title: Cell surface display yields evolvable, clickable antibody fragments.
abstract: Non-canonical amino acids (ncAAs) provide powerful tools for engineering the chemical and physical properties of proteins. However, introducing ncAAs into proteins can affect protein properties in unpredictable ways, thus necessitating screening efforts to identify mutants with desirable properties. In this work, we describe an Escherichia coli cell surface display platform for the directed evolution of clickable antibody fragments. This platform enabled isolation of antibody fragments with improved digoxigenin binding and modest affinity maturation in several different ncAA contexts. Azide-functionalized fragments exhibited improved binding kinetics relative to their methionine counterparts, facile chemical modification through azide-alkyne cycloaddition, and retention of binding properties after modification. The results described here suggest new possibilities for protein engineering, including modulation of molecular recognition events by ncAAs and direct screening of libraries of chemically modified proteins.
authors.affiliation: Division of Chemistry and Chemical Engineering, California Institute of Technology, 1200 E. California Boulevard, MC 210-41, Pasadena, CA 91125 (USA); The Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, MA 02139 (USA).
authors.forename: James A
authors.initials: JA
authors.lastname: Van Deventer
authors.forename: Kai P
authors.initials: KP
authors.lastname: Yuet
authors.forename: Tae Hyeon
authors.initials: TH
authors.lastname: Yoo
authors.forename: David A
authors.initials: DA
authors.lastname: Tirrell
mesh_terms: D000596:Amino Acids / Q000737:chemistry / Q000276:immunology; D000915:Antibody Affinity / Q000276:immunology; D057930:Click Chemistry; D004076:Digoxigenin / Q000737:chemistry / Q000276:immunology; D019020:Directed Molecular Evolution; D004926:Escherichia coli / Q000166:cytology / Q000276:immunology*; D005434:Flow Cytometry; D007128:Immunoglobulin Fragments / Q000737:chemistry* / Q000276:immunology* / Q000302:isolation & purification; D008958:Models, Molecular; D015394:Molecular Structure
chemical_list: D000596:Amino Acids; D007128:Immunoglobulin Fragments; D004076:Digoxigenin
keywords: antibodies; click chemistry; directed evolution; high-throughput screening; non-canonical amino acids