Transport of pyruvate in Saccharomyces cerevisiae and cloning of the gene encoded pyruvate permease.
Pyruvate uptake in Saccharomyces cerevisiae was not observed at 0 degrees C and was prevented by the uncoupler carbonyl cyanide m-chlorophenylhydrazone (CCCP). The initial uptake rate of S. cerevisiae kyokai No. 901 was maximum at pH 6 and Km = 4.1 mM. It seemed that lactate inhibited the pyruvate uptake competitively from the results of the Lineweaver-Burk plots. The inhibition constant (Ki) in the presence of 3 mM lactate was 1.6 mM. The pyruvate uptake was inhibited by D-glucose and deoxyglucose, but not by L-glucose, acetate or ethanol. Mutants of laboratory strain No. 5022 ((a) his(2,6), ura3) deficient in pyruvate uptake were isolated from fluoropyruvate resistant mutants. Transformation of the mutant with a yeast genomic library allowed the isolation of the gene JEN1 (YKL217w), which restored pyruvate uptake. Disruption of JEN1 abolished the uptake of pyruvate and gained the resistance against fluoropyruvate. The results indicate that no other monocarboxylate permease is able to efficiently transport pyruvate in S. cerevisiae.
Akita
O
O
National Research Institute of Brewing, Higashi-Hiroshima, Japan. o.akita@nrib.go.jp
Nishimori
C
C
Shimamoto
T
T
Fujii
T
T
Iefuji
H
H
D001692:Biological Transport; D002352:Carrier Proteins / Q000235:genetics* / Q000378:metabolism*; D003001:Cloning, Molecular; D004252:DNA Mutational Analysis; D015698:Genomic Library; D007700:Kinetics; D026901:Membrane Transport Proteins / Q000235:genetics* / Q000378:metabolism; D027501:Monocarboxylic Acid Transporters; D019289:Pyruvic Acid / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D027981:Symporters; D014170:Transformation, Genetic
D002352:Carrier Proteins; C118697:JEN1 protein, S cerevisiae; D026901:Membrane Transport Proteins; D027501:Monocarboxylic Acid Transporters; D029701:Saccharomyces cerevisiae Proteins; D027981:Symporters; D019289:Pyruvic Acid