Alteration of channel characteristics by exchange of pore-forming regions between two structurally related Ca2+ channels.
Several types of structurally homologous high voltage-gated Ca2+ channels (L-, P- and N-type) have been identified via biochemical, pharmacological and electrophysiological techniques. Among these channels, the cardiac L-type and the brain BI-2 Ca2+ channel display significantly different biophysical properties. The BI-2 channel exhibits more rapid voltage-dependent current activation and inactivation and smaller single-channel conductance compared to the L-type Ca2+ channel. To examine the molecular basis for the functional differences between the two structurally related Ca2+ channels, we measured macroscopic and single-channel currents from oocytes injected with wild-type and various chimeric channel alpha 1 subunit cRNAs. The results show that a chimeric channel in which the segment between S5-SS2 in repeat IV of the cardiac L-type Ca2+ channel, was replaced by the corresponding region of the BI-2 channel, exhibited macroscopic current activation and inactivation time-courses and single-channel conductance, characteristic of the BI-2 Ca2+ channel. The voltage-dependence of steady-state inactivation was not affected by the replacement. Chimeras, in which the SS2-S6 segment in repeat III or IV of the cardiac channel was replaced by the corresponding BI-2 sequence, exhibited altered macroscopic current kinetics without changes in single-channel conductance. These results suggest that part of the S5-SS2 segment plays a critical role in determining voltage-dependent current activation and inactivation and single-channel conductance and that the SS2-S6 segment may control voltage-dependent kinetics of the Ca2+ channel.
Yatani
A
A
Department of Pharmacology and Cell Biophysics, University of Cincinnati, College of Medicine, Ohio 45267-0575.
Bahinski
A
A
Wakamori
M
M
Tang
S
S
Mori
Y
Y
Kobayashi
T
T
Schwartz
A
A
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001921:Brain / Q000502:physiology; D015220:Calcium Channels / Q000096:biosynthesis / Q000737:chemistry / Q000502:physiology*; D005260:Female; D006321:Heart / Q000502:physiology*; D015640:Ion Channel Gating; D046911:Macromolecular Substances; D008564:Membrane Potentials; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009865:Oocytes / Q000502:physiology; D016133:Polymerase Chain Reaction; D017433:Protein Structure, Secondary; D018075:RNA, Complementary; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry
D015220:Calcium Channels; D046911:Macromolecular Substances; D018075:RNA, Complementary; D011993:Recombinant Fusion Proteins