title: Generating conformation-specific synthetic antibodies to trap proteins in selected functional states.
abstract: A set of phage display sorting strategies and validation methodologies are presented that are capable of producing high performance synthetic antibodies (sABs) with customized properties. Exquisite control of antigen and conditions during the phage display selection process can yield sABs that: (1) recognize conformational states, (2) target specific regions of the surface of a protein, (3) induce conformational changes, and (4) capture and stabilize multi-protein complexes. These unique capabilities open myriad opportunities to study complex macromolecular processes inaccessible to traditional affinity reagent technology. We present detailed protocols for de novo isolation of binders, as well as examples of downstream biophysical characterization. The methods described are generalizable and can be adapted to other in vitro direct evolution approaches based on yeast or mRNA display.
authors.affiliation: Department of Biochemistry and Molecular Biology, The University of Chicago, Chicago, IL 60637, USA.
authors.forename: Marcin
authors.initials: M
authors.lastname: Paduch
authors.forename: Akiko
authors.initials: A
authors.lastname: Koide
authors.forename: Serdar
authors.initials: S
authors.lastname: Uysal
authors.forename: Shahir S
authors.initials: SS
authors.lastname: Rizk
authors.forename: Shohei
authors.initials: S
authors.lastname: Koide
authors.forename: Anthony A
authors.initials: AA
authors.lastname: Kossiakoff
mesh_terms: D000906:Antibodies / Q000737:chemistry* / Q000378:metabolism; D000918:Antibody Specificity; D004797:Enzyme-Linked Immunosorbent Assay; D008968:Molecular Conformation; D019151:Peptide Library; D011506:Proteins / Q000737:chemistry / Q000378:metabolism*
chemical_list: D000906:Antibodies; D019151:Peptide Library; D011506:Proteins