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Role of aromatic and charged ectodomain residues in the P2X 4 receptor functions
Author(s) -
Zemkova Hana,
Yan Zonghe,
Liang Zhaodong,
Jelinkova Irena,
Tomic Melanija,
Stojilkovic Stanko S.
Publication year - 2007
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/j.1471-4159.2007.04616.x
Subject(s) - ectodomain , mutant , receptor , chemistry , agonist , alanine , biophysics , biochemistry , microbiology and biotechnology , biology , amino acid , gene
The localization of ATP binding site(s) at P2X receptors and the molecular rearrangements associated with opening and closing of channels are still not well understood. At P2X 4 receptor, substitution of the K67, F185, K190, F230, R278, D280, R295, and K313 ectodomain residues with alanine generated low or non‐responsive mutants, whereas the F294A mutant was functional. The loss of receptor function was also observed in K67R, R295K, and K313R mutants, but not in F185W, K190R, F230W, R278K, and D280E mutants. To examine whether the loss of function reflects decreased sensitivity of mutants for ATP, we treated cells with ivermectin, an antiparasitic agent that enhances responsiveness of P2X 4 R. In the presence of ivermectin, all low or non‐responsive mutants responded to ATP in a dose‐dependent manner, with the EC 50 values for ATP of about 1, 2, 4, 20, 60, 125, 270, 420, 1000 and 2300 μmol/L at D280A, R278A, F185A, K190A, R295K, K313R, R295A, K313A, K67A and K67R mutants, respectively. These results indicate that lysines 67 and 313 and arginine 295 play a critical role in forming the proper three‐dimensional structure of P2X 4 R for agonist binding and/or channel gating.