Advanced In Vivo Cross-Linking Mass Spectrometry Platform to Characterize Proteome-Wide Protein Interactions

Martial Jean-Pierre Rey (1) , Jonathan Dhenin (1) , Youxin Kong (2) , Lucienne Nouchikian (1) , Isaac Filella (3) , Magalie Duchateau (1) , Mathieu Dupré (1) , Riccardo Pellarin (3) , Guillaume Duménil (2) , Julia Chamot-Rooke (1)

1 UTechS MSBio - Spectrométrie de Masse pour la Biologie – Mass Spectrometry for Biology
2 Pathogénèse des Infections vasculaires / Pathogenesis of Vascular Infections
3 Bioinformatique structurale - Structural Bioinformatics

Chemical cross-linking (XL) coupled to mass spectrometry (MS) has become a powerful approach to probe the structure of protein assemblies. Although most of the applications concerned purified complexes, latest developments focus on large-scale in vivo studies. Pushing in this direction, we developed an advanced in vivo cross-linking mass spectrometry platform to study the cellular interactome of living bacterial cells. It is based on in vivo labeling and involves a one-step enrichment by click chemistry on a solid support. Our approach shows an impressive efficiency on Neisseria meningitidis, leading to the identification of about 3300 cross-links for the LC-MS/MS analysis of a biological triplicate using a benchtop high-resolution Orbitrap mass spectrometer. Highly dynamic multiprotein complexes were successfully captured and characterized in all bacterial compartments, showing the great potential and precision of our proteome-wide approach. Our workflow paves new avenues for the large-scale and nonbiased analysis of protein-protein interactions. All raw data, databases, and processing parameters are available on ProteomeXchange via PRIDE repository (data set identifier PXD021553).

Domaines: Biologie structurale [q-bio.BM]

Martial Jean-Pierre Rey, Jonathan Dhenin, Youxin Kong, Lucienne Nouchikian, Isaac Filella, et al.. Advanced In Vivo Cross-Linking Mass Spectrometry Platform to Characterize Proteome-Wide Protein Interactions. Analytical Chemistry, 2021, 93 (9), pp.4166-4174. ⟨10.1021/acs.analchem.0c04430⟩. ⟨hal-03157343⟩