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

• Published in: Molecular & cellular proteomics Vol. 15; no. 9; pp. 3071 - 3080
• Main Authors: Sarpe, Vladimir, Rafiei, Atefeh, Hepburn, Morgan, Ostan, Nicholas, Schryvers, Anthony B., Schriemer, David C.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.09.2016; The American Society for Biochemistry and Molecular Biology
• Subjects: Actinobacillus pleuropneumoniae; Actinobacillus pleuropneumoniae - chemistry; Actinobacillus pleuropneumoniae - genetics; Actinobacillus pleuropneumoniae - metabolism; Animals; Bacterial Proteins - chemistry; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Cross-Linking Reagents - chemistry; Models, Molecular; Mutation; Peptides - analysis; Peptides - chemistry; Protein Binding; Protein Conformation; Software; Swine; Tandem Mass Spectrometry; Technological Innovation and Resources; Transferrin - chemistry; Transferrin - metabolism; Transferrin-Binding Protein B - chemistry; Transferrin-Binding Protein B - genetics; Transferrin-Binding Protein B - metabolism
• ISSN: 1535-9476; 1535-9484
• DOI: 10.1074/mcp.O116.058685

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.