Site-specific N-terminal labeling of proteins using sortase-mediated reactions

Christopher S. Theile, Martin D. Witte, Annet E. M. Blom, Lenka Kundrat, Hidde L. Ploegh*, Carla P. Guimaraes*

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review

125 Citations (Scopus)

Abstract

This protocol describes the use of sortase-mediated reactions to label the N terminus of any given protein of interest. The sortase recognition sequence, LPXTG (for Streptococcus aureus sortase A) or LPXTA (for Staphylococcus pyogenes sortase A), can be appended to a variety of probes such as fluorophores, biotin or even to other proteins. The protein to be labeled acts as a nucleophile by attacking the intermediate formed between the probe containing the LPXTG/A motif and the sortase enzyme. If sortase, the protein of interest and a suitably functionalized label are available, the reactions usually require less than 3 h.

Original languageEnglish
Pages (from-to)1800-1807
Number of pages8
JournalNature protocols
Volume8
Issue number9
DOIs
Publication statusPublished - Sep-2013
Externally publishedYes

Keywords

  • GRAM-POSITIVE BACTERIA
  • SURFACE-PROTEINS
  • SOLID-PHASE
  • STAPHYLOCOCCUS-AUREUS
  • LIVING CELLS
  • PEPTIDE
  • DOMAIN
  • IDENTIFICATION
  • CONJUGATION
  • MALEIMIDE

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