RELATIONSHIP BETWEEN EFFECT OF ETHANOL ON PROTON FLUX ACROSS PLASMA-MEMBRANE AND ETHANOL TOLERANCE, IN PICHIA-STIPITIS

Pichia stipitis efficiently converts glucose or xylose into ethanol bu t is inhibited by ethanol concentrations exceeding 30 g/L. In Saccharo myces cerevisiae, ethanol has been shown to alter the movement of prot ons into and out of the cell. In P. stipitis the passive entry of prot ons into either glucose- or xylose-grown cells is unaffected at physio logical ethanol concentrations. In contrast, active proton extrusion i s affected differentially by ethanol, depending on the carbon source c atabolized. In fact, in glucose-grown cells, the H+-extrusion rate is reduced by low ethanol concentrations, whereas, in xylose-grown cells, the H+-extrusion rate is reduced only at non-physiological ethanol co ncentrations. Thus, the ethanol inhibitory effect on growth and ethano l production, in glucose-grown cells, is probably caused by a reductio n in H+-extrusion. Comparison of the rates of H+-flux with the related in vitro H+-ATPase activity suggests a new mechanism for the regulati on of the proton pumping plasma membrane ATPase (EC 3.6.1.3) of P. sti pitis, by both glucose and ethanol. Glucose activates both the ATP hyd rolysis and the proton-pumping activities of the H+-ATPase, whereas et hanol causes an uncoupling between the ATP hydrolysis and the proton-p umping activities. This uncoupling may well be the cause of ethanol in duced growth inhibition of glucose grown I? stipitis cells. (C) 1997 A cademic Press.