Nuclear thiol peroxidase as a functional alkyl-hydroperoxide reductase necessary for stationary phase growth of Saccharomyces cerevisiae.
Yeast nucleus-localized thiol peroxidase (nTPx) was characterized as a functional peroxidase. There are two cysteine residues in nTPx. Replacement of Cys-106 or Cys-111 with serine resulted in a complete loss of thioredoxin-linked peroxidase activity. However, when their activities were measured in terms of the ability to inhibit oxidation of glutamine synthetase, C111S showed the same antioxidant activity as the wild type protein. SDS-PAGE gel analysis revealed that only C111S existed as the dimer form. In addition to the identification of Cys-106 as the primary catalytic site, these data suggest the formation of the intradisulfide bond as a part of the catalytic cycle between nTPx and thioredoxin. nTPx preferentially reduced alkyl-hydroperoxides rather than H2O2. Furthermore, a nTPx mutant strain showed higher sensitivity toward alkyl-hydroperoxide than hydrogen peroxide. Also, reduction of the viability of nTPx mutant strain against various oxidants supports an in vivo antioxidant role for nTPx. nTPx transcriptional activity was not significantly detectable in log phase yeast, but the activity was exponentially increased after the diauxic shift. The transcriptional activity was highly induced even in the log phase yeast grown in nonfermentable carbon source. Deletion of Tor1p, Ras1p, and Ras2p resulted in considerable induction when compared with their parent strains, demonstrating the activation of the transcription of nTPx gene at the diauxic shift. Transcription of nTPx gene was induced in response to oxidative stress. Viability of a stationary phase nTPx mutant was considerably reduced compared with the isogenic strain. Collectively, these data demonstrate that nTPx is a thiol peroxidase family acting as alkyl-hydroperoxide reductase in the nucleus during post-diauxic growth.
Department of Biochemistry, Paichai University, Taejon 302-735, Republic of Korea.
Mee-Kyung
MK
Cha
Yong-Soo
YS
Choi
Seung-Keun
SK
Hong
Won-Cheol
WC
Kim
Kyoung Tai
KT
No
Il-Han
IH
Kim
D015966:Gene Expression Regulation, Fungal / Q000502:physiology; D009154:Mutation; D010544:Peroxidases / Q000235:genetics / Q000502:physiology*; D054464:Peroxiredoxins; D011401:Promoter Regions, Genetic; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000254:growth & development; D013438:Sulfhydryl Compounds / Q000378:metabolism; D014158:Transcription, Genetic
D013438:Sulfhydryl Compounds; D010544:Peroxidases; D054464:Peroxiredoxins