CHARACTERIZATION OF GLUCOSE-TRANSPORT ACTIVITY RECONSTITUTED FROM HEART AND OTHER TISSUES

We examined several aspects of glucose transport reconstituted in lipo somes, with emphasis on transporters of rat heart (mostly GLUT4) compa red to those of human erythrocytes (GLUT1), and on effects of agents t hat modulate transport in intact cells. Several types of samples gave higher reconstituted activity using liposomes of egg lipids rather tha n soybean lipids. Diacylglycerol, proposed to activate transporters di rectly as part of the mechanism of insulin action, increased the intri nsic activity of heart transporters by only 25%, but increased the siz e of the reconstituted liposomes by 90%. The dipeptide Cbz-Gly-Phe-NH2 inhibited GLUT4 with a K-i of 0.2 mM, compared to 2.5 mM for GLUT1, w hich explains its preferential inhibition of insulin-stimulated glucos e transport in adipocytes. Verapamil, which inhibits insulin- and hypo xia-stimulated glucose transport in muscle, had no effect on reconstit uted transporters. Heart transporters had a higher K-m for glucose upt ake (13.4) than did GLUT1 (1.6 mM), in agreement with a recent study o f GLUT1 and GLUT4 expressed in yeast and reconstituted in liposomes. T ransporters reconstituted from heart and adipocytes were 40-70% inacti vated by external trypsin, suggesting the presence of trypsin-sensitiv e sites on the cytoplasmic domain of GLUT4. NaCl and KCl both reduced reconstituted transport activity, but KCI had a much smaller effect on the size of the liposomes. (C) 1998 Elsevier Science B.V. All rights reserved.