SUMMARYGlycolysis of yeast is inhibited by the polyene antibiotic nystatin. This inhibition was prevented or reversed at or above pH 5.8 (the approximate intracellular pH of yeast) by adding N H : , Kf or Rb' (in order of decreasing effectiveness), These ions do not interfere with the absorption of nystatin by the yeast cell. At pH 4 both glycolysis and respiration are highly sensitive to nystatin but are not protected by the monovalent cations. At pH 7 respiration of intact cells was insensitive; only with yeast protoplasts could respiration be inhibited by nystatin. Temporary protection was obtained with NH,+ or Kf.K+ and NH,+ did not prevent the fungicidal or fungistatic actions of nystatin even under conditions where the protection of glycolysis by these ions was essentially complete. The original yeast cells were not dependent upon added K'. After a brief treatment with nystatin at pH 7.0, the cells showed an absolute requirement for K+ or N H f ; it was shown that K' was rapidly lost by cells following contact with nystatin. It is concluded that nystatin directly damages the cell membrane and thus produces a rapid increase in permeability to small ions. The resulting depletion of cellular Kf halts glycolysis. The addition of K+ or NHZ restores glycolysis but does not reverse the membrane alterations.
The polyene antifungal antibiotics nystatin and N-acetylcandidin were bound rapidly at 0 C by isolated cell walls and derived polysaceharides or by protoplast membranes from Saccharomyces cerevisiae strain LK2G12. These binding sites were relatively inaccessible or unreactive in the intact cell, since polyene uptake by protoplasts, log-phase cells, or stationary-phase cells was slow, especially at 0 C. Binding by the membrane appears to be the critical event in cell damage; thus, uptake of nystatin by the cell wall may actually be protective. B3inding by all cell forms showed little rev-ersibility. Bound radioactive nystatin or N-acetyl (1-C14) candidin was not displaced during incubation of log-phase yeast cells with a large excess of unlabeled polyene. Saturation of the polyenic moiety of nystatin to form the perhy-(Iro compound eliminated almost completely the affinity of the molecule for the yeast cell. Essentially all of the polyene bound by protoplasts was present on the membrane. It was removed by treatment of the protoplasts with the sterol-comp)lexing agent digitonin. A variety of evidence is offered that the binding site on the membrane contains a sterol. (The membrane sterol was mostly unesterified ergosterol.) This hypothesis is consistent with the ability of certain exogenous sterols to complex with polyenes and prevent their binding and growth-inhibiting action for fungi. The bound sterol of the wall structure may also participate in polyene binding, although a lpossible function of the wall polysaceharides cannot be excluded. The specific binding structure(s) appears to be absent from bacteria, since nystatin was not taken up even by heated or benzene
Nystatin, a polyene antibiotic produced by Streptomyces noursei (Brown and Hazen, 1957), is highly active in inhibiting the growth of a wide variety of fungi, but has little or no effect on other types of microorganisms or on animal cells. It inhibits both endogenous metabolic
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