Oral Presentation 2014 International Biophysics Congress

High-resolution structural studies of membrane proteins by cryo-electron microscopy: Observing potassium ion channels and bacterial secretion systems in action (#195)

Henning Stahlberg 1
  1. Centre for Cellular Imaging and NanoAnalytics (C-INA), Biozentrum, University of Basel, Basel, Switzerland

Electron microscopy (EM) is a versatile tool for structural studies of biological samples at various resolution scales, ranging from cellular tissue to macromolecular complexes. Electron tomography of frozen hydrated bacteria can reveal insights into the cellular context of membrane protein systems, while high-resolution cryo-EM can reveal the ultrastructure of membrane proteins in the lipid-membrane embedded state, and reach sufficient resolution to determine the atomic resolution structure of the proteins.

This lecture will present structural studies by EM of MloK1, a cyclic-nucleotide modulated potassium channel that features putative voltage sensor domains. The structure of the lipid-membrane reconstituted bacterial channel was determined in the presence and absence of its ligand cAMP, revealing significant movements of the ligand binding domains, which interact with the channel’s putative “voltage sensor domains”, thereby presumably altering the channel at the selectivity filter domain 1

In another study on bacterial secretion systems, electron tomography was used to study the type-III secretion system of Yersinia enterocolitica, revealing the native conformation and surprisingly large structural elasticity of this large multi-protein complex in situ2. In another study, the contractile sheath of the type-VI secretion system of Vibrio cholera was studied by high-resolution cryo-EM and helical image processing, revealing at a resolution of 3.2 Ångstrom the structure of the sheath in its contracted form, which allowed the de novo protein structure of the sheath proteins VipA and VipB to be determined3.

  1. Kowal, J., Chami, M., Baumgartner, P., Arheit, M., Chiu, P.-L., Schröder, G.F., Nimigean, C.M., and Stahlberg, H. (2014) Ligand-induced structural changes in the cyclic nucleotide-modulated potassium channel MloK1. Nature Communications 5:3106
  2. Kudryashev, M., Stenta, M., Schmelz, S., Amstutz, M., Wiesand, U., Castaño-Díez, D., Degiacomi, M., Bleck, C. K. E., Kowal, J., Diepold, A., Heinz, D.W., Dal Peraro, M., Cornelis, G.R., and Stahlberg, H. (2013) In situ structural analysis of the Yersinia enterocolitica injectisome, eLife 2:e00792
  3. Kudryashev, M., Wang, R., Ionescu, M., Brackmann, M., Scherer, S., Maier, T., DiMaio, F., Baker D., Stahlberg, H., Egelman, E., and Basler, M. (2014) The structure of the type six secretion system contractile sheath solved by cryo-electron microscopy (submitted)