Construction of a high-affinity receptor site for dihydropyridine agonists and antagonists by single amino acid substitutions in a non-L-type Ca2+ channel

Gregory H. Hockerman, Blaise Z. Peterson, Elizabeth Sharp, Timothy N. Tanada, Todd Scheuer, William A. Catterall

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The activity of L-type Ca2+ channels is increased by dihydropyridine (DHP) agonists and inhibited by DHP antagonists, which are widely used in the therapy of cardiovascular disease. These drugs bind to the pore-forming α1 subunits of L-type Ca2+ channels. To define the minimal requirements for DHP binding and action, we constructed a high-affinity DHP receptor site by substituting a total of nine amino acid residues from DHP-sensitive L-type α1 subunits into the S5 and S6 transmembrane segments of domain III and the S6 transmembrane segment of domain IV of the DHP-insensitive P/Q-type α1A subunit. The resulting chimeric α1A/DHPS subunit bound DHP antagonists with high affinity in radioligand binding assays and was inhibited by DHP antagonists with high affinity in voltage clamp experiments. Substitution of these nine amino acid residues yielded 86% of the binding energy of the L-type α1C subunit and 92% of the binding energy of the L-type α1S subunit for the high-affinity DHP antagonist PN200-110. The activity of chimeric Ca2+ channels containing α1A/DHPS was increased 3.5 ± 0.7-fold by the DHP agonist (-)Bay K8644. The effect of this agonist was stereoselective as in L-type Ca2+ channels since (+) Bay K8644 inhibited the activity of α1A/DHPS. The results show conclusively that DHP agonists and antagonists bind to a single receptor site at which they have opposite effects on Ca2+ channel activity. This site contains essential components from both domains III and IV, consistent with a domain interface model for binding and allosteric modulation of Ca2+ channel activity by DHPs.

Original languageEnglish (US)
Pages (from-to)14906-14911
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Issue number26
Publication statusPublished - Dec 23 1997


All Science Journal Classification (ASJC) codes

  • General

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