The facilitative glucose transporter 1 (GLUT1) mediates the passive diffusion of D-glucose across the cell membrane, providing the energy resource in glycolysis in the erythrocytes. Anion exchanger 1 (band 3) is another important membrane protein that mediates rapid exchange of CO2 through Cl-/HCO3- exchange across the erythrocyte membrane. For verifying the presumption over a decade that GLUT1 and band 3 in the erythrocyte would be interacting with each other, we cloned and expressed both the cytoplasmic domains of GLUT1 and band 3 in Escherichia coli, and tested their binding ability. By coimmunoprecipitation we found that among the tested N-terminal, C-terminal, and loop fraction of GLUT1, only the C-terminal of GLUT1 can interact with cytoplasmic domain of band 3. The interaction was further verified by coimmunoprecipitation and pull-down assay using both proteins as bait and target. These results showed that GLUT1 and band 3 form a protein complex that can regulate the activities of the proteins within it. (c) 2005 Elsevier Inc. All rights reserved.