Detection of purine cytosine permease of S. cerevisiae: use of antibodies against a synthetic peptide corresponding to a predicted sequence in the N-terminal domain of the protein.
A synthetic peptide, selected in the predicted N-terminal amino-acid sequence of the purine cytosine permease (gene FCY2), linked to albumins proved a remarkably good immunogen in rabbits. In ELISA, sera reacted with the synthetic peptide and with specific proteins of plasma-membrane-enriched fractions of mutant Saccharomyces cerevisiae pAB strains (amplified FCY2 gene) with high titers and high avidity. Western blots of plasma membrane proteins of pAB strain probed with antisera showed two bands: a major (45 kDa) and minor band (50 kDa). On the contrary, plasma-membrane-enriched fractions of mutant S. cerevisiae pJDB strain (deficient in FCY2 gene) gave no signal when probed in the same conditions. These results demonstrate the specificity of the antisera and also suggest that the 45 kDa and 50 kDa proteins are both products of the FCY2 gene.
Grandier-Vazeille
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Institut de Biochimie Cellulaire, Centre National de la Recherche Scientifique, Bordeaux, France.
Neaud
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Geoffre
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D000595:Amino Acid Sequence; D000906:Antibodies; D000936:Antigen-Antibody Complex; D015153:Blotting, Western; D002352:Carrier Proteins / Q000032:analysis* / Q000235:genetics / Q000276:immunology; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D017353:Gene Deletion; D005800:Genes, Fungal; D007106:Immune Sera; D007700:Kinetics; D026901:Membrane Transport Proteins / Q000032:analysis* / Q000235:genetics / Q000276:immunology; D008969:Molecular Sequence Data; D033702:Nucleobase Transport Proteins; D010455:Peptides / Q000138:chemical synthesis* / Q000276:immunology; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins
D000906:Antibodies; D000936:Antigen-Antibody Complex; D002352:Carrier Proteins; C043181:FCY2 protein, S cerevisiae; D007106:Immune Sera; D026901:Membrane Transport Proteins; D033702:Nucleobase Transport Proteins; D010455:Peptides; D029701:Saccharomyces cerevisiae Proteins