3-O-methyl-D-glucose transport in rat red cells: effects of heavy water

Transport of 3-O-methyl-D-glucose (3-OMG) in rat red blood cells (RBCs) has been examined at 24 degrees C. The Km and Vm of zero-trans net uptake are 2.3 +/- 0.48 mM and 0.055 +/- 0.003 mumol (ml cell water)-1) min-1, whereas the Km and Vm for net exit are 2.1 +/- 0.12 mM and 0.12 +/- 0.01 mumol (ml cell water)-1 min-1. The Km and Vm for infinite-trans exchange uptake are 2.24 +/- 0.14 mM and 0.20 +/- 0.04 mumol (ml cell water)-1 min-1. In agreement with Whitesell et al. (Abumrad, N.A., Briscoe, P., Beth, A.H. and Whitesell, R.R. (1988) Biochim. Biophys. Acta 938, 222-230), we find that there is no significant acceleration of the rate of exchange exit over net exit. Substitution of D2O for water results in an increase in the Vm for zero-trans net uptake to 0.091 +/- 0.004 mumol (ml cell water)-1 min-1. There is no change in the Vm or Km for exchange uptake or net or exchange exit. Counterflow experiments indicate, in agreement with Helgerson and Carruthers (1989) Biochemistry 28, 4580-4594), that there is some compartmentalization of 3-OMG within the cells, perhaps resulting from slow complexation of the sugar with some intracellular component. The data can be simulated by assuming that transport across the membrane is mediated by either a fixed 2-site, or an alternating 1-site symmetrical transporter. With both models the observed asymmetries in net and exchange kinetics and in counterflow can be ascribed entirely to the complexation reaction of the sugar to an intracellular component. Also the D2O effects can entirely be attributed to an increase in the rate of sugar movement between bound and free compartments.