title: Proteins interacting with Saccharomyces cerevisiae type 1 protein phosphatase catalytic subunit identified by single-step affinity purification and mass spectrometry.
abstract: The catalytic subunit of type 1 protein phosphatase (PP1C) interacts with a large number of polypeptides in eukaryotic cells from yeast to man and these regulatory subunits can both modulate the activity of PP1C and target it to different subcellular locations. Thus, PP1 is really a family of protein phosphatases that share a common catalytic subunit, and identifying and characterizing the PP1-associated proteins is therefore critical to understanding the cellular roles of PP1 and its ability to dephosphorylate specific substrates. Here we describe methods for affinity isolation of PP1C-containing protein complexes in the yeast Saccharomyces cerevisiae and the identification of the associated polypeptides by mass spectrometry. The basic method we describe could be easily adapted to study PP1C-associated proteins in other lower or higher eukaryotes and for characterizing the protein-protein interactions of other protein phosphatases in yeast.
authors.affiliation: University of Newcastle, UK.
authors.forename: Edmund P
authors.initials: EP
authors.lastname: Walsh
authors.forename: Douglas J
authors.initials: DJ
authors.lastname: Lamont
authors.forename: Kenneth A
authors.initials: KA
authors.lastname: Beattie
authors.forename: Michael J R
authors.initials: MJ
authors.lastname: Stark
mesh_terms: D000595:Amino Acid Sequence; D020134:Catalytic Domain; D002846:Chromatography, Affinity; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010749:Phosphoprotein Phosphatases / Q000378:metabolism*; D011485:Protein Binding; D054645:Protein Phosphatase 1; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*
chemical_list: D029701:Saccharomyces cerevisiae Proteins; D010749:Phosphoprotein Phosphatases; D054645:Protein Phosphatase 1