Abstract
A protein that binds the calcium-channel antagonist verapamil has been partially purified from maize (Zea mays) coleoptile membranes. The protein was solubilized with the detergent CHAPS ([ 3-(3-cholamidopropyl)dimethylammonio]propane-1-sulphonate) and purified by a combination of ion-exchange, gel-filtration and hydrophobic-interaction chromatography. This resulted in a 120-fold purification. SDS/polyacrylamide-gel-electrophoretic analysis of the polypeptides from the final purification step indicated that the verapamil-binding protein may have a major component of Mr 169,000. The dissociation constants for specific binding of [3H]verapamil to crude and CHAPS-solubilized maize coleoptile membrane fractions are 72 nM and 158 nM respectively, with respective binding-site concentrations of 135 pmol/mg of protein and 78 pmol/mg of protein. In both cases the Scatchard plots are linear, indicating a single class of binding sites. [3H]Verapamil binding to crude maize coleoptile membrane fractions could not be displaced by unlabelled desmethoxyverapamil or by nifedipine, but could be displaced by unlabelled methoxyverapamil.