title: Partition and turnover of glutathione reductase from Saccharomyces cerevisiae: a proteomic approach.
abstract: Glutathione reductase (Glr1) is a low abundance protein involved in defense mechanisms against reactive oxygen species. Expressed on cytosolic ribosomes, the same gene, GLR1, uses alternative start codons to generate two forms of Glr1. Translation from the first AUG codon generates the mitochondrial form incorporating a presequence necessary for import; translation from the second AUG codon yields the cytosolic counterpart. Proteomic strategies were used to analyze the N-terminal sequences and the turnover of Saccharomyces cerevisiae Glr1. The N-terminus of cytosolic Glr1 was found normally to be N-acetylserine. When a Glr1-overproducing strain was employed, unprocessed mitochondrial Glr-1 with N-terminal acetylmethionine also accumulated in the cytosol. The processed mitochondrial Glr1 was surprisingly found to have three alternative N-termini, none of them acetylated. Mitochondrial Glr1 was turned over faster than the cytosolic form by a factor of about 2, consistent with the importance of redox homeostasis in the mitochondria. These experiments also allowed us to estimate the extent of "leaky scanning" in the synthesis of Glr1. Surprisingly, the second AUG appears to be responsible for most of the cellular Glr1. This is the first report of protein turnover measurements of a low-abundance protein distributed in different compartments of a eukaryotic cell.
authors: Manchester Institute of Biotechnology, University of Manchester, 131 Princess Street, Manchester M1 7DN, United Kingdom.
authors: Narciso
authors: N
authors: Couto
authors: Naglis
authors: N
authors: Malys
authors: Simon J
authors: SJ
authors: Gaskell
authors: Jill
authors: J
authors: Barber
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D018387:Codon, Initiator; D003600:Cytosol / Q000737:chemistry / Q000201:enzymology; D015966:Gene Expression Regulation, Fungal; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D008928:Mitochondria / Q000737:chemistry / Q000201:enzymology; D008969:Molecular Sequence Data; D010442:Peptide Chain Initiation, Translational / Q000235:genetics*; D017434:Protein Structure, Tertiary; D040901:Proteomics; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
chemical_list: D018387:Codon, Initiator; C508746:GLR1 protein, S cerevisiae; D007527:Isoenzymes; D029701:Saccharomyces cerevisiae Proteins