Cryo-EM samples of gas-phase purified protein assemblies using native electrospray ion-beam deposition.
Esser TK., Böhning J., Fremdling P., Bharat T., Gault J., Rauschenbach S.
An increasing number of studies on biomolecular function indirectly combine mass spectrometry (MS) with imaging techniques such as cryo electron microscopy (cryo-EM). This approach allows information on the homogeneity, stoichiometry, shape, and interactions of native protein complexes to be obtained, complementary to high-resolution protein structures. We have recently demonstrated TEM sample preparation via native electrospray ion-beam deposition (ES-IBD) as a direct link between native MS and cryo-EM. This workflow forms a potential new route to the reliable preparation of homogeneous cryo-EM samples and a better understanding of the relation between native solution-phase and native-like gas-phase structures. However, many aspects of the workflow need to be understood and optimized to obtain performance comparable to that of state-of-the-art cryo-EM. Here, we expand on the previous discussion of key factors by probing the effects of substrate type and deposition energy. We present and discuss micrographs from native ES-IBD samples with amorphous carbon, graphene, and graphene oxide, as well as landing energies in the range between 2 and 150 eV per charge.