L-TYPE CALCIUM CHANNELS - BINDING DOMAINS FOR DIHYDROPYRIDINES AND BENZOTHIAZEPINES ARE LOCATED IN CLOSE PROXIMITY TO EACH OTHER

We investigated the binding of a fluorescent diltiazem analogue -indac en-3-yl]propionyl]amino]propyl]amino]ethyl]- yphenyl)-6-(trifluorometh yl)-2H-1-benzazepin-2-one (DMBODIPY-BAZ) to L-type Ca2+ channels in th e presence of different 1,4-dihydropyridines (DHPs) by using fluoresce nce resonance energy transfer (FRET) [Brauns, T., Cai, Z.-W., Kimball, S. D., Kang, H.-C., Haugland, R. P., Berger, W., Berjukov, S., Hering , S., Glossmann, H., Br Striessnig, J. (1995) Biochemistry 34, 3461]. When channels are occupied with DMBODIPY-BAZ, a rapid fluorescence cha nge occurred upon addition of different DHPs. The direction of this in tensity modulation was found to be only dependent on the chemical comp osition of the dihydropyridine employed. DHPs containing a nitro group decreased, whereas others (e.g., isradipine) enhanced the fluorescenc e signal. In addition, all DHPs markedly decreased the association rat e constant for DMBODIPY-BAZ without affecting equilibrium binding. Bot h observations together are best explained by a steric model where the DHP binding site is located in close proximity to the accession pathw ay of DMBODIPY-BAZ.