Isoform-Selective Inhibition of Facilitative Glucose Transporters: Elucidation of the Molecular Mechanism of HIV Protease Inhibitor Binding [Cell Biology]

April 4th, 2014 by Hresko, R. C., Kraft, T. E., Tzekov, A., Wildman, S. A., Hruz, P. W.

Pharmacologic inhibitors of the HIV Protease (PIs) and structurally related oligopeptides are known to reversibly bind and inactivate the insulin-responsive facilitative glucose transporter GLUT4. Several PIs exhibit isoform selectivity with little effect on GLUT1. The ability to target individual GLUT isoforms in an acute and reversible manner provides novel means both to investigate the contribution of individual GLUTs to health and disease and to develop targeted treatment of glucose-dependent diseases. To determine the molecular basis of transport inhibition, a series of chimeric proteins containing transmembrane (TM) and cytosolic (Cyt) domains from GLUT1 and GLUT4 and/or point mutations were generated and expressed in HEK293 cells. Structural integrity was confirmed via measurement of ATB-BMPA labeling of the chimeric proteins in low-density microsome fractions isolated from stably transfected 293 cells. Functional integrity was assessed via measurement of zero-trans 2-deoxyglucose uptake (2-DOG). ATB-BMPA labeling studies and 2-DOG uptake revealed that TM helices 1 and 5 contain amino acid residues that influence inhibitor access to the transporter binding domain. Substitution of Thr30 and His160 in GLUT1 to the corresponding positions in GLUT4 is sufficient to completely transform GLUT1 into GLUT4 with respect to indinavir inhibition of 2-DOG uptake and ATB-BMPA binding. These data provide a structural basis for the selectivity of PIs toward GLUT4 over GLUT1 that can be used in ongoing novel drug design.
  • Posted in Journal of Biological Chemistry, Publications
  • Comments Off on Isoform-Selective Inhibition of Facilitative Glucose Transporters: Elucidation of the Molecular Mechanism of HIV Protease Inhibitor Binding [Cell Biology]