Translocase mediated biogenesis and assembly of the F0-sector of the F1F0-ATPase
09 / 2003 - 08 / 2007
Nederlandse Organisatie voor Wetenschappelijk Onderzoek (NWO)
About 30% of all cellular proteins encoded by the genome are integral membrane proteins. To fulfill their biological function, many of these proteins assemble into multi-subunit complexes together with other integral and/or peripheral membrane proteins. In bacteria, most nascent membrane proteins are targeted to the membrane by the signal recognition particle, and subsequently membrane-inserted by the translocase. Translocase is a large membrane-embedded protein complex that consists of a molecular motor (SecA) and seven membrane-integrated subunits (SecYEGDFyajC-YidC). Translocase mediates both the translocation of proteins across the membrane and the integration of membrane proteins into the lipid bilayer. An unresolved question is how membrane proteins assemble into large multisubunit complexes and how translocase is involved in this process. The F1F0-ATPase is a well-known example of a membrane protein complex. In Escherichia coli, the membrane-integrated F0-domain consists of a ring of ten to twelve c-subunits, and one a- and two b-subunits. Although the ring can self-assemble from the purified c-subunits, recent in vivo evidence indicates that the functional assembly of the F0-domain is extremely dependent on YidC. YidC is a newly identified integral membrane protein that may function as a membrane protein assembly factor or integrase. In this project, we will use an in vitro approach to analyze the membrane insertion of the E. coli F0-subunits and their assembly into a functional proton-conducting channel. These studies will provide detailed insight into how insertion and assembly are linked, the order of subunit assembly and the role of YidC and other translocase components in this process. The results will deepen our understanding of the mechanism of membrane protein biogenesis.