Molecular analysis of voltage-gated α-Na⁺ channel toxin binding site 3 to determine key residues that confer resistance to α-scorpion toxin

Includes bibliographical references.Voltage-gated Na⁺ channels (Nav1.2) which are proteins that posses 4 (I-IV) domains and six transmembrane (S1-S6) helices in each domain display abnormally slowed fast inactivation when an α-scorpion toxin binds to it. However, voltage gated Na⁺ channels of the Mesobuthus martensii Karsch (MmK) scorpion that produces α-scorpion toxins are resistant to their toxins. The previously described α-toxin binding site 3 of the MmK voltage-gated Na⁺ channels were examined by a comparative method using an alignment of many sodium channel protein sequences (multi-alignment) generated using the rat Nav1.2 α-toxin sequence of binding site 3 as a template for the alignment. The α-toxin binding site 3 is comprised of 4 regions or segments and found in domains I and IV of the Na⁺ channel. An analysis suggested that key residues exist that presumably are essential for the α-scorpion toxin binding at each of the four segments of the toxin binding site 3. The most significant mutations either altered the charge distribution and or polarity of the binding region or likely introduced changes in secondary structure (introduction of a turn). Mutations observed in the first segment of domain I S5-S6 that correspond to protein sequence positions in the rat and scorpion Na⁺ channel (rat/Mmk) at K355/S339, A356/N340, -/Y341 (- refers to a missing residue), R358/P343, Y362/H347, and G371-S371/Y356 were the most significant. Significant mutations in the second segment of domain I S5-S6 were evident at K399/P384 and T400/W385. The fourth segment of domain IV S5-S6 had significant mutations at K1687/H1599, E168/R1600, and M1694/N1606. The third segment of domain IV S3-S4 was suspected of having significant mutations at E1616/A1528 and K1617/S1529. Changes in these key resides may introduce resistance to binding and action of α-scorpion toxins to the Na⁺ channel. Future experimentation and or analysis will be needed to determine which of these residues are the most important for binding of the toxin to the channel.B.S. (Bachelor of Science