Mechanism of Water and Solute Cotransport by the Sodium Glucose Cotransporter SGLT1
Sprache des Vortragstitels:
The molecular mechanism of carrier mediated water and solute cotransport is unresolved. It has long been believed that water may be engulfed into proteinaceous cavities and thus be moved along with the solute in a fixed stoichiometric ratio from one side of the membrane to the other. Since solute and solvent flux measurements through the potassium-chloride-cotransporter or the glucose sodium cotransporter (SGLT1) in epithelial monolayers and/or reconstituted lipid bilayers did not confirm stoichiometric coupling, we tested the hypothesis that water and glucose share part of their pathway through SGLT1. To this end we reconstituted the purified SGLT1 into lipid vesicles and exposed the proteoliposomes to osmotic gradients of different origin. SGLT1 facilitates passive water flow. Its unitary water permeability pf was higher in the presence of sucrose than in the presence of glucose indicating a reflection coefficient ? of glucose smaller than one. The decrease pf was due to occlusion of the water pathway by bound glucose molecules. Mutation of the glucose binding pocket released the block. The glucose concentration at which pf dropped to 50 % of its maximal value characterizes the glucose affinity of SGLT1's inwardly open conformation. It was two orders of magnitude larger than the affinity in the outwardly open conformation, i.e. the previously reported concentration at which SGLT1 achieves half of its maximum rate. Thus glucose transport requires conformational transitions while water is channeled as long as the glucose binding site is unoccupied - explaining as to why both substrates are transported at despairingly different rates.