Changes in glucose transport and water permeability resulting from the T310I pathogenic mutation in Glut1 are consistent with two transport channels per monomer

Abstract

We studied glucose and water passage across wild type (WT) glucose transporter Glut1 and its T310I pathogenic mutant, expressing them in Xenopus laevis oocytes. We found that the T310I mutation produced a 8-fold decrease in glucose transport (zero-trans influx, 13 ± 2% compared with WT), accompanied by a 2.8-fold increase in the osmotic water permeability (Pf 280 ± 40% compared with WT), and no change in the diffusional water permeability (Pd). The dependence of glucose and water transports on the amounts of mutant cRNA injected was identical exponential buildups (k = 19.7 ng), suggesting that they depend similarly on the quaternary structure. The Ea values for Pf were 16 ± 0.4 (WT) and 11 ± 1 kcal mol-1 (T310I). We report for the first time that 10 mM D-glucose and L-glucose inhibit Pf by ∼45% in the WT but not in the T310I mutant. In addition, 10 mM maltose reduces Pf (15-20%) in both cases. However, 5 mM L-glucose increased the Pf of T310I, consistent with a cooperative effect. These experimental observations and an analysis of our three-dimensional model strongly suggest the presence of two channels per Glut1 monomer, one of which can be blocked by the mutation T310I.