Synthesis of multi-domain proteins using Expressed Protein Ligation: Strategies for segmental isotopic labeling of internal regions

Ulrich K. Blaschke, Graham J. Cotton, Tom W. Muir

Research output: Contribution to journalArticlepeer-review

24 Scopus citations

Abstract

Here we describe how a sequential version of the protein semi-synthesis technique, Expressed Protein Ligation (EPL), can be used to assemble multiple (i.e. 3 or more) recombinantly-derived polypeptides segments into a target protein. Sequential EPL was successfully used to assembly the 304 amino acid eukaryotic adaptor protein, Crk-II, from three recombinant polypeptide segments in good yield. Moreover, the resulting multi-component ligation product was found to possess the expected biological activity in a series of ligand binding studies. By allowing the controlled assembly of 3 or more recombinant polypeptide segments, sequential EPL opens the door to the segmental isotopic labeling of internal regions of large proteins with NMR probe-nuclei. (C) 2000 Elsevier Science Ltd.

Original languageEnglish (US)
Pages (from-to)9461-9470
Number of pages10
JournalTetrahedron
Volume56
Issue number48
DOIs
StatePublished - Nov 24 2000
Externally publishedYes

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Drug Discovery
  • Organic Chemistry

Keywords

  • Expressed protein ligation
  • Isotopic labeling
  • Semi-synthesis technique

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