Purification and characterization of human erythrocyte glucose transporterin decylmaltoside detergent solution

The facilitative glucose transporter from human erythrocyte membrane, Glut1 , was purified by a novel method. The nonionic detergent decylmaltoside was selected for solubilization on the basis of its efficiency to extract Glut 1 from the erythrocyte membrane and its ability to maintain the protein in a monodisperse state. A positive, anion-exchange chromatography protocol pr oduced a Glut1 preparation of 95% purity with little copurified lipid. This protein preparation exhibited cytochalasin B binding in detergent solution , as measured by tryptophan fluorescence quenching. The transporter existed as a monomer in decylmaltoside, with a Stokes radius of 50 Angstrom and a molecular mass of 147 kDa for the protein-detergent complex. We screened de tergent, pH, additive, and lipid and have found conditions to maintain Glut 1 monodispersity for 8 days at 25 degreesC or over 5 weeks at 4 degreesC, T his Glut1 preparation represents the best available material for two- and t hree-dimensional crystallization trials of the human glucose transporter pr otein. (C) 2001 Academic Press.