Proximity-Triggered Covalent Stabilization of Low-Affinity Protein Complexes In Vitro and In Vivo

Marko Cigler, Thorsten G. Müller, Daniel Horn-Ghetko, Marie Kristin von Wrisberg, Maximilian Fottner, Roger S. Goody, Aymelt Itzen, Matthias P. Müller, Kathrin Lang

Research output: Contribution to journalArticlepeer-review

50 Scopus citations


The characterization of low-affinity protein complexes is challenging due to their dynamic nature. Here, we present a method to stabilize transient protein complexes in vivo by generating a covalent and conformationally flexible bridge between the interaction partners. A highly active pyrrolysyl tRNA synthetase mutant directs the incorporation of unnatural amino acids bearing bromoalkyl moieties (BrCnK) into proteins. We demonstrate for the first time that low-affinity protein complexes between BrCnK-containing proteins and their binding partners can be stabilized in vivo in bacterial and mammalian cells. Using this approach, we determined the crystal structure of a transient GDP-bound complex between a small G-protein and its nucleotide exchange factor. We envision that this approach will prove valuable as a general tool for validating and characterizing protein–protein interactions in vitro and in vivo.

Original languageEnglish
Pages (from-to)15737-15741
Number of pages5
JournalAngewandte Chemie International Edition in English
Issue number49
StatePublished - 4 Dec 2017


  • covalent stabilization
  • protein crosslinking
  • protein-protein interactions
  • proximity effects
  • unnatural amino acids


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