The renal brush border membrane sodium/sulfate cotransporter functions in situ as a homotetramer

The functional molecular size of the renal Na+/SO4(2-) cotransporter was analysed with the radiation inactivation and fragmentation method. Purified brush border membrane vesicles preserved in a cryoprotective medium were exposed to gamma-radiations. Initial rates of SO4(2-) influx into these vesicles were estimated with membranes irradiated with 0, 4 and 8 Mrad. In each case, SO4(2-) uptake by irradiated membranes was significantly reduced but remained linear during the first 5 sec of incubation. To avoid artifacts arising from a decrease in the driving force caused by modifications in membrane permeability, this incubation period was chosen to measure the effect of irradiation on the SO4(2-) transport activity. Increasing irradiation doses resulted in a monoexponential decrease in transport activity allowing the molecular size to be estimated at 238 +/- 6 kDa (SD, n = 3). Recently, a cDNA for the Na+/SO4(2-) cotransporter was cloned and expressed in Xenopus laevis oocytes (Markovich D. et al. (1993) Proc. Natl Acad. Sci. U.S.A. 90, 8073-8077). The deduced amino acid sequence of this cotransporter predicts a molecular weight of 66 kDa. We suggest that the in situ activity of the renal brush border membrane Na+/SO4(2-) cotransporter requires the presence of four intact and identical subunits arranged as a homotetramer.