Membrane Protein Insertion Papers
Jens Frauenfeld, James Gumbart, Eli O. van der Sluis, Soledad Funes, Marco Gartmann, Birgitta Beatrix, Thorsten Mielke, Otto Berninghausen, Thomas Becker, Klaus Schulten, and Roland Beckmann. Cryo-EM structure of the ribosome-SecYE complex in the membrane environment. Nature Structural & Molecular Biology, 2011. In press.
The ubiquitous SecY/Sec61-complex acts as protein-conducting channel (PCC) translocating nascent secretory proteins across and integrating membrane proteins into lipid bilayers. This can occur cotranslationally with the ribosome directly bound to the PCC. We present a cryo-electron microscopy structure of the ribosome-bound SecYEG complex in the lipid environment, reconstituted into so-called Nanodiscs. In addition to the canonical ribosome-SecYEG contacts, the presence of a membrane allowed the identification of ribosome-lipid interactions. The rRNA helix 59 (H59) directly contacts the lipid surface and appears to modulate the membrane in immediate vicinity to the proposed lateral gate of the PCC. Combining existing biochemical and structural data with our results, we propose a model of a signal anchor-gated PCC in the membrane.
James Gumbart, Christophe Chipot, and Klaus Schulten. Free-energy cost for translocon-assisted insertion of membrane proteins. Proceedings of the National Academy of Sciences, USA, 2011. In press.
Nascent membrane proteins typically insert in a sequential fashion into the membrane via a protein-conducting channel, the Sec translocon. How this process occurs is still unclear, although a thermodynamic partitioning between the channel and the membrane environment has been proposed. Experiment- and simulation-based scales for the insertion free energy of various amino acids are, however, at variance, the former appearing to lie in a narrower range than the latter. Membrane insertion of arginine, for instance, requires 14-17 kcal/mol according to MD simulations, but only 2-3 kcal/mol according to experiment. We suggest that this disagreement is resolved by assuming a two-stage insertion process wherein the first step, the insertion into the translocon, is energized by protein synthesis and, therefore, has an effectively zero free-energy cost; the second step, the insertion into the membrane, invokes the translocon as an intermediary between the fully hydrated and the fully inserted locations. Using free- energy perturbation calculations, the effective transfer free energies from the translocon to the membrane have been determined for both arginine and leucine amino acids carried by a background poly.leucine helix. Indeed, the insertion penalty for arginine as well as the insertion gain for leucine from the translocon to the membrane are found to be significantly reduced compared to direct insertion from water, resulting in the same compression as observed in the experiment-based scale.
