Multiple Roles of the Extracellular Vestibule Amino Acid Residues in the Function of the Rat P2X4 Receptor
Figure 1
Effect of alanine and cysteine point mutations on the maximum current amplitude.
(A) Alanine and cysteine scanning mutagenesis of residues V47–V61 and F324–N338 that contain the upper parts of the TM1 and TM2 helices and the β-sheets in the open state. The maximum amplitude of currents (Imax) induced by 100 µM ATP in the wild type (WT; white bars) and cysteine (dark bars) and alanine (gray bars) mutant receptors. The receptors most affected had mutations at positions V49, Y54, Q55 (blue), F324 and G325 (red). The data are expressed as the mean ± SEM from 10–96 cells; **p<0.01 between WT and alanine or cysteine mutants. (B) The pattern of the ATP-induced currents by the WT and low-responsive rP2X4R alanine mutants. The horizontal bars indicate the duration of the application of 100 µM ATP (60 s). Notice the variable Y-scales for the WT and mutant receptors. (C) The topology of low-active residue mutants in the zfP2X4R in the apo-closed state (left) and ATP-bound open state (right); the mutated regions containing the upper parts of the TM1 and TM2 are shown in blue and red, respectively; affected mutated residues (rP2X4 numbering) are shown in red and blue spheres. Identity between rat and zebrafish P2X4R in amino acid sequences V47–V61 and F324–N338 is 67% and all functionally important residues are identical in both receptors.