Modeling of the outer vestibule and selectivity filter of the L-type Ca2+ channel

Using the KcsA bacterial K+ channel crystal structure [Doyle, D.A., et al. (1998) Science 280, 69-74] and the model of the outer vestibule of the Nachannel [Lipkind, G. M., and Fozzard, I-I. A. (2000) Biochemistry 39, 8161- 8170] as structural templates, we propose a structural model of the outer v estibule and selectivity filter of the pore of the Ca2+ channel (alpha (1c) or Ca(V)1.2). The Ca2+ channel P loops were modeled by alpha -helix-turn-b eta -strand motifs, with the glutamate residues of the EEEE motif located i n the turns. P loops were docked in the extracellular part of the inverted teepee structure formed by S5 and S6 alpha -helices with backbone coordinat es from the M1 and M2 helices of the KcsA crystal structure. This construct ion results in a conical outer vestibule that tapers to the selectivity fil ter at the bottom. The modeled selectivity ring forms a wide open pore (sim ilar to6 Angstrom) in the absence of Ca2+. When Ca2+ is present (similar to 1 muM), all four glutamate side chains move to the center and form a cage a round the dehydrated Ca2+ ion, blocking the pore. In the millimolar concent ration range, Ca2+ also interacts with two low-affinity sites located exter nally and internally, which were modeled by the same carboxylate groups of the selectivity filter. Calculation of the resulting electrostatic potentia ls show that the single Ca2+ ion is located in an electrostatic trap. Only when three Ca2+ ions are bound simultaneously in the high- and low-affinity sites of the selectivity filter is Ca2+ able to overcome electrostatic att raction, permitting Ca2+ flux.