Biophysical characteristics of the pig kidney Na+/glucose cotransporter SGLT2 reveal a common mechanism for SGLT1 and SGLT2

Abstract

The Na+-dependent, low affinity glucose transporter SGLT2 cloned from pig kidney is 76% identical (at the amino acid level) to its high affinity homologue SGLT1. Using two-microelectrode voltage clamp, we have characterized the presteady-state and steady-state kinetics of SGLT2 expressed in Xenopus oocytes. The kinetic properties of the steady-state sugar-evoked currents as a function of external Na+ and α-methyl-D- glucopyranoside (αMG) concentrations were consistent with an ordered, simultaneous transport model in which Na+ binds first. Na+ binding was voltage-dependent and saturated with hyperpolarizing voltages. Phlorizin was a potent inhibitor of the sugar-evoked currents (K(i)/(Pz) ≃ 10 μM) and blocked an inward Na+ current in the absence of sugar. SGLT2 exhibited Na+- dependent presteady-state currents with time constants 3-7 ms. Charge movements were described by Boltzmann relations with apparent valence ≃ 1 and maximal charge transfer ≃ 11 nC, and were reduced by the addition of sugar or phlorizin. The differences between SGLT1 and SGLT2 were that (i) the apparent affinity constant (K(0.5)) for αMG (≃3 mM) was an order of magnitude higher for SGLT2; (ii) SGLT2 excluded galactose, suggesting discrete sugar binding; (iii) K(0.5) for Na+ was lower in SGLT2; and (iv) the Hill coefficient for Na+ was 1 for SGLT2 but 2 for SGLT1. Simulations of the six-state kinetic model previously proposed for SGLT1 indicated that many of the kinetic properties observed in SGLT2 are expected by simply reducing the Na+/glucose coupling from 2 to 1.