Two peptides, ProTx-I and ProTx-II, from the venom of the tarantula
Thrixopelma pruriens,have been isolated and characterized. These peptides were purified on the basis of their ability to reversiblyinhibit the tetrodotoxin-resistant Na channel, Na
V 1.8, and are shown to belong to the inhibitory cystineknot (ICK) family of peptide toxins interacting with voltage-gated ion channels. The family has severalhallmarks: cystine bridge connectivity, mechanism of channel inhibition, and promiscuity across channelswithin and across channel families. The cystine bridge connectivity of ProTx-II is very similar to that ofother members of this family, i.e., C
2 to C
16, C
9 to C
21, and C
15 to C
25. These peptides are the first high-affinity ligands for tetrodotoxin-resistant peripheral nerve Na
V channels, but also inhibit other Na
V channels(IC
50's < 100 nM). ProTx-I and ProTx-II shift the voltage dependence of activation of Na
V 1.5 to morepositive voltages, similar to other gating-modifier ICK family members. ProTx-I also shifts the voltagedependence of activation of Ca
V 3.1 (
1G, T-type, IC
50 = 50 nM) without affecting the voltage dependenceof inactivation. To enable further structural and functional studies, synthetic ProTx-II was made; it adoptsthe same structure and has the same functional properties as the native peptide. Synthetic ProTx-I wasalso made and exhibits the same potency as the native peptide. Synthetic ProTx-I, but not ProTx-II, alsoinhibits K
V 2.1 channels with 10-fold less potency than its potency on Na
V channels. These peptidesrepresent novel tools for exploring the gating mechanisms of several Na
V and Ca
V channels.