Unit

Site-Specific Protein Bioconjugation via a Pyridoxal 5′-Phosphate-Mediated N-Terminal Transamination Reaction

  1. Leah S. Witus,
  2. Matthew Francis

Published Online: 1 JUN 2010

DOI: 10.1002/9780470559277.ch100018

Current Protocols in Chemical Biology

Current Protocols in Chemical Biology

How to Cite

Witus, L. S. and Francis, M. 2010. Site-Specific Protein Bioconjugation via a Pyridoxal 5′-Phosphate-Mediated N-Terminal Transamination Reaction. Current Protocols in Chemical Biology. 2:125–134.

Author Information

  1. University of California, Berkeley, Department of Chemistry, Berkeley, California

Publication History

  1. Published Online: 1 JUN 2010
  2. Published Print: JUN 2010

Abstract

The covalent attachment of chemical groups to proteins is a critically important tool for the study of protein function and the creation of protein-based materials. Methods of site-specific protein modification are necessary for the generation of well defined bioconjugates possessing a new functional group in a single position in the amino acid sequence. This article describes a pyridoxal 5′-phosphate (PLP)–mediated transamination reaction that is specific for the N-terminus of a protein. The reaction oxidizes the N-terminal amine to a ketone or an aldehyde, which can form a stable oxime linkage with an alkoxyamine reagent of choice. Screening studies have identified the most reactive N-terminal residues, facilitating the use of site-directed mutagenesis to achieve high levels of conversion. Additionally, this reaction has been shown to be effective for a number of targets that are not easily accessed through heterologous expression, such as monoclonal antibodies. Curr. Protoc. Chem. Biol. 2:125-134 © 2010 by John Wiley & Sons, Inc.

Keywords:

  • bioconjugation;
  • N-terminus;
  • pyridoxal phosphate;
  • PLP;
  • oximation