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Substitution of conserved tyrosine residues in helix 4 (Y143) and 7 (Y293) affects the activity, but not IAPS‐forskolin binding, of the glucose transporter GLUT4
Author(s) -
Wandel Sonja,
Schürmann Annette,
Becker Walter,
Summers Scott A.,
Shanahan Michael F.,
Joost Hans G.
Publication year - 1994
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(94)00558-3
Subject(s) - tyrosine , glut4 , chemistry , transporter , glucose transporter , biochemistry , forskolin , helix (gastropod) , stereochemistry , biology , endocrinology , insulin , receptor , gene , ecology , snail
Six tyrosine residues (Y28, Y143, Y292, Y293, Y308, Y432 1 ) which are conserved in all mammalian glucose transporters were substituted for phenylalanine by site‐directed mutagenesis, and mutant glucose transporters were transiently expressed in COS‐7 cells. Glucose transport activity as assessed by reconstitution of the solubilized transporters into lecithin liposomes was reduced by 70% in the mutant Y143F and appeared to be abolished in Y293F, but was not affected by substitution of Y28, Y292, Y308 and Y432. In contrast, covalent binding of the photolabel 125 IAPS‐forskolin was normal in all mutants. Stable expression of the mutants Y143F, Y293F, and Y292F in LTK cells yielded identical results. These data indicate that only two of the 6 conserved helical tyrosines residues, located in helices 4 and 7, are essential for full activity, but not for IAPS‐forskolin binding of the GLUT4.