title: Simplified Enrichment of Plasma Membrane Proteins from Arabidopsis thaliana Seedlings Using Differential Centrifugation and Brij-58 Treatment.
abstract: The plasma membrane (PM) forms a barrier between a plant cell and its environment. Proteins at this subcellular location play diverse and complex roles, including perception of extracellular signals to coordinate cellular changes. Analyses of PM proteins, however, are often limited by the relatively low abundance of these proteins in the total cellular protein pool. Techniques traditionally used for enrichment of PM proteins are time consuming, tedious, and require extensive optimization. Here, we provide a simple and reproducible enrichment procedure for PM proteins from Arabidopsis thaliana seedlings starting from total microsomal membranes isolated by differential centrifugation. To enrich for PM proteins, total microsomes are treated with the nonionic detergent Brij-58 to decrease the abundance of contaminating organellar proteins. This protocol combined with the genetic resources available in Arabidopsis provides a powerful tool that will enhance our understanding of proteins at the PM.
authors.affiliation: Division of Biochemistry, Interdisciplinary Plant Group (IPG), University of Missouri, Columbia, MO, 65211, USA.
authors.forename: Carina A
authors.initials: CA
authors.lastname: Collins
authors.affiliation: Division of Biochemistry, Interdisciplinary Plant Group (IPG), University of Missouri, Columbia, MO, 65211, USA.
authors.forename: Michelle E
authors.initials: ME
authors.lastname: Leslie
authors.affiliation: Division of Biochemistry, Interdisciplinary Plant Group (IPG), University of Missouri, Columbia, MO, 65211, USA. pecks@missouri.edu.
authors.forename: Scott C
authors.initials: SC
authors.lastname: Peck
authors.affiliation: Division of Biochemistry, Interdisciplinary Plant Group (IPG), University of Missouri, Columbia, MO, 65211, USA.
authors.forename: Antje
authors.initials: A
authors.lastname: Heese
mesh_terms: D017360:Arabidopsis / Q000737:chemistry* / Q000378:metabolism; D029681:Arabidopsis Proteins / Q000302:isolation & purification*; D002458:Cell Fractionation / Q000379:methods*; D002462:Cell Membrane / Q000737:chemistry* / Q000378:metabolism; D002498:Centrifugation / Q000295:instrumentation / Q000379:methods; D002592:Cetomacrogol / Q000737:chemistry; D008565:Membrane Proteins / Q000302:isolation & purification*; D008861:Microsomes / Q000737:chemistry / Q000378:metabolism; D059828:Plant Cells / Q000737:chemistry / Q000378:metabolism; D036226:Seedlings / Q000737:chemistry* / Q000378:metabolism; D013501:Surface-Active Agents / Q000737:chemistry
chemical_list: D029681:Arabidopsis Proteins; D008565:Membrane Proteins; D013501:Surface-Active Agents; D002592:Cetomacrogol
keywords: Arabidopsis; Brij-58 detergent; Cell fractionation; Differential centrifugation; Plasma membrane; Protein enrichment; Receptor-like kinases (RLKs); Seedlings; Ultracentrifugation