Cytochalasin-B-binding proteins related to glucose transport across the basolateral membrane of the intestinal epithelial cell

Abstract

Basolateral membrane vesicles were isolated from mouse enterocytes. The vesicles showed Na+-independent uptake of D-glucose. The uptake was inhibited by cytochalasin B and phloretin with 50% inhibition at 2.0 microM and 25 microM, respectively. 2-Deoxy-D-glucose inhibited D-glucose transport by 50% at 0.5 M. The basolateral membranes bound 13.5 pmol mg protein-1 of 0.1 microM-cytochalasin B. The effects of various monosaccharides on cytochalasin B binding were examined; the strongest inhibitor was 2-deoxy-D-glucose (20-30%). Sodium dodecyl sulphate-polyacrylamide gel electrophoresis of the basolateral membranes labelled with [3H]cytochalasin B revealed two components with Mr values of 52(+/- 2) and 30(X 10(3)). Phloretin and 2-deoxy-D-glucose inhibited the photo-incorporation of [3H]cytochalasin B into these components. While phloretin inhibited the photolabelling of the two components to a similar extent, 2-deoxy-D-glucose seemed to inhibit preferentially that into the 52 X 10(3) Mr component. The similar sensitivity to 2-deoxy-D-glucose of the photolabelling of the 52 X 10(3) Mr component and of D-glucose transport, together with the fact that dithiothreitol removal increased the incorporation into the 52 X 10(3)Mr component and decreased that into the 30 X 10(3) Mr component, seems to suggest that the 52 X 10(3) Mr component is the major glucose transporter of the basolateral membrane.