Structural Bioinformatics Research On The Diverse Animal Toxin-potassium Channel Interactions

For their survival, scorpions, snakes, bees, spiders, sea anemones and cone snails have developed efficient venoms for prey capture and their defense against predators. These venoms contain peptide toxins that target potassium channels which represent the largest and most diverse group of ion channels, including voltage-gated potassium channels, inward-rectifier potassium channels, small conductance Ca2+-activated potassium channels, intermediate conductance Ca2+-activated potassium channels, large conductance Ca2+-activated potassium channels, and so on. These toxins are useful pharmacological tools to probe the structure and function of potassium channels. In addition, they might represent potential peptide drugs to treating potassium channel-associated diseases.First, we investigate the unique mechanism of the interaction between honey bee toxin TPNQ and rKir1.1potassium channel through structural bioinformatics. Inward rectifier potassium channels (Kir channels) are an important class of potassium channels, they serve a variety of important physiological functions. The21-residue compact tertiapin-Q (TPNQ) toxin, a derivative of honey bee toxin tertiapin (TPN), is a potent blocker of inward-rectifier K+channel subtype, rat Kir1.1(rKir1.1) channel, and their interaction mechanism remains unclear. Based on the flexible feature of potassium channel turrets, a good starting rKir1.1channel structure was modeled for the accessibility of rKir1.1channel turrets to TPNQ toxin. In combination with experimental alanine scanning mutagenesis data, computational approaches were further used to obtain a reasonable TPNQ toxin-rKir1.1channel complex structure, which was completely different from the known binding modes between animal toxins and potassium channels. TPNQ toxin mainly adopted its helical domain as the channel-interacting surface together with His12as the pore-blocking residue. The important GIn13residue mainly contacted channel residues near the selectivity filter, and Lys20residue was surrounded by a polar "groove" formed by Arg118, Thr119, Glu123, and Asn124in the channel turret. On the other hand, four turrets of rKirl.1channel gathered to form a narrow pore entryway for TPNQ toxin recognition. The Phel46and Phe148residues in the channel pore region formed strong hydrophobic protrusions, and produced dominant nonpolar interactions with toxin residues. These specific structure features of rKir1.1channel vestibule well matched the binding of potent TPNQ toxin, and likely restricted the binding of the classical animal toxins. Based on the the TPNQ toxin-rKir1.1channel complex structure, we further investigate the structural basis of TPNQ toxin selectivity for Kir channel subtypes. Therefore, the TPNQ toxin-rKir1.1channel complex structure not only revealed their unique interaction mechanism, but also would highlight the diverse animal toxin-potassium channel interactions.Second, we investigate the role of animal toxin acidic residue polymorphism in the diversity of toxin-potassium channel interactions.Venomous animals, such as scorpions, snakes, sea anemones and cone snails, developed potassium channel-blocking toxins with a variety of3D structures. At the molecular level, various modes of interactions between toxins and potassium channels have been described, but the intimate molecular basis remains poorly understood. Here, we report the functional importance of the polymorphism in animal toxin acidic residues. To investigate this question, we selected BmKTX, a natural37-residue scorpion toxin with two native acidic Asp19and Asp33residues. Through adjusting the distribution of these acidic residues, we designed three closely related analogs of BmKTX with a polymorphism in the distribution of acidic residues. The wild-type BmKTX and its analogs (BmKTX-D33H, BmKTX-D19K and BmKTX-D19K/K6D) have similar3D structures and block Kv1.3channel currents with respective IC50values of0.091nM,0.015nM,0.375nM and7.3nM. Alanine scanning mutagenesis and computational simulations on these toxins further show that the four peptides adopt dissimilar binding interfaces to recognize Kvl.3channel as highlighted also by the involvement of different critical channel pore-blocking residues (Arg23in BmKTX, Lys26in BmKTX-D33H, Lys8in BmKTX-D19K and Lys15in BmKTX-D19K/K6D). By swapping the position of these acid residues, we identified four distinct modes of peptide-Kvl.3channel interactions. These findings illustrate the'evolutionary' function of the polymorphism in toxin acidic residue distribution in setting the diversity of toxin-potassium channel interactions. In addition, these observations define an innovative strategy for generating new peptide drugs by simply altering acidic residue distribution in toxin amino acid sequences.In conclusion, we not only revealed the novel molecular mechanisms of the diverse toxin-potassium channel interactions, but also further exhibited its prospect in the discovery of peptide drugs targeting potassium channels.