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Chembiochem. 2014 Aug 18;15(12):1777-81. doi: 10.1002/cbic.201402184. Epub 2014 Jul 17.

Cell surface display yields evolvable, clickable antibody fragments.

Author information

1
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).

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.

KEYWORDS:

antibodies; click chemistry; directed evolution; high-throughput screening; non-canonical amino acids

PMID:
25045032
PMCID:
PMC4199383
DOI:
10.1002/cbic.201402184
[Indexed for MEDLINE]
Free PMC Article

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