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

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.

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PNAS-2014-Leitner-9455-60.pdf (Any fulltext), 779KB
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Leitner, Alexander1, Author
Joachimiak, Lukasz A.1, Author
Unverdorben, Pia2, Author              
Walzthoeni, Thomas1, Author
Frydman, Judith1, Author
Förster, Friedrich2, Author              
Aebersold, Ruedi1, Author
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1external, ou_persistent22              
2Förster, Friedrich / Modeling of Protein Complexes, Max Planck Institute of Biochemistry, Max Planck Society, ou_1565148              

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Free keywords: MASS-SPECTROMETRY; 26S PROTEASOME; MOLECULAR ARCHITECTURE; REGULATORY PARTICLE; STRUCTURAL-ANALYSIS; LINKED PEPTIDES; CHAPERONIN; IDENTIFICATION; RESOLUTION; TOPOLOGIES
 Abstract: 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.

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Language(s): eng - English
 Dates: 2014
 Publication Status: Published in print
 Pages: 6
 Publishing info: -
 Table of Contents: -
 Rev. Type: Peer
 Identifiers: ISI: 000338118900039
DOI: 10.1073/pnas.1320298111
 Degree: -

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Title: PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
Source Genre: Journal
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Publ. Info: 2101 CONSTITUTION AVE NW, WASHINGTON, DC 20418 USA : NATL ACAD SCIENCES
Pages: - Volume / Issue: 111 (26) Sequence Number: - Start / End Page: 9455 - 9460 Identifier: ISSN: 0027-8424