A semisynthetic Eph receptor tyrosine kinase provides insight into ligand-induced kinase activation

Nikhil Singla, Hediye Erdjument-Bromage, Juha P. Himanen, Tom W. Muir, Dimitar B. Nikolov

Research output: Contribution to journalArticle

23 Scopus citations

Abstract

We have developed a methodology for generating milligram amounts of functional Eph tyrosine kinase receptor using the protein engineering approach of expressed protein ligation. Stimulation with ligand induces efficient autophosphorylation of the semisynthetic Eph construct. The in vitro phosphorylation of key Eph tyrosine residues upon ligand-induced activation was monitored via time-resolved, quantitative phosphoproteomics, suggesting a precise and unique order of phosphorylation of the Eph tyrosines in the kinase activation process. To our knowledge, this work represents the first reported semisynthesis of a receptor tyrosine kinase and provides a potentially general method for producing single-pass membrane proteins for structural and biochemical characterization.

Original languageEnglish (US)
Pages (from-to)361-371
Number of pages11
JournalChemistry and Biology
Volume18
Issue number3
DOIs
StatePublished - Mar 25 2011

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Medicine
  • Molecular Biology
  • Pharmacology
  • Drug Discovery
  • Clinical Biochemistry

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