Large-Scale Quantitative Cross-Linking and Mass Spectrometry Provides New Insight on Protein Conformational Plasticity within Organelles, Cells, and Tissues

Abstract

Many proteins can exist in multiple conformational states in vivo to achieve distinct functional roles. These states include alternative conformations, variable PTMs, and association with interacting protein, nucleotide, and ligand partners. Quantitative chemical cross-linking of live cells, organelles, or tissues together with mass spectrometry provides the relative abundance of cross-link levels formed in two or more compared samples, which depends both on the relative levels of existent protein conformational states in the compared samples as well as the relative likelihood of the cross-link originating from each. Because cross-link conformational state preferences can vary widely, one expects intra-protein cross-link levels from proteins with high conformational plasticity to display divergent quantitation among samples with differing conformational ensembles. Here we use the large volume of quantitative cross-linking data available on the public XLinkDB database to cluster intra-protein cross-links according to their quantitation in many diverse compared samples to provide the first widescale glimpse of cross-links grouped according to the protein conformational state(s) from which they predominantly originate. We further demonstrate how cluster cross-links can be aligned with any protein structure to assess the likelihood that they were derived from it.