A SITE ACCESSIBLE TO EXTRACELLULAR TEA+ AND K+ INFLUENCES INTRACELLULAR MG2+ BLOCK OF CLONED POTASSIUM CHANNELS

The members of the RCK family of cloned voltage-dependent K+ channels are quite homologous in primary structure, but they are highly diverse in functional properties. RCK4 channels differ from RCK1 and RCK2 cha nnels in inactivation and permeation properties, the sensitivity to ex ternal TEA, and to current modulation by external K+ ions. Here we sho w several other interesting differences: While RCK1 and RCK2 are block ed in a voltage and concentration dependent manner by internal Mg2+ io ns, RCK4 is only weakly blocked at very high potentials. The single-ch annel current-voltage relations of RCK4 are rather linear while RCK2 e xhibits an inwardly rectifying single-channel current in symmetrical K + solutions. The deactivation of the channels, measured by tail curren t protocols, is faster in RCK4 by a factor of two compared with RCK2. In a search for the structural motif responsible for these differences , point mutants creating homology between RCK2 and RCK4 in the pore re gion were tested. The single-point mutant K533Y in the background of R CK4 conferred the properties of Mg2+ block, tail current kinetics, and inward ion permeation of RCK2 to RCK4. This mutant was previously sho wn to be responsible for the alterations in external TEA sensitivity a nd channel regulation by external K+ ions. Thus, this residue is expec ted to be located at the external side of the pore entrance. The data are consistent with the idea that the mutation alters the channel occu pancy by K+ and thereby indirectly affects internal Mg2+ block and cha nnel closing.