The active transport of monosaccharides by the yeast Metschnikowia reukaufii: Evidence for an electrochemical gradient of H + across the cell membrane

Abstract

The facultative anaerobic yeast Metschnikowia reukaufii was found to take up the monosaccharides d-glucose, d-xylose, and 3- O-methyl- d-glucose by a constitutive mobile membrane carrier. The uptake was dependent on the supply of metabolic energy. The half-saturation constants of transport, K T , amounted to 0.9, 2.0, and 1.0 m m, respectively. Cell suspensions of M. reukaufii catabolized d-glucose and d-xylose, but not 3- O-methyl- d-glucose. d-Xylose was accumulated about 100-fold above the diffusion equilibrium in spite of its catabolism. 3- O-Methyl- d-glucose was accumulated about 40-fold. There was no accumulation of monosaccharides in the presence of uncouplers like carbonylcyanide m-chlorophenylhydrazone or dinitrophenol. For d-glucose utilization, its transport was the ratelimiting step. All three monosaccharides were transported by the same carrier; in a mixture they competed for the binding site at the carrier. An electrochemical gradient of H + ions was measured across the plasma membrane of M. reukaufii cells. The pH difference between the cell interior and the medium of an unbuffered suspension (pH 4–5) reached the value of 2.0 pH units (inside alkaline). The membrane potential measured with the lipid-soluble cation tetraphenylphosphonium at pH 9.0 was about - 100 mV (inside negative). The onset of sugar transport brought about a short alkalinization of unbuffered cell suspensions or a depolarization of the membrane potential in buffered suspensions (pH 9.0). The H +/sugar stoichiometry was one H + ion per each sugar molecule taken up. The data are interpreted as evidence in favor of the active monosaccharide transport in M. reukaufii being an H + symport, energized by the electrochemical gradient of H + across the plasma membrane.