title: Molecular determinants of Ca2+ selectivity and ion permeation in L-type Ca2+ channels.
abstract: Voltage-gated Ca2+ channels link changes in membrane potential to the delivery of Ca2+, a key second messenger for many cellular responses. Ca2+ channels show selectivity for Ca2+ over more plentiful ions such as Na+ or K+ by virtue of their high-affinity binding of Ca2+ within the pore. It has been suggested that this binding involves four conserved glutamate residues in equivalent positions in the putative pore-lining regions of repeats I-IV in the Ca2+ channel a1 subunit. We have carried out a systematic series of single amino-acid substitutions in each of these positions and find that all four glutamates participate in high-affinity binding of Ca2+ or Cd2+. Each glutamate carboxylate makes a distinct contribution to ion binding, with the carboxylate in repeat III having the strongest effect. Some single glutamate-to-lysine mutations completely abolish micromolar Ca2+ block, indicating that the pore does not possess any high-affinity binding site that acts independently of the four glutamate residues. The prevailing model of Ca2+ permeation must thus be modified to allow binding of two Ca2+ ions in close proximity, within the sphere of influence of the four glutamates. The functional inequality of the glutamates may be advantageous in allowing simultaneous interactions with multiple Ca2+ ions moving single-file within the pore. Competition among Ca2+ ions for individual glutamates, together with repulsive ion-ion electrostatic interaction, may help achieve rapid flux rates through the channel.
authors.affiliation: Department of Molecular and Cellular Physiology, Beckman Center, Stanford University Medical Center, California 94305.
authors.forename: J
authors.initials: J
authors.lastname: Yang
authors.forename: P T
authors.initials: PT
authors.lastname: Ellinor
authors.forename: W A
authors.initials: WA
authors.lastname: Sather
authors.forename: J F
authors.initials: JF
authors.lastname: Zhang
authors.forename: R W
authors.initials: RW
authors.lastname: Tsien
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D002463:Cell Membrane Permeability; D002478:Cells, Cultured; D005971:Glutamates / Q000378:metabolism; D007477:Ions; D008969:Molecular Sequence Data; D009154:Mutation; D009206:Myocardium / Q000378:metabolism; D011817:Rabbits; D014981:Xenopus
chemical_list: D015220:Calcium Channels; D005971:Glutamates; D007477:Ions; D002118:Calcium