CHARACTERIZATION OF THE BINDING OF OMEGA-CONOPEPTIDES TO DIFFERENT CLASSES OF NON-L-TYPE NEURONAL CALCIUM CHANNELS

The interaction of two synthetic omega-conopeptides SNX-111 (MVIIA) and SNX-230 (MVIIC) both derived from the marine snail Conus magus, with non-L-type neuronal voltage-sensitive calcium channels (VSCC) in rat-brain synaptosomal preparations has been investigated with the aid of well-characterized I-125 derivatives of the two peptides. To assess the effects of iodination on the binding characteristics of SNX-111 and SNX-230, the corresponding peptides containing monoiodotyrosine in place of tyrosine, namely, SNX-259 ([I-127]SNX-111) and SNX-260 ([I-127]SNX-230), respectively, were prepared by solid-phase synthesis. Saturation analysis showed that [I-125]SNX-111 and [I-125]SNX-230 bound to two distinct classes of high-affinity sites with apparent K-d's Of 9 and II pM and B-max's of 0.54 and 2.2 pmol/mg protein, respectively. Kinetic analysis revealed that both peptides exhibited; high association rates as well as rapid dissociation rates in contrast to the I-125 derivative of the synthetic omega-conopeptide from Conus geographus, GVIA (SNX-124), which binds irreversibly to N-type channels in rat brain synaptosomes. Competition binding experiments with [I-125]SNX-111 and [I-125]SNX-124 established that both of them bind to the same site, namely, N-type VSCC. The site detected by the binding of [I-125]SNX-230 is distinct from N-type VSCC since SNX-111 has very low affinity (K-i = 135 nM) in competition studies. Recent findings that a novel high-voltage-activated calcium channel in rat cerebellar granule neurons is resistant to blockers of L-, N-, and P-type VSCC but is highly sensitive to SNX-230 suggest that the [I-125]SNX-230 binding site may represent this novel type of calcium channel or another, as yet undescribed, VSCC. (C) 1994 Academic Press, Inc.