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

Autor: Cigler M; Center for Integrated Protein Science Munich (CIPSM), Department of Chemistry, Group of Synthetic Biochemistry, Technical University of Munich, Institute for Advanced Study, Lichtenbergstr. 4, 85748, Garching, Germany., Müller TG; Center for Integrated Protein Science Munich (CIPSM), Department of Chemistry, Group of Synthetic Biochemistry, Technical University of Munich, Institute for Advanced Study, Lichtenbergstr. 4, 85748, Garching, Germany., Horn-Ghetko D; Center for Integrated Protein Science Munich (CIPSM), Department of Chemistry, Group of Synthetic Biochemistry, Technical University of Munich, Institute for Advanced Study, Lichtenbergstr. 4, 85748, Garching, Germany., von Wrisberg MK; Center for Integrated Protein Science Munich (CIPSM), Department of Chemistry, Group of Synthetic Biochemistry, Technical University of Munich, Institute for Advanced Study, Lichtenbergstr. 4, 85748, Garching, Germany., Fottner M; Center for Integrated Protein Science Munich (CIPSM), Department of Chemistry, Group of Synthetic Biochemistry, Technical University of Munich, Institute for Advanced Study, Lichtenbergstr. 4, 85748, Garching, Germany., Goody RS; Department of Structural Biochemistry, MPI of Molecular Physiology, Otto-Hahn Srasse 11, 442277, Dortmund, Germany., Itzen A; Center for Integrated Protein Science Munich (CIPSM), Department of Chemistry, Group of Protein Biochemistry, Technical University of Munich, Lichtenbergstr. 4, 85748, Garching, Germany., Müller MP; Faculty of Chemistry and Chemical Biology, Technical University Dortmund, Otto-Hahn Str. 4a, 44227, Dortmund, Germany., Lang K; Center for Integrated Protein Science Munich (CIPSM), Department of Chemistry, Group of Synthetic Biochemistry, Technical University of Munich, Institute for Advanced Study, Lichtenbergstr. 4, 85748, Garching, Germany.
Jazyk: angličtina
Zdroj: Angewandte Chemie (International ed. in English) [Angew Chem Int Ed Engl] 2017 Dec 04; Vol. 56 (49), pp. 15737-15741. Date of Electronic Publication: 2017 Nov 09.
DOI: 10.1002/anie.201706927
Abstrakt: 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.
(© 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)
Databáze: MEDLINE
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