Cross-linking analysis of protein complexes and structures by tandem mass spectrometry (MS/MS) has advantages in speed, sensitivity, specificity, and the capability of handling complicated protein assemblies. However, detection and accurate assignment of the cross-linked peptides are often challenging due to their low abundance and complicated fragmentation behavior in collision-induced dissociation (CID). To simplify the MS analysis and improve the signal-to-noise ratio of the cross-linked peptides, we developed a novel peptide enrichment strategy that utilizes a cross-linker with a cryptic thiol group and using beads modified with a photocleavable cross-linker. The functional cross-linkers were designed to react with the primary amino groups in proteins. Human serum albumin was used as a model protein to detect intra- and intermolecular cross-linkages. Use of this protein-free selective retrieval method eliminates the contamination that can result from avidin - biotin based retrieval systems and simplifies data analysis. These features may make the method suitable to investigate protein - protein interactions in biological samples.
ASJC Scopus subject areas
- Analytical Chemistry