Expression cloning and cDNA sequencing of the Na+/glucose co-transporter

Abstract

Organic substrates (sugars, amino acids, carboxylic acids and neutrotransmitters) are actively transported into eukaryotic cells by Na+ co-transport. Some of the transport proteins have been identified--for example, intestinal brush border Na+/glucose and Na+/proline transporters and the brain Na+/CI-/GABA transporter--and progress has been made in locating their active sites and probing their conformational states. The archetypical Na+-driven transporter is the intestinal brush border Na+/glucose co-transporter (see ref. 8), and a defect in the co-transporter is the origin of the congenital glucose-galactose malabsorption syndrome. Here we describe cloning of this co-transporter by a method new to membrane proteins. We have sequenced the cloned DNA and have found no homology between the Na+/glucose co-transporter and either the mammalian facilitated glucose carrier or the bacterial sugar transport proteins. This suggests that the mammalian Na+-driven transporter has no evolutionary relationship to the other sugar transporters.

Ontogenetic expression and regulation of Na+-d-glucose cotransporter in jejunum of domestic chicken

“…The molecular mass was very similar to that in adult chickens (11) and to those reported (13,23,26) for SGLT1 in various species. These results indicate that there were no age-related structural changes in SGLT1 that could explain the reduction in methyl ␣-D-glucopyranoside transport.…”

“…Blots and tissue samples were incubated with a rabbit polyclonal antibody, donated by Dr. M. Kasahara (Teikyo University), raised against the synthetic peptide corresponding to amino acids 564-575 of the deduced amino acid sequence of rabbit intestinal SGLT1 (13). Purified BBMV from rabbit small intestine was used as a reference material throughout.…”

Glycyl-l-proline transport in rabbit enterocyte basolateral-membrane vesicles

“…It is absent from basolateral membranes [19,25,26]. The 75 kDa Na+/glucose symporter is also specific to the brush-border membrane [28,29]. The major histocompatibility antigen (anti-RLA Class 1) is uniquely located on the basolateral membrane [19,25].…”

“…After electrotransfer on to cellulose nitrate sheets, the potential of the proteins to interact with the monoclonal antibody 23B921 raised against the 140 kDa antigen [19], the polyclonal antibodies raised against the major Vol. 269 histocompatibility antigen (anti-RLA class I) [30,31] and polyclonal antibodies raised against the Na+/glucose symporter [28,29] was assessed. As shown in Fig.…”

Kinetics of the Reverse Mode of the Na+/Glucose Cotransporter

“…The cloning of SGLT1 and its over-expression in Xenopus laevis oocytes allowed for a detailed characterization of the kinetics of cotransport (Hediger et al, 1987;Parent et al, 1992a,b;Loo et al, 1993;Hazama et al, 1997;Loo et al, 1998). Electrophysiological and tracer uptake experiments in intact oocytes expressing SGLT1 focused on the forward mode, where the substrates are transported from the extracellular to intracellular compartments.…”