Streamlining Cross-Linking Mass Spectrometry Analysis
- Author(s): Huszagh, Alex Scott
- Advisor(s): Huang, Lan
- et al.
Cross-linking mass spectrometry maps the structural topology of protein complexes by using
the distance between linked residues as spatial constraints, complementing other structural
biology techniques. However, the identification of cross-linked peptides scales poorly with the
number of proteins analyzed. Our lab has previously developed MS-cleavable cross-linkers to
enable the separation of cross-linked peptides prior to sequencing, enabling peptide identifica-
tion using standard peptide search databases. We describe the design and implementation of
platform and application named XLTools for the automated identification of MS-cleavable
cross-linked peptides. XLTools supports open and proprietary data formats and common
peptide search databases, facilitating its integration into existing workflows. Furthermore,
we developed peak-picking and validation algorithms to enable the accurate quantitation of
cross-linked peptides in complex samples. We demonstrate the application of XLTools to
the quantitative analysis of the 26S proteasome cross-linked in vivo and in vitro.