(full paper is archived in the Miller Library)
Title: Tityustoxin-Ka block of K+ current in giant fiber lobe neurons of Loligo opalescens is pH dependent
Student Author(s): Pittson, Sky
Faculty Advisor(s): Gilly, William F.
Location: Final Papers Biology 175H
Date: June 1996
Abstract: Tityustoxin-Ka (TsTX-Ka) is a 37 amino acid peptide component of the venom of the scorpion, Tityus serrulatus, and blocks voltage-gated K+ current (Ik) in highly specific channel types. TsTX-Ka is structurally similar to charybdotoxin, agitoxin-2, and other scorpion toxins that bind to the extracellular surface of the pore-forming region of Kv1-type (Shaker) channels. The action of TsTX-Ka was studied on squid K+ channels using the whole-cell patch clamp technique with dissociated giant fiber lobe (GFL) neurons and with inscet Sf9 cells infected with a baculovirus carrying the cDNA for the corresponding channel (Brock et al., 1996). Under standard conditions (12 C, pH 7.6, 20 mM external K+), TsTX-Ka reduces Ik for a strong voltage step (+40 mV) in GFL neurons with an estimated Kd of 50 nM. Steady state voltage dependence and activation kinetics of residual Ik are minimally affected in 300 nM TsTX-Ka, but inactivation kinetics are accelerated. Similar results were obtained in SF9 cells. Block of Ik in GFL neurons by TsTX-ka (300 nM) depends on external pH. Less block occurs at low pH, with half-block occurring at pH 6.0, approximately the pKa of histidine (6.02). This amino acid is located in the SqKv1A channel (H351) at a position homologous to F425 in Shaker B, a residue critical to binding of charybdotoxin (Goldstein, et al., 1994) and agitoxin-2 (Gross & MacKinnon, 1996). These results suggest that H351 in the external mouth of the squid SqKv1A channel is also a determinant of TsTX-ka binding, and that protonation of the side chain of this residue may decrease toxin sensitivity by electrostatic repulsion.