Biophysical characteristics of the pig kidney Na+/glucose cotransporter SGLT2 reveal a common mechanism for SGLT1 and SGLT2.
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 alpha-methyl-D-glucopyranoside (alphaMG) 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 approximately 10 microM) 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 approximately 1 and maximal charge transfer approximately 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 alphaMG (approximately 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.
Mackenzie
B
B
UCLA School of Medicine, Department of Physiology, Los Angeles, California 90095-1751, USA. dloo@physiology.medsch.ucla.edu
Loo
D D
DD
Panayotova-Heiermann
M
M
Wright
E M
EM
D000818:Animals; D001692:Biological Transport; D055592:Biophysical Phenomena; D001703:Biophysics; D004553:Electric Conductivity; D004594:Electrophysiology; D005947:Glucose / Q000378:metabolism*; D007668:Kidney / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000378:metabolism; D008564:Membrane Potentials; D009004:Monosaccharide Transport Proteins / Q000737:chemistry* / Q000378:metabolism / Q000502:physiology; D010695:Phlorhizin / Q000494:pharmacology; D012964:Sodium / Q000378:metabolism*; D051269:Sodium-Glucose Transporter 1; D013379:Substrate Specificity; D013552:Swine; D014982:Xenopus laevis
D008562:Membrane Glycoproteins; D009004:Monosaccharide Transport Proteins; D051269:Sodium-Glucose Transporter 1; D012964:Sodium; D010695:Phlorhizin; D005947:Glucose