Dimeric E. coli YidC forms a translocation pore in the membrane
Sprache des Vortragstitels:
Lukas Winter, Andreas Vogt, Christine Siligan, Denis Knyazev, Roland Kuttner, Hans-Georg Koch and Peter Pohl
The universally conserved protein YidC functions as a membrane protein insertase. It (i) facilitates transmembrane domains? passage out of SecYEG?s lateral gate and (ii) inserts short or closely spaced membrane proteins independent of SecYEG. According to a recent cryo-EM density map of a ribosome nascent chain complex bound to YidC in detergent in addition to X-ray structures of YidC in a monoolein membrane, monomeric YidC assists in membrane protein insertion by offering a hydrophilic groove that surrounds a charged amino acid located at the membrane midplane. These structures raise the question: how is the membrane barrier to protons maintained? We now show that purified E. coli YidC forms an aqueous pore upon reconstitution. The pore is closed to ions in the resting state and opens upon binding of ribosome nascent chain complexes. The observation indicates that YidC is able to preserve the membrane barrier in its idle state. As monomeric YidC cannot form a gated ion channel, we exploited fluorescence correlation spectroscopy to count the number of particles upon dissolution of proteoliposomes by a mild detergent and subsequent exposure of the resulting micelles to a harsh detergent1. The number of particles doubled, indicating that YidC was reconstituted as a dimer. To confirm dimer formation and the causal connection between ribosome binding and channel activity, we monitored single-molecule FRET between labeled ribosomes and labeled YidC. Unitary channel conductivity was decreased as compared to channels activated by the ribosome nascent chain complex, suggesting that the nascent chain is located at the interface between the two pore-forming monomers and the lipid. Thus, our data suggest that YidC forms an ion-conducting pore in the membrane, which is opened by ribosome binding similar to SecYEG2.