SUBUNIT INTERACTIONS IN COORDINATION OF NI2-GATED CHANNELS( IN CYCLICNUCLEOTIDE)

Cyclic nucleotide-gated (CNG) channels present a unique model for stud ying the molecular mechanisms of channel gating, We have studied the m echanism of potentiation of expressed rod CNG channels by Ni2+ as a fi rst step toward understanding the channel gating process. Here we repo rt that coordination of Ni2+ between histidine residues (H420) on adja cent channel subunits occurs when the channels are open, Mutation of H 420 to lysine completely eliminated the potentiation by Ni2+ but did n ot markedly alter the apparent cGMP affinity of the channel, indicatin g that the introduction of positive charge at the Ni2+-binding site wa s not sufficient to produce potentiation, Deletion or mutation of most of the other histidines present in the channel did not diminish poten tiation by Ni2+, We studied the role of subunit interactions in Ni2+ p otentiation by generating heteromultimeric channels using tandem dimer s of the rod CNG channel sequence, Injection of single heterodimers in which one subunit contained H420 and the other did not (wt/H420Q or H 420Q/wt) resulted in channels that were not potentiated by Ni2+, Howev er, coinjection of both heterodimers into Xenopus oocytes resulted in channels that exhibited potentiation, The H420 residues probably occur red predominantly in nonadjacent subunits when each heterodimer was in jected individually, but, when the two heterodimers were coinjected, t he H420 residues could occur in adjacent subunits as well, These resul ts suggest that the mechanism of Ni2+ potentiation involves intersubun it coordination of Ni2+ by H420. Based on the preferential binding of Ni2+ to open channels, we suggest that alignment of H420 residues of n eighboring subunits into the Ni2+-coordinating position may be associa ted with channel opening.