Mutants that show increased sensitivity to hydrogen peroxide reveal an important role for the pentose phosphate pathway in protection of yeast against oxidative stress.
We have isolated several mutants of Saccharomyces cerevisiae that are sensitive to oxidative stress in a screen for elevated sensitivity to hydrogen peroxide. Two of the sixteen complementation groups obtained correspond to structural genes encoding enzymes of the pentose phosphate pathway. Allelism of the pos10 mutation (POS for peroxide sensitivity) to the zwf1/met1 mutants in the structural gene for glucose 6-phosphate dehydrogenase was reported previously. The second mutation, pos18, was complemented by transformation with a yeast genomic library. The open reading frame of the isolated gene encodes 238 amino acids. No detectable ribulose 5-phosphate epimerase activity was found in the pos18 mutant, suggesting that the corresponding structural gene is affected in this mutant. For that reason the gene was renamed RPE1 (for ribulose 5-phosphate epimerase). RPE1 was localized to chromosome X. The predicted protein has a molecular mass of 25966 Daltons, a codon adaptation index (CAI) of 0.32, and an isoelectric point of 5.82. Database searches revealed 32 to 37% identity with ribulose 5-phosphate epimerases of Escherichia coli, Rhodospirillum rubrum, Alcaligenes eutrophus and Solanum tuberosum. We have characterized RPE1 by testing enzyme activities in rpe1 deletion mutants and in strains that overexpress RPE1, and compared the hydrogen peroxide sensitivity of rpe1 mutants to that of other mutants in the pentose phosphate pathway. Interestingly, all mutants tested (glucose 6-phosphate dehydrogenase, gluconate 6-phosphate dehydrogenase, ribulose 5-phosphate epimerase, transketolase, transaldolase) are sensitive to hydrogen peroxide.
Juhnke
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H
Institut für Mikrobiologie, der Johann Wolfgang Goethe-Universität Frankfurt, Biozentrum Niederursel, Frankfurt am Main, Germany.
Krems
B
B
Kötter
P
P
Entian
K D
KD
D050539:3-Isopropylmalate Dehydrogenase; D000429:Alcohol Oxidoreductases / Q000235:genetics / Q000378:metabolism; D000595:Amino Acid Sequence; D002238:Carbohydrate Epimerases / Q000235:genetics / Q000378:metabolism; D002455:Cell Division / Q000235:genetics; D003001:Cloning, Molecular; D004352:Drug Resistance, Microbial / Q000235:genetics; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism; D017353:Gene Deletion; D005800:Genes, Fungal; D006836:Hydro-Lyases / Q000235:genetics / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D008969:Molecular Sequence Data; D009154:Mutation; D016877:Oxidants / Q000494:pharmacology; D018384:Oxidative Stress; D010427:Pentose Phosphate Pathway / Q000235:genetics*; D010734:Phosphogluconate Dehydrogenase / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000187:drug effects* / Q000235:genetics* / Q000378:metabolism; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D014153:Transaldolase / Q000235:genetics / Q000378:metabolism
D005656:Fungal Proteins; D016877:Oxidants; D006861:Hydrogen Peroxide; D000429:Alcohol Oxidoreductases; D010734:Phosphogluconate Dehydrogenase; C029757:phosphogluconate dehydrogenase (decarboxylating); D050539:3-Isopropylmalate Dehydrogenase; D014153:Transaldolase; D006836:Hydro-Lyases; C021434:imidazoleglycerolphosphate dehydratase; D002238:Carbohydrate Epimerases; C022554:L-ribulosephosphate 4-epimerase