The respective absorptive and reabsorptive roles of the small intestine and the proximal kidney in nutrient accumulation and conservation are to a major extent subserved by Na ion-dependent carriers in the brush border membranes of their epithelial linings. There are over a dozen carriers of this general class differentiated by their specific substrates which range from sugars, amino acids, bile salts or vitamins to anions such as phosphate. The malfunction or absence of some of these carriers have been shown or imputed to underly such human diseases as glucose-galactose malabsorption, cystinuria and vitamin-D resistant, X-linked hypophosphatemia, among others. The Na ion-dependent glucose carrier may be taken as representative of the general class and our long-term objective is to arrive at an understanding of this transport modality in terms of its structure and its mechanism of operation in the brush border membrane. We have developed an assay for isolated active carriers based upon their reconstitution into liposomes and with its aid have identified a single protein band of the brush border membrane as being present in all active preparations. We are proceeding to purify this band and to resolve its components using electrophoretic, centrifugal and specific adsorption methods where applicable. We also studied the kinetics of the Na ion-dependent glucose carrier in the natural brush border membrane prepared in vesicular form. We have developed computer programs to resolve the data and have identified the kinetic type of the reaction. The kinetics newly seen have also led to the formulation of a concept of mechanism of Na ion-dependent glucose transport quite different from conventional mobile carrier ideas. The coupling between the fluxes of Na ion and substrate which characterizes Na ion-dependent carriers is in mobile carrier theory substantially mechanical. In our model it is substantially chemical. Current objectives, based upon the kinetics of the system, are to visualize carrier interactions by such means as fluorescent probes, ion inhibition and the effect of alterations in membrane potentials so as to discriminate the predictions of the carrier and chemical coupling models.
NIH ExPorter
NIH ExPorter