Energetics of polypeptide partitioning from the translocon into the lipid bilayer
Sprache des Vortragstitels:
3. 19th IUPAB Congress and 11th EBSA Congress
Sprache des Tagungstitel:
The heterotrimeric bacterial translocon SecYEG facilitates the secretion of hydrophilic proteins across the cytoplasmic membrane and the insertion of hydrophobic polypeptides into the membrane. Mounting evidence suggests that the energetic costs for protein partitioning into the membrane from (i) the aqueous lumen of SecYEG and (ii) the aqueous bulk solution are different. Here we tested the hypothesis that the restricted residence time of the nascent chain in the channel is responsible. Therefore we first reconstituted SecYEG into planar lipid bilayers and stalled translocation intermediates of different hydrophobicities in the channel. We prevented back-sliding of the transmembrane segments out of the translocon by sandwiching them between the ribosome on one side of the membrane and calmodulin on the other. In the second step we forced SecYEG closure by applying a membrane potential1. Hydrophobic transmembrane helices easily exited the lateral gate as indicated by zero residual ion conductivity, whereas hydrophilic ones remained in the region of the lateral gate or in SecYEG?s lumen giving rise to ion conductivity. This approach allows assaying the cost for membrane partitioning at thermodynamic equilibrium.