3D Proteomics

Protein complexes and interactions

We are working on novel methods for identifying and quantifying the interactions and the accurate sites of interaction of proteins with other proteins, DNA and RNA. This makes use of chemical cross-linking to preserve non-covalent interactions for their analysis by mass spectrometry and novel computer algorithms to automatically interpret the mass spectrometric data. We are further developing our methods to address larger complexes, dynamic structures and to couple our results with other technologies such as modelling and cryo-electron microscopy. Ultimately, we aim to analyze chromatin, a large, fragile, insoluble and dynamic protein assembly that is unlikely accessible to structural analysis by current technologies any time soon.

Publications

  • Architecture of the RNA polymerase II-TFIIF complex revealed by cross-linking and mass spectrometry.
    Chen ZA, Jawhari A, Fischer L, Buchen C, Tahir S, Kamenski T, Rasmussen M, Lariviere L, Bukowski-Wills JC, Nilges M, Cramer P, Rappsilber J.
    EMBO J. 2010 Feb 17;29(4):717-26.
    Higher-order multi-protein complexes such as RNA polymerase II (Pol II) complexes with transcription initiation factors are often not amenable to X-ray structure determination. Here, we show that protein cross-linking coupled to mass spectrometry (MS) has now sufficiently advanced as a tool to extend the Pol II structure to a 15-subunit, 670 kDa complex of Pol II with the initiation factor TFIIF at peptide resolution. This work establishes cross-linking/MS as an integrated structure analysis tool for large multi-protein complexes.
  • Implications for kinetochore-microtubule attachment from the structure of an engineered Ndc80 complex.
    Ciferri C, Pasqualato S, Screpanti E, Varetti G, Santaguida S, Dos Reis G, Maiolica A, Polka J, De Luca JG, De Wulf P, Salek M, Rappsilber J, Moores CA, Salmon ED, Musacchio A.
    Cell. 2008 May 2;133(3):427-39.
  • Structural analysis of multi-protein complexes by cross-linking, mass spectrometry and database searching.
    Maiolica A, Cittaro D, Borsotti D, Sennels L, Ciferri C, Tarricone C, Musacchio A, Rappsilber J.
    Mol Cell Proteomics. 2007 Dec;6(12):2200-11.
  • A generic strategy to analyze the spatial organization of multi-protein complexes by cross-linking and mass spectrometry.
    Rappsilber J, Siniossoglou S, Hurt EC, Mann M.
    Anal Chem. 2000 Jan 15;72(2):267-75.