KINETIC AND SPECIFICITY DIFFERENCES BETWEEN RAT, HUMAN, AND RABBIT NA-GLUCOSE COTRANSPORTERS (SGLT-1)()

The Na+ activation and substrate specificity of human, rabbit, and rat Na+-glucose cotransporter (SGLT-1) isoforms were characterized using the Xenopus oocyte expression system and the two-electrode voltage-cla mp method. We find that there are differences, major and minor, in bot h the kinetics and substrate specificities between these isoforms; the substrate concentration at half-maximal current (K-0.5) for hexoses v aries from 0.2 to >40 mM, depending on the species and sugar; the affi nity constant (K-i) for phlorizin, the classic competitive inhibitor o f SGLT-1, varies over two orders of magnitude (rat K-i = 0.03 mu M vs. rabbit K-i = 1.4 mu M); and some glucoside inhibitors of the rabbit i soform, p-nitrophenyl glucose and beta-naphthyl glucose, are transport ed by the human and rat transporters. Na+ activation is more sensitive to membrane potential in the human and rat isoforms compared with rab bit. The rabbit isoform has a higher apparent affinity for alpha-methy lglucose and 3-O-methylglucose by a factor of two than either human or rat. These results can be quantitatively fitted by our six-state kine tic model of SGLT-1, providing insight into the processes involved in these changes. For example, the model predicts that Na+ binding (rate constant, k(12)) in human and rat SGLT-1 is similar but is fourfold la rger than in rabbit, whereas sugar binding (k(23)) in rabbit and rat i s similar but double the value in human SGLT-1. The differences in the primary amino acid sequences between these three homologous proteins must account for the kinetic and substrate specificity differences, an d comparisons of the functional properties and amino acid sequences of SGLT-1 isoforms provide useful information about structure/function r elationships.