Hydrogen/deuterium exchange: moving from easy proteins to real life stories

Petr Man^1,2, Alan Kádek^1,2, Jiří Hausner^1,2, Daniel Kavan^1,2, Hynek Mrázek¹, Martin Stráňava², Markéta Martínková², Filip Trčka³, Michal Ďurech³, Petr Müller³, Martial Rey⁴, Petr Pompach^1,2, Petr Novák^1,2

¹Institute of Microbiology CAS, Vídeňská 1083, Prague

²Faculty of Science, Charles University in Prague, Hlavova 8, Prague

³ Masaryk Memorial Cancer Institute, Regional Centre for Applied Molecular Oncology, Žlutý kopec 7, Brno

⁴ Institut Pasteur, 28 rue du Dr. Roux, Paris

pman@biomed.cas.cz

H/D exchange coupled to mass spectrometry is already a well-established technique of structural biology. It allows monitoring of protein interactions and structure changes via time resolved kinetics of amide hydrogen exchange. It is often presented as a technique with virtually no limitations in terms of protein size, complexity, or buffer requirements.  In this presentation we will critically review these statements by demonstrating methodological developments in our laboratory and their application to several non-trivial protein problems including monitoring of a large number of unusual experimental conditions, big modified proteins, membrane proteins and complex protein mixtures.

This work was funded by Czech Science Foundation (P206/12/0503); European Regional Development Funds (CZ.1.07/2.3.00/30.0003 and CZ.1.05/1.1.00/02.0109), OPPK project CZ.2.16/3.1.00/24023; Institutional Research Concept of the Institute of Microbiology RVO61388971; Research Project of Charles University (UNCE 204025/2012).