Large-scale analysis of the yeast proteome by multidimensional protein identification technology

We describe a largely unbiased method for rapid and large-scale proteome an alysis by multidimensional liquid chromatography, tandem mass spectrometry, and database searching by the SEQUEST algorithm, named multidimensional pr otein identification technology (MudPIT). MudPIT was applied to the proteom e of the Saccharomyces cerevisiae strain BJ5460 grown to mid-log phase and yielded the largest proteome analysis to date. A total of 1,484 proteins we re detected and identified. Categorization of these hits demonstrated the a bility of this technology to detect and identify proteins rarely seen in pr oteome analysis, including low-abundance proteins like transcription factor s and protein kinases. Furthermore, we identified 131 proteins with three o r more predicted transmembrane domains, which allowed us to map the soluble domains of many of the integral membrane proteins. MudPIT is useful for pr oteome analysis and may be specifically applied to integral membrane protei ns to obtain detailed biochemical information on this unwieldy class of pro teins.