Identification by mass spectrometry of two-dimensional gel electrophoresis-separated proteins extracted from lager brewing yeast

Abstract

As two-dimensional (2-D) electrophoresis allows the separation of several hundred proteins in a single gel, this technique has become an important tool for proteome studies and for investigating the cellular physiology. In order to take advantage of information provided by the comparison of proteome pictures, the mass spectrometry technique is the way chosen for a rapid and an accurate identification of proteins of interest. Unfortunately, in the case of industrial yeasts, due to the high level of complexity of their genome, the whole DNA sequence is not yet available and all encoded protein sequences are still unknown. Nevertheless, this study presents here 30 lager brewing yeast proteins newly identified with matrix assisted laser desorption/ionization-time of flight (MALDI-TOF), tandem mass spectrometry (MS/MS) and database searching against the protein sequences of Saccharomyces cerevisiae. The identified proteins of the industrial strain correspond to proteins which do not comigrate with known proteins of S. cerevisiae separated on 2-D gels. This study presents an application of the MS technique for the identification of industrial yeast proteins which are only homologous to the corresponding S. cerevisiae proteins.

Progress in Metabolic Engineering of Saccharomyces cerevisiae

“…Proteomics has also been hampered by the lack of sequences encoding the non-Saccharomyces genes/proteins in lager brewer's yeast. Researchers have put much effort into identifying non-Saccharomyces protein spots by sequence homologies (152). The identification of the entire genomic sequence of a commonly used lager brewer's yeast strain, i.e., Weihenstephan Nr.…”

Crucial Role of Antioxidant Proteins and Hydrolytic Enzymes in Pathogenicity of Penicillium expansum

“…Due to the lack of genome sequence information for this species, protein identification provided an analytical challenge. We used MS/MS, which has been reported to be the most successful technique to correctly identify proteins from organisms with the whole DNA sequence unknown (40), to identify these proteins. MS/MS spectra were submitted for database searching in Mascot search engine.…”

Development and application of proteomics technologies in Saccharomyces cerevisiae

“…Also, subproteome reference maps of, for example, yeast mitochondria, have been generated [30]. Moreover, 2D reference maps have been constructed for important industrial yeast strains, such as an ale-fermenting strain [31], a wine strain [32] and a lager-brewing strain [33,34]. These annotated reference maps are useful tools for yeast researchers because they can be used for 2D gel comparisons; however, because of poor gel-to-gel reproducibility and strain variation, protein spot identities should always be confirmed using MS.…”