title: When monomers are preferred: a strategy for the identification and disruption of weakly oligomerized proteins.
abstract: Oligomerization is important for the structure and function of many proteins, but frequently complicates their characterization. It is often desirable to obtain the protein in monomeric form. Here, we report a strategy that allows the generation of monomers from weakly associated oligomers but does not require knowledge of the three-dimensional structure of the protein. The dynamics of protein association are used in solution NMR spectroscopy to identify regions of the polypeptide chain that are likely to be responsible for the interaction. Protein sequence analysis further refines the selection, as conserved sites with moderate hydrophobicity are targeted for modification. Gel filtration and activity assays straightforwardly reveal the consequences of the change and are used to screen for the desired mutants. The strategy is demonstrated for the Rac1 binding domain of plexin-B1. A monomeric variant is generated which preserves the Rac1 binding activity and the wild-type protein structure.
authors.affiliation: Department of Physiology and Biophysics, Case Western Reserve University School of Medicine, Cleveland, Ohio 44106, USA.
authors.forename: Yufeng
authors.initials: Y
authors.lastname: Tong
authors.forename: David
authors.initials: D
authors.lastname: Hughes
authors.forename: Lisa
authors.initials: L
authors.lastname: Placanica
authors.forename: Matthias
authors.initials: M
authors.lastname: Buck
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D002850:Chromatography, Gel; D016384:Consensus Sequence; D019281:Dimerization; D057927:Hydrophobic and Hydrophilic Interactions; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D019906:Nuclear Magnetic Resonance, Biomolecular; D017354:Point Mutation; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D020539:Sequence Analysis, Protein; D017386:Sequence Homology, Amino Acid; D012996:Solutions
chemical_list: D011506:Proteins; D012996:Solutions