The role of N-glycosylation of GLUT1 for glucose transport activity

T. Asano, Hideki Katagiri, K. Takata, J. L. Lin, H. Ishihara, K. Inukai, K. Tsukuda, M. Kikuchi, H. Hirano, Y. Yazaki, Y. Oka

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135 Citations (Scopus)

Abstract

To elucidate a functional role of N-glycosylation in glucose transporters, we introduced oligonucleotide-directed mutagenesis in GLUT1 cDNA to remove the possible site for N-linked glycosylation. The wild-type and the mutated GLUT1 cDNAs which induced a mutation of Asn at residue 45 to Asp, Tyr, or Gln were transfected and stably expressed into Chinese hamster ovary cells. The expressed wild-type and the mutated GLUT1 was demonstrated to be a broad band of a 45-60-kDa form and a sharp band of a 38-kDa form on Western blot analysis, respectively, indicating no glycosylation in the mutated GLUT1. Although the cell surface labeling of the glucose transporters demonstrated the presence of the glycosylation-defective glucose transporters on the cells surface, photoaffinity labeling of glycosylation-defective GLUT1 with [3H]cytochalasin B and a photoreactive mannose derivative, [3H]2-N-4-(1-azi-2,2,2,trifluoroethyl)benzoyl-1,3-bis(D-mannos-4- yloxy)-2-propylamine in the membranes was observed to be 40-70 and 15-30% of that of the wild-type GLUT1, respectively. The kinetic study of 2-deoxyglucose uptake revealed that the glycosylation-defective GLUT1 had a 2-2.5-fold greater K(m) value for 2-deoxyglucose uptake compared with the wild-type GLUT1. These observations strongly suggest that 1) N-glucosylation of GLUT1 glucose transporter is only on Asn 45 and 2) N-glycosylation plays an important role in maintaining a structure of glucose transporter with high affinity for glucose, thus, with high transport activity.

Original languageEnglish
Pages (from-to)24632-24636
Number of pages5
JournalJournal of Biological Chemistry
Volume266
Issue number36
Publication statusPublished - 1991 Dec 1
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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