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Proc Natl Acad Sci U S A. 2017 Feb 14;114(7):1732-1737. doi: 10.1073/pnas.1617220114. Epub 2017 Jan 27.

Mitochondrial protein interactome elucidated by chemical cross-linking mass spectrometry.

Author information

1
Department of Genome Sciences, University of Washington, Seattle, WA 98105.
2
Department of Bioengineering, University of Washington, Seattle, WA 98105.
3
Department of Anesthesiology and Pain Medicine, University of Washington, Seattle, WA 98105.
4
Mitochondria and Metabolism Center, University of Washington, Seattle WA 98105.
5
Department of Radiology, University of Washington, Seattle, WA 98105.
6
Department of Pathology Yale School of Medicine, New Haven, CT 06510.
7
Department of Genetics, Yale School of Medicine, New Haven, CT 06510.
8
Department of Genome Sciences, University of Washington, Seattle, WA 98105; jimbruce@uw.edu.

Abstract

Mitochondrial protein interactions and complexes facilitate mitochondrial function. These complexes range from simple dimers to the respirasome supercomplex consisting of oxidative phosphorylation complexes I, III, and IV. To improve understanding of mitochondrial function, we used chemical cross-linking mass spectrometry to identify 2,427 cross-linked peptide pairs from 327 mitochondrial proteins in whole, respiring murine mitochondria. In situ interactions were observed in proteins throughout the electron transport chain membrane complexes, ATP synthase, and the mitochondrial contact site and cristae organizing system (MICOS) complex. Cross-linked sites showed excellent agreement with empirical protein structures and delivered complementary constraints for in silico protein docking. These data established direct physical evidence of the assembly of the complex I-III respirasome and enabled prediction of in situ interfacial regions of the complexes. Finally, we established a database and tools to harness the cross-linked interactions we observed as molecular probes, allowing quantification of conformation-dependent protein interfaces and dynamic protein complex assembly.

KEYWORDS:

cross-linking; interactome; mass spectrometry; mitochondria; protein interaction reporter

PMID:
28130547
PMCID:
PMC5321032
DOI:
10.1073/pnas.1617220114
[Indexed for MEDLINE]
Free PMC Article

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