Structural determinants for the glucose transport kinetics of the erythrocyte glucose transporter have not been established. In this work the role of the cytosolic carboxy-terminal tail in the expression and function of the human GLUT1 isoform in Xenopus oocytes was investigated. Oocyte plasma membrane expression of GLUT1 was a saturable function of the amount of mRNA injected. Transport activity increased as a linear function of the amount of immunoreactive transporter in the plasma membrane. Transport kinetics of human GLUT1 expressed in oocytes resembled those of human erythrocyte GLUT1. Addition of up to 31 extra amino acids to the carboxy-terminal tail of GLUT1 was without effect on its function in oocytes. Removal of the carboxy-terminal 21 amino acids also did not affect GLUT1 expression or transport kinetics in oocytes. Removal of the entire carboxy-terminal tail to Phe-450 resulted in a transporter that had moderately decreased plasma membrane expression compared to that of GLUT1. However, transport activity of this construct was less than 5% of that of GLUT1, and was associated with loss of its outward-facing inhibitor binding site. When the carboxy-terminal 29 amino acids of GLUT1 were replaced with the corresponding region of GLUT4, transporter expression in the plasma membrane and the transport Vmaxfell to low levels, similar to those of native GLUT4. When the carboxy-terminal 29 or 73 amino acids of GLUT1 were swapped into the corresponding region of GLUT4, the transport Vmaxmarkedly increased to about one-third to one-half that of GLUT1, although the affinity for substrate was halved. These results show that the carboxy-terminal tail of the GLUT1 is not critical for targeting of the protein to the plasma membrane, but that this region is an important determinant of transport function.