Dihydropyridines (DHPs) are an important class of drugs, used extensively in the treatment of angina pectoris, hypertension, and arrhythmia. The molecular mechanism by which DHPs modulate Ca2+ channel function is not known in detail. We have found that DHP binding is allosterically coupled to Ca 2+ binding to the selectivity filter of the skeletal muscle Ca 2+ channel CaV1.1, which initiates excitation-contraction coupling and conducts L-type Ca2+ currents. Increasing Ca 2+ concentrations from approximately 10 nM to 1 mM causes the DHP receptor site to shift from a low-affinity state to a high-affinity state with an EC50 for Ca2+ of 300 nM. Substituting each of the four negatively charged glutamate residues that form the ion selectivity filter with neutral glutamine or positively charged lysine residues results in mutant channels whose DHP binding affinities are decreased up to 10-fold and are up to 150-fold less sensitive to Ca2+ than wild-type channels. Analysis of mutations of amino acid residues adjacent to the selectivity filter led to identification of Phe-1013 and Tyr-1021, whose mutation causes substantial changes in DHP binding. Thermodynamic mutant cycle analysis of these mutants demonstrates that Phe-1013 and Tyr-1021 are energetically coupled when a single Ca2+ ion is bound to the channel pore. We propose that DHP binding stabilizes a nonconducting state containing a single Ca2+ ion in the pore through which Phe-1013 and Tyr-1021 are energetically coupled. The selectivity filter in this energetically coupled high-affinity state is blocked by bound Ca2+, which is responsible for the high-affinity inhibition of Ca2+ channels by DHP antagonists.
All Science Journal Classification (ASJC) codes
- Molecular Medicine