RT Journal Article
SR Electronic
T1 Polar Residues in the Second Transmembrane Domain of the Rat P2X2 Receptor That Affect Spontaneous Gating, Unitary Conductance, and Rectification
JF The Journal of Neuroscience
JO J. Neurosci.
FD Society for Neuroscience
SP 14257
OP 14264
DO 10.1523/JNEUROSCI.4403-09.2009
VO 29
IS 45
A1 Cao, Lishuang
A1 Broomhead, Helen E.
A1 Young, Mark T.
A1 North, R. Alan
YR 2009
UL http://www.jneurosci.org/content/29/45/14257.abstract
AB Membrane ion channels activated by extracellular ATP (P2X receptors) are widely distributed in the nervous system. Their molecular architecture is fundamentally distinct from that of the nicotinic or glutamate receptor families. We have measured single-channel currents, spontaneous gating, and rectification of rat P2X2 receptor in which polar and charged residues of the second transmembrane domain (TM2) were systematically probed by mutagenesis. The results suggest that Asn333 and Asp349 lie respectively in external and internal vestibules. Substitutions at Asn333, Thr336, and Ser340 were particularly likely to cause spontaneously active channels. At Thr336, Thr339, and Ser340, the introduction of positive charge (Arg, Lys, or His, or Cys followed by treatment with 2-aminoethyl methanethiosulphonate) greatly enhanced outward currents, suggesting that side-chains of these three residues are exposed in the permeation pathway of the open channel. These functional findings are interpreted in the context of the recently reported 3.1 Å crystal structure of the zebrafish P2X4.1 receptor in the closed state. They imply that the gate is formed by residues Asn333 to Thr339 and that channel opening involves a counter-clockwise rotation and separation of the TM2 helices.