Unit

UNIT 15.3 Site-Specific Protein Labeling via Sortase-Mediated Transpeptidation

  1. Maximilian Wei-Lin Popp1,2,
  2. John M. Antos1,
  3. Hidde L. Ploegh1,2

Published Online: 1 APR 2009

DOI: 10.1002/0471140864.ps1503s56

Current Protocols in Protein Science

Current Protocols in Protein Science

How to Cite

Popp, M. W.-L., Antos, J. M. and Ploegh, H. L. 2009. Site-Specific Protein Labeling via Sortase-Mediated Transpeptidation. Current Protocols in Protein Science. 56:15.3:15.3.1–15.3.9.

Author Information

  1. 1

    Whitehead Institute for Biomedical Research, Cambridge, Massachusetts

  2. 2

    Department of Biology, Massachusetts Institute of Technology, Cambridge, Massachusetts

Publication History

  1. Published Online: 1 APR 2009
  2. Published Print: APR 2009

Abstract

Creation of functional protein bioconjugates demands methods for attaching a diverse array of probes to target proteins with high specificity, under mild conditions. The sortase A transpeptidase enzyme from Staphylococcus aureus catalyzes the cleavage of a short 5-aa recognition sequence (LPXTG) with the concomitant formation of an amide linkage between an oligoglycine peptide and the target protein. By functionalizing the oligoglycine peptide, it is possible to incorporate reporters into target proteins in a site-specific fashion. This reaction is applicable to proteins in solution and on the living cell surface. The method described in this unit only requires incubation of the target protein, which has been engineered to contain a sortase recognition site either at the C terminus or within solvent-accessible loops, with a purified sortase enzyme and a suitably functionalized oligoglycine peptide. Curr. Protoc. Protein Sci. 56:15.3.1-15.3.9. © 2009 by John Wiley & Sons, Inc.

Keywords:

  • sortase;
  • transpeptidation;
  • site-specific labeling;
  • chemoenzymatic labeling