title: Isolation of an scFv targeting BRG1 using phage display with characterization by AFM.
abstract: Remodeling of chromatin is a vitally important event in processes such as transcription and replication. Brahma-related gene 1 (BRG1) protein is the major ATPase subunit in the human Swi/Snf complex (hSwi/Snf), an important example of the family of enzymes that carry out such remodeling events. We have used a recently developed technique, recognition imaging, to better understand the role of BRG1 in remodeling chromatin. In such experiments, a specific antibody against BRG1 is needed. However, we have found that the commercially available polyclonal (CAP) antibodies interact non-specifically with nucleosomes, making it impossible to identify hSwi/Snf (BRG1) in their presence. Here antibody phage display technology is employed for development of an antibody specifically targeting BRG1. The Tomlinson I and J single chain variable fragment (scFv) libraries were used for successful isolation of an anti-BRG1 scFv. We demonstrate that the scFv binds more strongly and with less nonspecific interactions than the CAP antibody. This work lays the groundwork for future studies involving chromatin remodeling.
authors.affiliation: Department of Physics and Astronomy, Arizona State University, Tempe, AZ 85287, USA. wdmarcus@alum.mit.edu
authors.forename: W D
authors.initials: WD
authors.lastname: Marcus
authors.forename: H
authors.initials: H
authors.lastname: Wang
authors.forename: D
authors.initials: D
authors.lastname: Lohr
authors.forename: M R
authors.initials: MR
authors.lastname: Sierks
authors.forename: S M
authors.initials: SM
authors.lastname: Lindsay
mesh_terms: D002478:Cells, Cultured; D002843:Chromatin / Q000276:immunology* / Q000648:ultrastructure*; D042002:Chromatin Assembly and Disassembly / Q000502:physiology; D004265:DNA Helicases; D006801:Humans; D007089:Image Enhancement / Q000379:methods; D007128:Immunoglobulin Fragments / Q000276:immunology / Q000648:ultrastructure; D018625:Microscopy, Atomic Force / Q000379:methods*; D009687:Nuclear Proteins / Q000276:immunology* / Q000648:ultrastructure*; D019151:Peptide Library; D014157:Transcription Factors / Q000276:immunology* / Q000648:ultrastructure*
chemical_list: D002843:Chromatin; D007128:Immunoglobulin Fragments; D009687:Nuclear Proteins; D019151:Peptide Library; D014157:Transcription Factors; C084108:SMARCA4 protein, human; D004265:DNA Helicases