasma membrane proton pumping ATPase (H(+)-ATPase) was investigated as a potential molecular target for antifungal drug therapy by examining the inhibitory effects of the sulfhydryl-reactive reagent omeprazole on cell growth, glucose-induced medium acidification and H(+)-ATPase activity. Omeprazole inhibits the growth of Saccharomyces cerevisiae and the human pathogenic yeast Candida albicans in a pH dependent manner. Omeprazole action is closely correlated with inhibition of the H(+)-ATPase and is fungicidal. Glucose-dependent medium acidification is correspondingly blocked by omeprazole and appears to require the H(+)-ATPase to proceed through its reaction cycle. A strong correlation is observed between inhibition of medium acidification and H(+)-ATPase activity in plasma membranes isolated from treated cells. The inhibitory properties of omeprazole are blocked by pre-treatment of activated drug with beta-mercaptoethanol, which is consistent with the expected formation of a sulfhydryl-reactive sulfenamide derivative. Mutagenesis of the three putative membrane sector cysteine residues (C148S, C312S, C867A) in the S. cerevisiae H(+)-ATPase suggests that covalent modification of the conserved C148 residue may be important for inhibition of ATPase activity and cell growth. Other mutations (M128C and G158D/G156C) mapping near C148 support the importance of this region by modulating omeprazole inhibition of the H(+)-ATPase. These findings suggest that the plasma membrane H(+)-ATPase may serve as an important molecular target for antifungal intervention.
Monk
B C
BC
Public Health Research Institute, New York, NY 10016, USA.
Mason
A B
AB
Abramochkin
G
G
Haber
J E
JE
Seto-Young
D
D
Perlin
D S
DS
D002176:Candida albicans / Q000187:drug effects* / Q000201:enzymology; D002462:Cell Membrane / Q000201:enzymology; D003545:Cysteine / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005947:Glucose / Q000502:physiology; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008623:Mercaptoethanol / Q000494:pharmacology; D009154:Mutation; D009853:Omeprazole / Q000494:pharmacology*; D054328:Proton Pump Inhibitors; D017494:Proton Pumps / Q000378:metabolism; D006180:Proton-Translocating ATPases / Q000037:antagonists & inhibitors* / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000187:drug effects* / Q000201:enzymology; D029701:Saccharomyces cerevisiae Proteins; D013439:Sulfhydryl Reagents / Q000494:pharmacology*
D004791:Enzyme Inhibitors; C434364:PMA2 protein, S cerevisiae; D054328:Proton Pump Inhibitors; D017494:Proton Pumps; D029701:Saccharomyces cerevisiae Proteins; D013439:Sulfhydryl Reagents; D008623:Mercaptoethanol; C092477:PMA1 protein, S cerevisiae; D006180:Proton-Translocating ATPases; D005947:Glucose; D003545:Cysteine; D009853:Omeprazole