Nutrient acquisition in microbial pathogens: riding the elevator

Sialic acid is a sugar molecule which plays an essential role in infections by human pathogens such as Haemophilus influenzae and Vibrio cholerae. Because both bacteria are not able to de novo synthesize sialic acid, they rely on the uptake of this molecule from their environment. This is accomplished by secondary active transporters from the group of tripartite ATP-independent periplasmic (TRAP) transporters. TRAP transporters consist of three different domains, a small and a large membrane domain and a periplasmic substrate-binding protein. While the structure and function of the substrate-binding protein is well-characterized, the membrane domains are not well studied. We used cryo-electron microscopy to solve the first structure of a TRAP transporter membrane domains in lipid nanodiscs. The structure revealed surprising and unexpected similarities of TRAP transporters to elevator-like secondary transporters. Using a combined approach of AlphaFold modelling, in vivo activity assays and single molecule fluorescence microscopy, we postulated a combined mechanism of the elevator movement of the membrane domains with the substrate-binding and tripartite complex formation of the substrate-binding protein. In addition we showed that specific-nanobodies, which were used as molecular marker to identify the TRAP transporter on the EM-micrographs, also have an inhibiting effect on the transporters function.