To gain success in the evolutionary “arms race”, venomous animals such as scorpions produce diverse neurotoxins selected to hit targets in the nervous system of prey. Scorpion α-toxins affect insect and/or mammalian voltage-gated sodium channels (Nav’s) and thereby modify the excitability of muscle and nerve cells. Although more than a hundred α-toxins are known and a number of them have been studied into detail, the molecular mechanism of their interaction with Nav’s is still poorly understood. Here, we employ extensive molecular dynamics simulations and spatial mapping of hydrophobic/hydrophilic properties distributed over the molecular surface of α-toxins. It is revealed that in spite of the small size and relatively rigid structure, these toxins possess modular organization from structural, functional and evolutionary perspectives. The more conserved and rigid “core module” is supplemented with the “specificity module” (SM) that is comparatively flexible and variable, and determines the taxon (mammal vs. insect) specificity of α-toxin activity. We further show that SMs in mammal toxins are more flexible and hydrophilic than in insect toxins. Concomitant sequence-based analysis of Nav’s extracellular loops suggests that α-toxins recognize the channels using both modules. We propose that the core module binds to the voltage-sensing domain of repeat IV, whereas the more versatile SM interacts with the pore domain in repeat I of Nav’s. These findings corroborate and expand the hypothesis on different functional epitopes of toxins that has been reported previously. In effect, we propose that the modular structure in toxins evolved to match the domain architecture of Nav’s.
The elastin binding protein (EBP), a spliced variant of lysosomal β-galactosidase, is the primary receptor of elastin peptides that have been linked to emphysema, aneurysm and cancer progression. The sequences recognized by EBP share the XGXXPG consensus pattern found in numerous matrix proteins, notably in elastin where the VGVAPG motif is repeated. To delineate the elastin binding site of human EBP, we built a homology model of this protein and docked VGVAPG on its surface. Analysis of this model suggested that Gln-97 and Asp-98 were required for interaction with VGVAPG because they contribute to the definition of a pocket thought to represent the elastin binding site of EBP. Additionally, we proposed that Leu-103, Arg-107, and Glu-137 were essential residues because they could interact with VGVAPG itself. Site-directed mutagenesis experiments at these key positions validated our model. This work therefore provides the first structural data concerning the interaction of the VGVAPG with its cognate receptor. The present structural data should now allow the development of EBP-specific antagonists.
Tk-hefu is an artificial peptide designed based on the α-hairpinin scaffold, which selectively blocks voltage-gated potassium channels Kv1.3. Here we present its spatial structure resolved by NMR spectroscopy and analyze its interaction with channels using computer modeling. We apply protein surface topography to suggest mutations and increase Tk-hefu affinity to the Kv1.3 channel isoform. We redesign the functional surface of Tk-hefu to better match the respective surface of the channel pore vestibule. The resulting peptide Tk-hefu-2 retains Kv1.3 selectivity and displays ∼15 times greater activity compared with Tk-hefu. We verify the mode of Tk-hefu-2 binding to the channel outer vestibule experimentally by site-directed mutagenesis. We argue that scaffold engineering aided by protein surface topography represents a reliable tool for design and optimization of specific ion channel ligands.