Abstract
We isolated a Pichia pastoris mutant that was unable to grow on the peroxisome-requiring media, methanol and oleate. Cloning the gene by complementation revealed that the encoded protein, Pex22p, is a new peroxin. A Deltapex22 strain does not grow on methanol or oleate and is unable to import peroxisomal matrix proteins. However, this strain targets peroxisomal membrane proteins to membranes, most likely peroxisomal remnants, detectable by fluorescence and electron microscopy. Pex22p, composed of 187 amino acids, is an integral peroxisomal membrane protein with its NH2 terminus in the matrix and its COOH terminus in the cytosol. It contains a 25-amino acid peroxisome membrane-targeting signal at its NH2 terminus. Pex22p interacts with the ubiquitin-conjugating enzyme Pex4p, a peripheral peroxisomal membrane protein, in vivo, and in a yeast two-hybrid experiment. Pex22p is required for the peroxisomal localization of Pex4p and in strains lacking Pex22p, the Pex4p is cytosolic and unstable. Therefore, Pex22p anchors Pex4p at the peroxisomal membrane. Strains that do not express Pex4p or Pex22p have similar phenotypes and lack Pex5p, suggesting that Pex4p and Pex22p act at the same step in peroxisome biogenesis. The Saccharomyces cerevisiae hypothetical protein, Yaf5p, is the functional homologue of P. pastoris Pex22p.
Publication types
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, Non-P.H.S.
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Amino Acid Sequence
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Base Sequence
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Biological Transport
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Carrier Proteins / analysis
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Carrier Proteins / chemistry
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Carrier Proteins / genetics
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Carrier Proteins / metabolism*
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Cloning, Molecular
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Conserved Sequence / genetics
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Cytosol / chemistry
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Cytosol / metabolism
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Cytosol / ultrastructure
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Fungal Proteins*
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Gene Deletion
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Genes, Fungal / genetics
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Genes, Fungal / physiology
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Genetic Complementation Test
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Intracellular Membranes / chemistry
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Intracellular Membranes / enzymology
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Intracellular Membranes / metabolism*
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Intracellular Membranes / ultrastructure
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Ligases / genetics
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Ligases / metabolism*
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Membrane Proteins / analysis
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Membrane Proteins / chemistry
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Membrane Proteins / genetics
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Membrane Proteins / metabolism*
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Membrane Transport Proteins*
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Methanol / metabolism
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Microbodies / chemistry
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Microbodies / enzymology
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Microbodies / metabolism*
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Microbodies / ultrastructure
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Molecular Sequence Data
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Oleic Acid / metabolism
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Phenotype
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Pichia / cytology
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Pichia / genetics
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Pichia / metabolism*
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Pichia / ultrastructure
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Protein Sorting Signals / genetics
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Protein Sorting Signals / physiology
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Recombinant Fusion Proteins / analysis
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Recombinant Fusion Proteins / biosynthesis
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Ubiquitins*
Substances
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Carrier Proteins
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Fungal Proteins
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Membrane Proteins
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Membrane Transport Proteins
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Pex22 protein, Pichia pastoris
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Protein Sorting Signals
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Recombinant Fusion Proteins
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Ubiquitins
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Oleic Acid
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Ligases
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Methanol