Glycine scanning mutagenesis of the second TM helix of the G344A mutant. Amino acid sequence of the second TM helix of the P2X₂ channel. Macroscopic currents were recorded as in Fig. 2. The pulse protocol used is shown at the bottom left. Current traces recorded in the presence of 10 μM ATP are shown at positions corresponding to the introduced glycine in the TM helix. Current traces are shown after subtracting data obtained in the absence of ATP. In the drawing of the helix, the positions of the amino acid residues that when mutated to glycine rescued the voltage-dependent activation phases are highlighted.