Accelerated net efflux of 3-O-methylglucose from rat adipocytes: a reevaluation

In a study of 3-O-methylglucose transport in insulin-stimulated rat adipocytes (catalyzed primarily by the GLUT4 isoform), it was reported that at 37 degrees C the Km and Vmax were 2.8-fold higher for net efflux than for equilibrium exchange (Vinten, J. (1984) Biochim. Biophys. Acta 772, 244-250). Because of its implications for the relative sizes of steps in the transport cycle, we reinvestigated this phenomenon. Accelerated net efflux was apparent when the extracellular methylglucose was diluted 26-fold but not when it was diluted 11-fold. When analyzed according to the one-site alternating conformation model, the data indicate about a 1.7-fold higher Vmax for efflux than for exchange, only about 40% of the difference reported previously. Together with other results in the literature, the accelerated net flux indicates that the conformational change of the loaded transporter from its outward-facing to its inward-facing form is likely the slowest step in the transport cycle, in contrast to the case for GLUT1. Experiments at 25 degrees C indicate a lower degree of accelerated net flux than at 37 degrees C. This is consistent with the above conformational change being the step with the lowest activation energy, as for GLUT1.