RT Journal Article
SR Electronic
T1 Photoaffinity labeling of the partially purified mitochondrial phenylalkylamine calcium antagonist receptor.
JF Molecular Pharmacology
JO Mol Pharmacol
FD American Society for Pharmacology and Experimental Therapeutics
SP 1010
OP 1013
VO 42
IS 6
A1 G Zernig
YR 1992
UL http://molpharm.aspetjournals.org/content/42/6/1010.abstract
AB Mitochondria contain specific Ca2+ antagonist binding sites that are associated with an inner mitochondrial membrane anion channel. These mitochondrial Ca2+ antagonist receptors can be solubilized with digitonin and partially purified [as assessed by postreversible (+/-)-[3H]nitrendipine binding] using ion exchange chromatography and sucrose density gradient centrifugation. In the present study, reversible binding of the phenylakylamine Ca2+ antagonist [3H]ludopamil, an optically pure photoaffinity analog of verapamil, to the partially purified mitochondrial Ca2+ antagonist receptor complex (Kd, 9 +/- 4 microM; Bmax, 1.2 +/- 0.5 nmol/mg of protein) depended on NaNO3 and was inhibited by the 1,4-dihydropyridine niludipine and by ATP. Accordingly, the unlabeled racemic analog of [3H]ludopamil, (+/-)-LU 47781, dose-dependently inhibited the binding of the 1,4-dihydropyridine (+/-)-[3H]nitrendipine to the purified mitochondrial receptors (IC50, 2.1 +/- 0.1 microM). After UV irradiation, [3H]ludopamil specifically incorporated into two polypeptides of 12.7 +/- 0.1 kDa and 11.7 +/- 0.1 kDa, with the pharmacological profile of [3H]ludopamil photoincorporation stimulation and protection being identical to that of reversible binding.