SKM 2023 – wissenschaftliches Programm
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BP: Fachverband Biologische Physik
BP 21: Bioimaging
BP 21.3: Vortrag
Donnerstag, 30. März 2023, 10:00–10:15, BAR Schö
Cryo-EM samples of gas-phase purified protein assemblies using native electrospray ion-beam deposition (ES-IBD) — •Tim Esser1, Lukas Eriksson2, Paul Fremdling2, and Stephan Rauschenbach2 — 1Thermo Fisher Scientific, 1 Boundary Park, Hemel Hempstead, HP2 7GE, UK — 2Department of Chemistry, University of Oxford, Oxford OX1 3TF, UK
Combining native mass spectrometry (MS) with cryo electron microscopy (cryo-EM) allows to correlate information on homogeneity, stoichiometry, shape, and interactions of native protein complexes, complementary to high-resolution protein structures. Cryo-EM samples are conventionally made by coating TEM grids with a protein-containing solution, blotting, and plunge freezing in liquid ethane, quenching proteins in their native state, embedded in ultra-thin films of vitreous ice. Reliable sample preparation remains a major challenge, in particular for heterogeneous samples. Here we demonstrate mass-selective cryo-EM sample preparation via native electrospray ion-beam deposition (ES-IBD), as a direct link between native MS and cryo-EM. Protein complexes are brought into the gas phase, mass-selected, and deposited on TEM grids with thin carbon films at defined temperature. By controlling interactions in solution, gas-phase, and on the surface, we probe protein conformations between native solution-phase and native-like gas-phase structures. We show sub-nanometer EM density maps obtained using native ES-IBD and discuss its potential to extend the scope of cryo-EM in structural biology.