Biophysical Characteristics of the Pig Kidney Na+/Glucose Cotransporter SGLT2 Reveal a Common Mechanism for SGLT1 and SGLT2

• Published in: The Journal of biological chemistry 1996-12, Vol.271 (51), p.32678-32683
• Main Authors: Mackenzie, Bryan, Loo, Donald D.F., Panayotova-Heiermann, Mariana, Wright, Ernest M.
• Format: Article
• Language: English
• Subjects: AMPHIBIEN; ANFIBIOS; Animals; Biological Transport; Biophysical Phenomena; Biophysics; CERDO; Electric Conductivity; ELECTRICIDAD; ELECTRICITE; Electrophysiology; ESTRUCTURA CELULAR; GLUCOSA; GLUCOSE; Glucose - metabolism; GLUCOSIDE; GLUCOSIDOS; ION; IONES; Kidney - metabolism; Membrane Glycoproteins - metabolism; Membrane Potentials; Monosaccharide Transport Proteins - chemistry; Monosaccharide Transport Proteins - metabolism; Monosaccharide Transport Proteins - physiology; OVULE; OVULO; Phlorhizin - pharmacology; PORCIN; PROTEINAS; PROTEINE; REIN; RINONES; SODIO; SODIUM; Sodium - metabolism; Sodium-Glucose Transporter 1; STRUCTURE CELLULAIRE; Substrate Specificity; Swine; Xenopus laevis
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.271.51.32678

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 (KiPz≈ 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 (K0.5) for αMG (≈3 mM) was an order of magnitude higher for SGLT2; (ii) SGLT2 excluded galactose, suggesting discrete sugar binding; (iii) K0.5 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.