Functional unit of 30 kDa for proximal tubule water channels as revealed by radiation inactivation

Abstract

The high water permeability of kidney proximal tubules is of paramount importance for isotonic reabsorption of 70% of the glomerular filtrate, and water channels have been postulated to account for the high water permeability. Target analysis following radiation inactivation was used to probe the molecular size of the water channel. Samples of brush border membranes from rat renal cortex were subjected to 3-MeV electron pulses from the Van de Graaff accelerator at a temperature of -130 degrees C. The inactivation of the renal brush border enzymes, alkaline phosphatase, and maltase was used for internal standardization of accumulated dose measurements in target analysis of the water channel. Osmotic water permeability was measured by following the change in scattered light intensity upon rapid mixing of vesicles with a hypertonic solution using stopped-flow spectrophotometry. The vesicle shrinkage response was biphasic and the rate of the fast phase decreased dose dependently by irradiation corresponding to a target size of 30 +/- 3.5 kDa. The total change in scattered light intensity was unaltered, indicating that irradiation did not destroy the lipid barrier. Our results provide strong support for the hypothesis that the high osmotic water permeability of renal proximal tubules results from a water channel-specific protein with a functional unit of 30 kDa.