Pseudechetoxin: A peptide blocker of cyclic nucleotide-gated ion channels

Ion channels activated by the binding of cyclic nucleotides first were disc overed in retinal rods where they generate the cell's response to light. In other systems, however, it has been difficult to unambiguously determine w hether cyclic nucleotide-dependent processes are mediated by protein kinase s, their classical effector enzymes, or cyclic nucleotide-gated (CNG) ion c hannels. Part of this difficulty has been caused by the lack of specific ph armacological tools. Here we report the purification from the venom of the Australian King Brown snake of a peptide toxin that inhibits current throug h CNG channels, This toxin, which we have named Pseudechetoxin (PsTx), was purified by cation exchange and RP-HPLC and has a molecular mass of about 2 4 kDa, When applied to the extracellular face of membrane patches containin g the a-subunit of the rat olfactory CNG channel, PsTx blocked the cGMP-dep endent current with a Ki of 5 nM. Block was independent of voltage and requ ired only a single molecule of toxin. PsTx also blocked CNG channels contai ning the bovine rod a-subunit with high affinity (100 nhl), but it was less effective on the heteromeric version of the rod channel (K-i approximate t o 3 mu M). We have obtained N-terminal and partial internal sequence data a nd the amino acid composition of PsTx. These data indicate that PsTx is a b asic protein that exhibits some homology with helothermine a toxin isolated from the venom of the Mexican beaded lizard, PsTx promises to be a valuabl e pharmacological tool for studies on the structure and physiology of CNG c hannels.