Glucose transport activity and photolabelling with 3-[125I]iodo-4-azidophenethylamido-7-0-succinyldeacetyl (IAPS)-forskolin of two mutants at tryptophan-388 and -412 of the glucose transporter GLUT1: dissociation of the binding domains of forskolin and glucose
| Autor: | F.M. Brown, Konrad Keller, Sonja Wandel, Michael F. Shanahan, Ingrid Monden, HG Joost, Annette Schürmann |
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| Rok vydání: | 1993 |
| Předmět: |
endocrine system
Forskolin biology Chemistry Tryptophan Glucose transporter nutritional and metabolic diseases Transporter Cell Biology Transfection Ligand (biochemistry) Biochemistry Molecular biology carbohydrates (lipids) chemistry.chemical_compound biology.protein GLUT1 Binding site Molecular Biology hormones hormone substitutes and hormone antagonists |
| Zdroj: | Biochemical Journal. 290:497-501 |
| ISSN: | 1470-8728 0264-6021 |
| DOI: | 10.1042/bj2900497 |
| Popis: | The tryptophan residues 388 and 412 in the glucose transporter GLUT1 were altered to leucine (L) by site-directed mutagenesis and were transiently expressed in COS-7 cells. As assessed by immunoblotting, comparable numbers of glucose transporters were present in plasma membranes from cells transfected with wild-type GLUT1, GLUT1-L388 or GLUT1-L412. Transfection of the wild-type GLUT1 gave rise to a 3-fold increase in the reconstituted glucose transport activity recovered from plasma membranes. In contrast, transfection of GLUT1-L412 failed to increase the reconstituted transport activity, whereas transfection of GLUT1-L388 produced only a 70% increase. Photolabelling of GLUT1-L412 with 3-[125I]iodo-4-azidophenethylamido-7-O-succinyldeacetyl (125IAPS)-forskolin was not different from that of the wild-type GLUT1, whereas the GLUT1-L388 incorporated 70% less photolabel than did the wild-type GLUT1. These data suggest a dissociation of the binding sites of forskolin and glucose in GLUT1. Whereas both tryptophan-388 and tryptophan-412 appear indispensable for the function of the transporter, only tryptophan-388 is involved in the binding of the inhibitory ligand forskolin. |
| Databáze: | OpenAIRE |
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