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Mol Cell Proteomics. 2016 Sep;15(9):3071-80. doi: 10.1074/mcp.O116.058685. Epub 2016 Jul 13.

High Sensitivity Crosslink Detection Coupled With Integrative Structure Modeling in the Mass Spec Studio.

Author information

1
From the ‡Department of Biochemistry and Molecular Biology.
2
§Department of Chemistry.
3
¶Department of Microbiology, Immunology and Infectious Diseases, University of Calgary, Alberta, T2N 1N4, Canada.
4
From the ‡Department of Biochemistry and Molecular Biology, ¶Department of Microbiology, Immunology and Infectious Diseases, University of Calgary, Alberta, T2N 1N4, Canada.
5
From the ‡Department of Biochemistry and Molecular Biology, §Department of Chemistry, dschriem@ucalgary.ca.

Abstract

The Mass Spec Studio package was designed to support the extraction of hydrogen-deuterium exchange and covalent labeling data for a range of mass spectrometry (MS)-based workflows, to integrate with restraint-driven protein modeling activities. In this report, we present an extension of the underlying Studio framework and provide a plug-in for crosslink (XL) detection. To accommodate flexibility in XL methods and applications, while maintaining efficient data processing, the plug-in employs a peptide library reduction strategy via a presearch of the tandem-MS data. We demonstrate that prescoring linear unmodified peptide tags using a probabilistic approach substantially reduces search space by requiring both crosslinked peptides to generate sparse data attributable to their linear forms. The method demonstrates highly sensitive crosslink peptide identification with a low false positive rate. Integration with a Haddock plug-in provides a resource that can combine multiple sources of data for protein modeling activities. We generated a structural model of porcine transferrin bound to TbpB, a membrane-bound receptor essential for iron acquisition in Actinobacillus pleuropneumoniae Using mutational data and crosslinking restraints, we confirm the mechanism by which TbpB recognizes the iron-loaded form of transferrin, and note the requirement for disparate sources of restraint data for accurate model construction. The software plugin is freely available at www.msstudio.ca.

PMID:
27412762
PMCID:
PMC5013318
DOI:
10.1074/mcp.O116.058685
[Indexed for MEDLINE]
Free PMC Article

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