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Chemical cross-linking/mass spectrometry targeting acidic residues in proteins and protein complexes

MPG-Autoren
http://pubman.mpdl.mpg.de/cone/persons/resource/persons78814

Unverdorben,  Pia
Förster, Friedrich / Modeling of Protein Complexes, Max Planck Institute of Biochemistry, Max Planck Society;

http://pubman.mpdl.mpg.de/cone/persons/resource/persons77965

Förster,  Friedrich
Förster, Friedrich / Modeling of Protein Complexes, Max Planck Institute of Biochemistry, Max Planck Society;

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Volltexte (frei zugänglich)

PNAS-2014-Leitner-9455-60.pdf
(beliebiger Volltext), 779KB

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Zitation

Leitner, A., Joachimiak, L. A., Unverdorben, P., Walzthoeni, T., Frydman, J., Förster, F., et al. (2014). Chemical cross-linking/mass spectrometry targeting acidic residues in proteins and protein complexes. PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 111(26), 9455-9460. doi:10.1073/pnas.1320298111.


Zitierlink: http://hdl.handle.net/11858/00-001M-0000-001A-1B73-6
Zusammenfassung
The study of proteins and protein complexes using chemical cross-linking followed by the MS identification of the cross-linked peptides has found increasingly widespread use in recent years. Thus far, such analyses have used almost exclusively homobifunctional, amine-reactive cross-linking reagents. Here we report the development and application of an orthogonal cross-linking chemistry specific for carboxyl groups. Chemical cross-linking of acidic residues is achieved using homobifunctional dihydrazides as cross-linking reagents and a coupling chemistry at neutral pH that is compatible with the structural integrity of most protein complexes. In addition to cross-links formed through insertion of the dihydrazides with different spacer lengths, zero-length cross-link products are also obtained, thereby providing additional structural information. We demonstrate the application of the reaction and the MS identification of the resulting cross-linked peptides for the chaperonin TRiC/CCT and the 26S proteasome. The results indicate that the targeting of acidic residues for cross-linking provides distance restraints that are complementary and orthogonal to those obtained from lysine cross-linking, thereby expanding the yield of structural information that can be obtained from cross-linking studies and used in hybrid modeling approaches.