On the cover of the EMBO journal! ''Dynamic coupling in the bacterial Sec translocon."
Faculty scientist Dr. Tomas Fessl led a research project that demonstrated the key role of fast protein dynamics in the mechanism of protein transport across membranes. The results, which were published in the January issue of the EMBO journal, were obtained in collaboration with teams from University of Bristol (Prof. Ian Collinson) and University of Leeds (Prof. Sheena Radford, FRS) and involved local colleagues Dr. Joel Crossley and Prof. Roman Tuma.
The machinery behind the protein transport investigated in this study is the ubiquitous Sec translocon, a highly conserved membrane assembly for polypeptide transport across, or into, lipid bilayers. In bacteria, secretion through the core channel complex—SecYEG in the inner membrane—is powered by the cytosolic ATPase SecA. While individual parts of the molecular machinery are well known, the mechanism by which SecYEG adapts to and performs these various tasks remains unresolved. Prior to this publication the prevailing view favoured direct coupling between the ATPase and translocation, the so-called power stroke mechanism.
In collaboration with the groups of Prof. Ian Collinson (Bristol, United Kingdom) and Prof. Sheena Radford (Leeds, United Kingdom), local researchers developed a state-of-the-art single-molecule essay for quantitative detection and analyses of Sec translocon dynamics. Investigation of the translocon dynamics in different nucleotide states of SecA revealed that SecYEG channel is subject to rapid opening and closing on a much faster time scale than that of the SecA ATPase turnover. Such dynamic coupling of SecA ATPase to the SecYEG channel opening is incompatible with the directly coupled, power stroke models and instead favours a Brownian-ratchet mechanism of translocation which relies on pre-protein diffusion through the pore.
The research team further exploited the well characterised mutant of SecY (PrlA4) to test the Brownian-ratchet mechanism. This variant exhibits faster transport while its ATPase rate remains the same, an observation compatible only with the Brownian-ratchet mechanism. Furthermore, the mutation increases the time the channel spends in the open state, facilitating pre-protein diffusion through the pore and thereby enhancing translocation efficiency. This finding links SecYEG channel dynamics with its biological function, i.e. protein transport.
Such a dynamic allosteric mechanism is not unique to the SecYEG translocon, since similar nucleotide-dependent modulation of rapid dynamics has been recently shown for adenylate kinase, and the AAA+ ring ATPase, ClpB. However, SecYEG is the first membrane complex for which dynamic allostery was demonstrated.
The local group was funded by the European Regional Development Fund-Project (CZ.02.1.01/0.0/0.0/15_003/0000441) and the Czech Science Foundation (20-11563Y) and collaborative funding from BBSRC (BB/T008059/1). Additional funds for collaborating partners were provided by BBSRC (BB/V001531/1 and BB/T006889/1) and Royal Society Professorial Research Fellowship (RSRP\R1\211057).
Contact: Mgr. Tomáš Fessl, Ph.D. (fessl at prf.jcu.cz)
Picture: Illustration of Sec translocon transporting preprotein across inner bacterial membrane
