Energy landscape steering mediates dynamic coupling in ATP-driven protein translocation by the bacterial Sec machinery

Abstract

The Sec translocon is a highly conserved membrane complex for transport of polypeptides across, or into, lipid bilayers. In bacteria, the core protein-channel SecYEG resides in the inner-membrane, through which secretion is powered by the cytosolic ATPase SecA. Here, we use single-molecule fluorescence to interrogate the dynamic state of SecYEG throughout the hydrolytic cycle of SecA. We show that the SecYEG channel fluctuates between open and closed states faster ([~]20-fold during transport) than ATP turnover; while the nucleotide status of SecA modulates the rates of opening and closure. Interestingly, a SecY variant (PrlA4), exhibiting faster protein transport, but unaffected ATPase rates, increases the dwell time in the open state, facilitating pre-protein diffusion through the pore; thereby improving the efficiency of translocation. Thus, contrary to prevailing structure-based models, SecYEG plays an integral part in the translocation mechanism through dynamic allosteric coupling in which SecA steers the energy landscape of the protein-channel.