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Optimized Protein Extraction for Quantitative Proteomics of Yeasts

BACKGROUND: The absolute quantification of intracellular protein levels is technically demanding, but has recently become more prominent because novel approaches like systems biology and metabolic control analysis require knowledge of these parameters. Current protocols for the extraction of protein...

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Detalles Bibliográficos
Autor principal: von der Haar, Tobias
Formato: Texto
Lenguaje:English
Publicado: Public Library of Science 2007
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2031916/
https://www.ncbi.nlm.nih.gov/pubmed/17957260
http://dx.doi.org/10.1371/journal.pone.0001078
Descripción
Sumario:BACKGROUND: The absolute quantification of intracellular protein levels is technically demanding, but has recently become more prominent because novel approaches like systems biology and metabolic control analysis require knowledge of these parameters. Current protocols for the extraction of proteins from yeast cells are likely to introduce artifacts into quantification procedures because of incomplete or selective extraction. PRINCIPAL FINDINGS: We have developed a novel procedure for protein extraction from S. cerevisiae based on chemical lysis and simultaneous solubilization in SDS and urea, which can extract the great majority of proteins to apparent completeness. The procedure can be used for different Saccharomycetes yeast species and varying growth conditions, is suitable for high-throughput extraction in a 96-well format, and the resulting extracts can easily be post-processed for use in non-SDS compatible procedures like 2D gel electrophoresis. CONCLUSIONS: An improved method for quantitative protein extraction has been developed that removes some of the sources of artefacts in quantitative proteomics experiments, while at the same time allowing novel types of applications.