title: Proteomic analysis of scaffold proteins in the ERK cascade.
abstract: ERK cascade scaffolds serve as docking platforms to coordinate the assembly of multiprotein complexes that contribute to the spatial and temporal control of ERK signaling. Given that protein-protein interactions are essential for scaffold function, determining the full repertoire of scaffold binding partners will likely provide new insight into the regulation and activities of the ERK cascade scaffolds. In this chapter, we describe methods to identify scaffold interacting proteins using a proteomics approach. This protocol is based on the affinity purification of scaffold complexes from tissue culture cells and utilizes mass spectrometry to identify the protein constituents of the complex.
authors.affiliation: Laboratory of Cell and Developmental Signaling, Center for Cancer Research, National Cancer Institute-Frederick, Frederick, MD, USA.
authors.forename: Melissa M
authors.initials: MM
authors.lastname: McKay
authors.forename: Deborah K
authors.initials: DK
authors.lastname: Morrison
mesh_terms: D048049:Extracellular Signal-Regulated MAP Kinases / Q000302:isolation & purification / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D020935:MAP Kinase Signaling System; D013058:Mass Spectrometry; D040901:Proteomics / Q000379:methods*; D012117:Resins, Synthetic
chemical_list: D012117:Resins, Synthetic; D048049:Extracellular Signal-Regulated MAP Kinases
references.citation: Science. 1998 Sep 11;281(5383):1668-71
references.pmid: 9733512
references.citation: Mol Cell. 2001 Nov;8(5):983-93
references.pmid: 11741534
references.citation: Genes Dev. 1996 Nov 1;10(21):2684-95
references.pmid: 8946910
references.citation: J Cell Sci. 2005 Aug 15;118(Pt 16):3569-72
references.pmid: 16105880
references.citation: J Biol Chem. 1982 Nov 10;257(21):12467-70
references.pmid: 6290466
references.citation: Dev Cell. 2004 Jul;7(1):33-44
references.pmid: 15239952
references.citation: Growth Factors. 2006 Mar;24(1):21-44
references.pmid: 16393692
references.citation: Nat Rev Mol Cell Biol. 2005 Nov;6(11):827-37
references.pmid: 16227978
references.citation: Annu Rev Cell Dev Biol. 2003;19:91-118
references.pmid: 14570565
references.citation: Mol Cell. 2004 Oct 22;16(2):257-67
references.pmid: 15494312
references.citation: Mol Cell Biol. 2005 Sep;25(18):7940-52
references.pmid: 16135787
references.citation: Curr Biol. 2007 Jan 23;17(2):179-84
references.pmid: 17174095
references.citation: Biochim Biophys Acta. 2007 Aug;1773(8):1213-26
references.pmid: 17112607
references.citation: Mol Cell Biol. 2009 Mar;29(5):1338-53
references.pmid: 19114553
references.citation: Curr Biol. 2003 Aug 19;13(16):1356-64
references.pmid: 12932319
references.citation: Mol Cell. 2009 Jun 26;34(6):652-62
references.pmid: 19560418
references.citation: Proc Natl Acad Sci U S A. 1985 Dec;82(23):7952-4
references.pmid: 2415976