Monosaccharide transport in protein-depleted vesicles from erythrocyte membranes.

M A Zoccoli,G E Lienhard

doi:10.1016/s0021-9258(17)40361-9

Abstract

Treatment of human erythrocyte membranes with dilute alkali (pH 11.5) generates sealed, protein-depleted vesicles that can be isolated by density gradient centrifugation. The vesicles are 0.5 to 2.0 micrometers in diameter, and their membranes are predominantly oriented inside-out. The vesicles lack protein bands 1, 2, 5, and 6 (nomenclature of Steck, T.L. (1974) J. Cell Biol. 62, 1-19) of the erythrocyte membrane. L-Sorbose, a substrate of the monosaccharide transport system in erythrocytes, is transported by the vesicles. Based on comparisons between erythrocytes and vesicles with regard to specificity, temparture dependence, and effects of inhibitors, we conclude that sorbose uptake into the vesicles occurs by way of the monosaccharide transport system. The specific activity of the transport system in vesicles, as determined by initial rate measurements of sorbose uptake, averaged 58% of that in erythrocytes. This finding indicates that the major polypeptides of Bands 1, 2, 5, and 6 do not play an obligatory role in monosaccharide transport.

Highlights

Treatment of human erythrocyte membranes with dilute alkali generates sealed, protein-depleted vesicles that can be isolated by density gradient centrifugation
Based on comparisons between erythrocytes and vesicles with regard to specificity, temperature dependence, and effects of inhibitors, we conclude that sorbose uptake into the vesicles occurs by way of the monosaccharide transport system
Vesicles - The pellet of protein-depleted membrane fragments that was obtained by the treatment of erythrocyte ghosts with dilute alkali separated into three fractions upon density gradient centrifugation in dextran T-70

Summary

Introduction

Treatment of human erythrocyte membranes with dilute alkali (pH 11.5) generates sealed, protein-depleted vesicles that can be isolated by density gradient centrifugation. A substrate of the monosaccharide transport system in erythrocytes, is transported by the vesicles. Based on comparisons between erythrocytes and vesicles with regard to specificity, temperature dependence, and effects of inhibitors, we conclude that sorbose uptake into the vesicles occurs by way of the monosaccharide transport system. The specific activity of the transport system in vesicles, as determined by initial rate measurements of sorbose uptake, averaged 58% of that in erythrocytes. This finding indicates that the major polypeptides of Bands 1, 2, 5, and 6 do not play an obligatory role in monosaccharide transport. The monosaccharide transport system of the human erythrocyte catalyzes the equilibration of n-glucose and other sugars across the cellular membrane. As an approach to this problem, we have made use of the finding of Steck and

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Conclusion