title: A peptide affinity reagent for isolating an intact and catalytically active multi-protein complex from mammalian cells.
abstract: We have developed an approach for directly isolating an intact multi-protein chromatin remodeling complex from mammalian cell extracts using synthetic peptide affinity reagent 4. FOG1(1-15), a short peptide sequence known to target subunits of the nucleosome remodeling and deacetylase (NuRD) complex, was joined via a 35-atom hydrophilic linker to the StreptagII peptide. Loading this peptide onto Streptactin beads enabled capture of the intact NuRD complex from MEL cell nuclear extract. Gentle biotin elution yielded the desired intact complex free of significant contaminants and in a form that was catalytically competent in a nucleosome remodeling assay. The efficiency of 4 in isolating the NuRD complex was comparable to other reported methods utilising recombinantly produced GST-FOG1(1-45).
authors: School of Molecular Bioscience, The University of Sydney, Sydney, NSW 2006, Australia.
authors: Hinnerk
authors: H
authors: Saathoff
authors: School of Molecular Bioscience, The University of Sydney, Sydney, NSW 2006, Australia.
authors: Mattias
authors: M
authors: Brofelth
authors: School of Molecular Bioscience, The University of Sydney, Sydney, NSW 2006, Australia.
authors: Anne
authors: A
authors: Trinh
authors: School of Molecular Bioscience, The University of Sydney, Sydney, NSW 2006, Australia.
authors: Benjamin L
authors: BL
authors: Parker
authors: John Curtin School of Medical Research, The Australian National University, Canberra, ACT 0200, Australia.
authors: Daniel P
authors: DP
authors: Ryan
authors: School of Molecular Bioscience, The University of Sydney, Sydney, NSW 2006, Australia.
authors: Jason K K
authors: JK
authors: Low
authors: School of Molecular Bioscience, The University of Sydney, Sydney, NSW 2006, Australia.
authors: Sarah R
authors: SR
authors: Webb
authors: School of Molecular Bioscience, The University of Sydney, Sydney, NSW 2006, Australia.
authors: Ana P G
authors: AP
authors: Silva
authors: School of Molecular Bioscience, The University of Sydney, Sydney, NSW 2006, Australia.
authors: Joel P
authors: JP
authors: Mackay
authors: School of Molecular Bioscience, The University of Sydney, Sydney, NSW 2006, Australia. Electronic address: nicholas.shepherd@sydney.edu.au.
authors: Nicholas E
authors: NE
authors: Shepherd
mesh_terms: D000345:Affinity Labels; D000595:Amino Acid Sequence; D000818:Animals; D002384:Catalysis; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D056245:Mi-2 Nucleosome Remodeling and Deacetylase Complex / Q000302:isolation & purification* / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014407:Tumor Cells, Cultured
chemical_list: D000345:Affinity Labels; D010455:Peptides; D056245:Mi-2 Nucleosome Remodeling and Deacetylase Complex
keywords: Affinity purification; Friend of GATA1 (FOG1); Multi-protein complex; Nucleosome remodeling and deacetylase complex peptides