P4 Structure and function of the betaine transporter BetP from Corynebacterium glutamicum

The Structure and function of the osmoregulated betaine transporter BetP from Corynebacterium glutamicum and functional relevant mutants were investigated by crystallography and betaine uptake measurements in cells and proteoliposomes. The X-ray structure of the fully osmoregulated BetP trimer has been determined to 3.3 Å. BetP has a symmetrical substrate-occluded conformation with a betaine bound in an occluded tryptophan pocket in each monomer. Aromatic side chains lining the transport pathway contribute to the exquisite substrate specificity of BetP. The overall fold of two structurally related inverted repeats and partially unwound helices is striking similar to other unrelated Na⁺-coupled transporters. The unique conformation of BetP allows a describition of the conformational changes during inward transport of ions and substrate. A 8Å 3D map of the deregulated, constitutively active transporter has been determined from two-dimensional crystals by electron cryo-microscopy revealing that BetP is an asymmetrical homotrimer of three different conformations in the membrane. The X-ray structure was fitted into the EM density and the three conformations were assigned according to the alternated access model as an outward facing and an inward facing conformation and an occluded state. Based on these structural data we aim to understand how BetP detects and responds to osmotic stress. In future, we will investigate the molecular details of sodium-coupled osmolyte transport and osmoregulation by electron cryo-microscopy and X-ray crystallography.

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