And Yet It Moves: Conformational States of the Ca2+ Channel Pore

Until recently, the two essential aspects of channel function seemed to be clearly distinct: a selectivity fi lter that discriminates among incoming ions and an intracellular gate that regulates the ion fl ux. As usual, reality is more complex, and this traditional view of ion channels is changing rapidly. Although a physical gate appears to operate at the intracellular part of the channel and undergo conformational transitions associated with the open-closed transitions of the channels, there is much more going on in the permeation pathway than was initially appreciated and the notion of a fairly “passive” or “rigid” selectivity fi lter is no longer adequate. Initial observations that different permeant ions could alter channel gating and/or inactivation rates pointed in that direction, but new data provides a compelling demonstration for an extended and more complex function of the pore, beyond its role in selectivity. Several pieces of evidence, obtained mostly in selective K+ pores, suggest that the selectivity fi lter can contribute substantially to channel gating, thereby indicating that the selectivity fi lter itself may also be undergoing conformational transitions. One of the fi rst studies supporting this view (Chapman et al., 1997), which was based on a careful analysis of the subconductance states in DRK1 channels, proposed that ion permeation and channel opening are coupled, meaning that the same structures that control selectivity also participate in channel opening and closing. More recently, Blunck et al. (2006) found that movement of the TM2 helix bundle, forming the inner gate in KcsA channels, is not the main determinant of the open probability. Fluorescence lifetime data led to the conclusion that the open probability of the bacterial channel is controlled by a second gate—likely the selectivity fi lter—in series with the bundle crossing gate. Opening of the intracellular gate may be a necessary, but not suffi cient, condition for ion conduction. In addition, insights provided by the crystal structure of the fi rst K+ pore (Morais-Cabral et al., 2001) has stimulated extensive theoretical work, including free energy molecular dynamic simulations of ion movement through a K+ selective pore, which subsequently led to the prediction of fl uctuations in the selectivity fi lter structure that likely increase the energy barrier for ion translocation, thereby de facto reproducing a nonconducting state without the involvement of an additional gate (Berneche and Roux, 2005). These “pore events” that underlie different gating mechanisms or inactivation processes (e.g., C-type inactivation) observed in voltage-gated K+ channels are direct consequences of a highly dynamic pore structure as opposed to a more static structure. Unfortunately, Ca2+ channel researchers do not (yet) enjoy the advantages associated with having a resolved crystal structure, so the molecular details of conduction, selectivity, and ion binding in the Ca2+ channels pore remain hazy. In addition, only limited assistance comes from the K+ pore structure because the Ca2+ pore (selectivity fi lter) seems to have molecular organization that differs from the K+ channels’ K+-binding carbonyl oxygens (provided by the pore TVGYG backbone chain). Rather, Ca2+ channel selectivity seems to rely on the carboxyl side chains of four glutamates (one from each of the four channel domains) that form a highly conserved EEEE locus that characterizes the pore of high voltage–activated channels (CaV1-CaV2 families; for review see Sather and McCleskey, 2003). Nevertheless, well designed experimental strategies can reveal important information about the likely mechanisms that underlie the function of these channels. In this issue, Babich et al. (p. 461 and 477) contribute two companion articles that report on molecular events that reveal possible structural rearrangement in the selectivity fi lter of L-type channels (CaV1.2). Babich et al. (2007a,b) cleverly exploited the Gd3+ block of the L-type channels to probe molecular events in the pore that occur during channel activation and inactivation. Submicromolar [Gd 3+] is known to produce both a tonic block and to accelerate decay of the L-type current during depolarizing steps (Biagi and Enyeart, 1990). The acceleration of current decay appears to be very similar to an enhancement of inactivation. Babich et al. (2007a,b) found that both these effects of Gd3+ blocks take place at the same site (which also binds Ca2+ or Ba2+ with a 1-mM affi nity), possibly located at the extracellular entrance of the selectivity fi lter, where the Gd3+