Biomembrane-affinity centrifugal analyses of solute interactions with membrane proteins

We have developed a rapid centrifugal method for analyzing solute interacti ons with membrane proteins in cytoskeleton-depleted membrane vesicles or pr oteoliposomes sterically immobilized in Superdex 200 gel beads. The size an d density of the gel beads allow fast sedimentation in a bench-top centrifu ge. Biospecific interactions of cytochalasin B and D-glucose with the human red cell glucose transporter, Glut1, were analyzed. The binding constants and the molar ratio of inhibitor sites per protein monomer agreed well with recent results obtained by frontal affinity chromatography. (C) 1999 Elsev ier Science B.V. All rights reserved.