Mechanism of metal-specific ECF transporters

• Applicant: Professor Dr. Thomas Eitinger
• Subject Area: Biochemistry ; Biophysics
• Term: from 2016 to 2021
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 315269648

Energy-coupling factor (ECF) transporters represent a large subgroup of the ATP-binding cassette (ABC) transporters and are involved in micronutrient uptake in prokaryotes. Substrates comprise water-soluble vitamins and the transition metal ions Co2+ and Ni2+. The common composition of all ECF transporters consisting of a substrate-binding integral membrane protein (S) and the ECF, composed of another integral membrane-protein (T) and two ABC ATPases, was originally described for metal-specific systems in 2006. In the following years, vitamin-specific ECF transporters were in the focus of experimental research. Recent results include the finding that S units for different substrates with negligible sequence similarity have very similar 3D topologies, and the conclusion/hypothesis that toppling over of S units within the membrane is required for substrate translocation across the lipid bilayer. The first 3D structure of a metal-specific S unit revealed a very similar topology compared to the vitamin-specific counterparts. Previous findings indicating a specific role for the additional N-terminal transmembrane helix in metal-specific S units were confirmed. The very N-terminal end extends deeply into the substrate-binding pocket, blocks access of larger molecules and provides three of the four metal ligands of the square planar ligand field. Questions of (i) how the N-terminus is positioned correctly during metal binding, (ii) how it is removed from the binding pocket for metal release into the cytoplasm, and (iii) how these processes are controlled by a helper protein, that has an essential but uncharacterized role and occurs only in metal-specific ECF transporters, are completely open. Those questions are within the focus of the proposal. Based on extensive preliminary work, we aim at elucidating the mechanism of metal binding and metal release and the underlying conformational changes of the S unit and the helper protein using spectroscopical and biochemical techniques. This research is of general interest for an understanding of the mechanism of ECF transporters. The distinct similarity among S, T and ATPase components of metal- and vitamin-specific ECF transporters suggests a basically similar transport mechanism but significant differences in the substrate-loading and -release steps.

• DFG Programme: Research Grants