Chapter 14 Sodium-D-Glucose Cotransport System: Biochemical Analysis of Active Sites

Abstract

This chapter describes the polymeric structure of the sodium-D-glucose co-transport system. To isolate the sodium-D-glucose co-transport system, negative extraction procedures are used, in which by successive use of proteases and detergents all membrane transport proteins except the one related to phlorizin binding or glucose transport are removed from the membrane, as well as positive extraction of the transport system from the membranes followed by enrichment through various chromatographic procedures. The chapter describes three functionally essential or active sites of the transport system: (1) the sugar binding site that also binds phlorizin, (2) the gating site that interacts with sodium, and (3) the portions of the protein that interact with the lipid phase of the membrane. Studies on the amino acid residues linked to the sugar and phlorizin-binding site have shown that organic and inorganic mercury compounds as well as N- ethylmaleimide (NEM) inhibit the binding of phlorizin to isolated brush border membranes. Under certain conditions, N-ethylmaleimide blocks only the high-affinity, sodium-dependent, D-glucose-inhibitable phlorizin binding sites related to the sodium-D-glucose co-transport system.