Identification of proteins of the yeast protein map using genetically manipulated strains and peptide-mass fingerprinting

Abstract

In this study we used genetically manipulated strains in order to identify polypeptide spots of the protein map of Saccharomyces cerevisiae. Thirty‐two novel polypeptide spots were identified using this strategy. They corresponded to the product of 23 different genes. We also explored the possibilities of using peptide‐mass fingerprinting for the identification of proteins separated on our gels. According to this strategy, proteins contained in spots are digested with trypsin and the masses of generated peptides are determined by matrix‐assisted laser desorption‐ionization mass spectrometry (MALDI‐MS). The peptide masses are then used to search a yeast protein database for proteins that match the experimental data. Application of this strategy to previously identified polypeptide spots gave evidence of the feasibility of this approach. We also report predictions on the identities of nine unknown spots using MALDI‐MS.