J Biol Chem 1997 Jan 24;2724:2110-5.

Cation effects on protein conformation and transport in the Na+/glucose cotransporter.

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Cation-driven cotransporters are essential membrane proteins in procaryotes and eucaryotes, which use the energy of the transmembrane electrochemical gradient to drive transport of a substrate against its concentration gradient. Do they share a common mechanism? Cation selectivity of the rabbit isoform of the Na+/glucose cotransporter (SGLT1) was examined using the twoelectrode voltage clamp and the Xenopus oocyte expression system. The effect of H+, Li+, and Na+ on kinetics of SGLT1 was compared to the effects of these cations on the bacterial melibiose. In SGLT1, substitution of H+ or Li+ for Na+ caused a kinetic penalty in that the apparent affinity for sugar (K0.5sugar) decreased by an order of magnitude or more (from 0.2 to 30 mM) depending on the membrane potential and cation. The effect of the cation on the K0.5sugar/V profiles was independent of the sugar for glucose and alpha-methyl-beta-D-glucose; this profile was maintained for galactose in Li+ and Na+, but was 2 orders of magnitude higher in H+, but the Imax for glucose, galactose, and alpha-methyl-beta-D-glucose in a given cation were identical. Li+ supported a lower maximal rate of transport (Imax) than Na+ (approximately 80% of ImaxNa), while the Imax in H+ was higher than Na+ (>/=180% of ImaxNa). Our interpretation of these results and simulations using a six-state mathematical model, are as follows. 1) Binding of the cation causes a conformational change in the sugar binding pocket, the exact conformation being determined by the specific cation. 2) Once the sugar is bound, it is transported at a characteristic rate determined by the cation. 3) Mathematical simulations suggest that the largest contribution to the kinetic variability of both cation and sugar transport is associated with cation binding. Similarity to the effects of cation substitution in MelB suggests that the mechanism of energy coupling has been evolutionarily conserved.

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Species referenced: Xenopus laevis
Genes referenced: slc5a1.2