Improving Identification of Protein-Protein Interactions Using an Affinity-enrichable, Isotopically Coded, and Mass Spectrometry-cleavable Chemical Crosslinker.

Karl A T Makepeace, Yassene Mohammed, Elena L Rudashevskaya, Evgeniy V Petrotchenko, F-Nora Vögtle, Chris Meisinger, Albert Sickmann, Christoph H Borchers

Journal: Molecular & cellular proteomics : MCP 2021;19(4):624-639

PMID: 32051233

Free Full Text on Elsevier Science View on PubMed

Abstract

An experimental and computational approach for identification of protein-protein interactions by chemical crosslinking and mass spectrometry (CLMS) has been developed that takes advantage of the specific characteristics of cyanurbiotindipropionylsuccinimide (CBDPS), an affinity-tagged isotopically coded mass spectrometry (MS)-cleavable crosslinking reagent. Utilizing this reagent in combination with a crosslinker-specific data-dependent acquisition strategy based on MS2 scans, and a software pipeline designed for integrating crosslinker-specific mass spectral information led to demonstrated improvements in the application of the CLMS technique, in terms of the detection, acquisition, and identification of crosslinker-modified peptides. This approach was evaluated on intact yeast mitochondria, and the results showed that hundreds of unique protein-protein interactions could be identified on an organelle proteome-wide scale. Both known and previously unknown protein-protein interactions were identified. These interactions were assessed based on their known sub-compartmental localizations. Additionally, the identified crosslinking distance constraints are in good agreement with existing structural models of protein complexes involved in the mitochondrial electron transport chain.

Address: Department of Biochemistry and Microbiology, University of Victoria, 3800 Finnerty Rd., Victoria, BC V8P 5C2, Canada; University of Victoria - Genome British Columbia Proteomics Centre, #3101-4464 Markham Street, Vancouver Island Technology Park, Victoria, BC V8Z7X8, Canada.; University of Victoria - Genome British Columbia Proteomics Centre, #3101-4464 Markham Street, Vancouver Island Technology Park, Victoria, BC V8Z7X8, Canada; Center for Proteomics and Metabolomics, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, The Netherlands.; Leibniz Institut für Analytische Wissenschaften - ISAS - e.V., Dortmund, Germany.; University of Victoria - Genome British Columbia Proteomics Centre, #3101-4464 Markham Street, Vancouver Island Technology Park, Victoria, BC V8Z7X8, Canada; Segal Cancer Proteomics Centre, Lady Davis Institute, Jewish General Hospital, McGill University, Montreal, Quebec, H3T 1E2, Canada.; Institute of Biochemistry and Molecular Biology, ZBMZ, Faculty of Medicine, University of Freiburg, Freiburg, Germany; Signalling Research Centres BIOSS and CIBSS, University of Freiburg, Germany.; Leibniz Institut für Analytische Wissenschaften - ISAS - e.V., Dortmund, Germany. Electronic address: [email protected].; Department of Biochemistry and Microbiology, University of Victoria, 3800 Finnerty Rd., Victoria, BC V8P 5C2, Canada; University of Victoria - Genome British Columbia Proteomics Centre, #3101-4464 Markham Street, Vancouver Island Technology Park, Victoria, BC V8Z7X8, Canada; Segal Cancer Proteomics Centre, Lady Davis Institute, Jewish General Hospital, McGill University, Montreal, Quebec, H3T 1E2, Canada; Gerald Bronfman Department of Oncology, Jewish General Hospital, Montreal, Quebec, H3T 1E2, Canada; Department of Data Intensive Science and Engineering, Skolkovo Institute of Science and Technology, Skolkovo Innovation Center, Nobel St., Moscow 143026, Russia. Electronic address: [email protected].

DOI: 10.1074/mcp.RA119.001839

Link outs

Free resources

Full Text Sources:

Molecular Biology Databases:

https://www.yeastgenome.org/reference/32051233

MeSH Terms: Biotin, Chemical Fractionation, Cross-Linking Reagents, Isotope Labeling, Mass Spectrometry, Mitochondria, Models, Molecular, Organelles, Peptides, Protein Interaction Mapping, Protein Interaction Maps, Saccharomyces cerevisiae, Succinimides