Expression of a differentiated transport function in apical membrane vesicles isolated from an established kidney
Abstract:
Electrogenic Na+ gradient-stimulated active accumulation of alpha-methyl-D-glucopyranoside was demonstrated using apical membrane vesicles isolated from an established kidney epithelial cell line (LLC-PK1). Apical membrane vesicles were purified from densely confluent cell cultures after cell disruption by nitrogen cavitation using a rapid Mg2+ differential precipitation procedure. Such preparations were 11-fold enriched in apical-specific enzyme markers, were not enriched in basolateral-specific markers, and were relatively free from contamination by intracellular membranes. Sugar accumulated within vesicles was sensitive to osmotic pressure differences across the membrane. alpha-Methyl-D-glucopyranoside transport activity in vesicles exhibited similar hexose specificities and sensitivity to inhibition by phlorizin to the corresponding transport system expressed in intact LLC-PK1 cells and in kidney proximal tubule. Vesicles prepared from subconfluent cultures, which have not yet developed the expression of this transport system, had diminished Na+-stimulated sugar transport activity yet expressed Na+-stimulated amino acid transport activity. Active accumulation as a function of increasing Na+ gradient and an interior-negative membrane potential suggested a stoichiometry between sugar and Na+ uptake of 1:2, respectively. Experimental generation of interior-negative membrane potentials in vesicles produced a further stimulation of sugar accumulation, consistent with an additive contribution from both a Na+ chemical gradient and an electrical membrane potential in energizing active sugar transport.
More Related Videos
Related Concept Videos
Secondary Active Transport
Membrane Transporters
Transporters are mainly composed of alpha-helices, built from bundles of ten or more helices traversing the plasma membrane. The solute-binding sites are located midway, where some of the helices are broken or distorted, making space for the binding site through...
Secondary Active Transport
Transcellular Transport of Solutes
Secondary Active Transport
Active Transport
Primary active transporters, like Na+, K+ and -ATPase, directly utilize ATP to move ions across the membrane. These transporters play significant roles in various physiological processes. For instance, Na+, K+ and -ATPase maintain...


