Replacement of intracellular C-terminal domain of GLUT1 glucose transporter with that of GLUT2 increases Vmax and Km of transport activity.
The intracellular C-terminal domain is diverse in size and amino acid sequence among facilitative glucose transporter isoforms. The characteristics of glucose transport are also divergent, and GLUT2 has far higher Km and Vmax values compared with GLUT1. To investigate the role of the intracellular C-terminal domain in glucose transport, we expressed in Chinese hamster ovary cells the mutated GLUT1 protein whose intracellular C-terminal domain was replaced with that of GLUT2 by means of engineering the chimeric cDNA. Cytochalasin B, for which GLUT2 protein has much lower affinity, bound to this chimeric protein in a fashion similar to GLUT1. In contrast, greater transport activity was observed in this chimeric glucose transporter compared with the wild-type GLUT1 at 10 mM 2-deoxy-D-glucose concentration. The kinetic studies on 2-deoxy-D-glucose uptake revealed a 3.8-fold increase in Km and a 4.3-fold increase in Vmax in this chimeric glucose transporter compared with the wild-type GLUT1. Thus, replacement of the intracellular C-terminal domain confers the GLUT2-like property on the glucose transporter. These results strongly suggest that the diversity of intracellular C-terminal domain contributes to the diversity of glucose transport characteristics among isoforms.
Katagiri
H
H
Third Department of Internal Medicine, Faculty of Medicine, University of Tokyo, Japan.
Asano
T
T
Ishihara
H
H
Tsukuda
K
K
Lin
J L
JL
Inukai
K
K
Kikuchi
M
M
Yazaki
Y
Y
Oka
Y
Y
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001692:Biological Transport; D016466:CHO Cells; D006224:Cricetinae; D003571:Cytochalasin B / Q000378:metabolism; D004247:DNA / Q000235:genetics; D003847:Deoxyglucose / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D009004:Monosaccharide Transport Proteins / Q000378:metabolism*; D011485:Protein Binding; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013329:Structure-Activity Relationship
D009004:Monosaccharide Transport Proteins; D011993:Recombinant Fusion Proteins; D003571:Cytochalasin B; D004247:DNA; D003847:Deoxyglucose