title: Electrostatic and steric contributions to block of the skeletal muscle sodium channel by mu-conotoxin.
abstract: Pore-blocking toxins are valuable probes of ion channels that underlie electrical signaling. To be effective inhibitors, they must show high affinity and specificity and prevent ion conduction. The 22-residue sea snail peptide, mu-conotoxin GIIIA, blocks the skeletal muscle sodium channel completely. Partially blocking peptides, derived by making single or paired amino acid substitutions in mu-conotoxin GIIIA, allow a novel analysis of blocking mechanisms. Replacement of one critical residue (Arg-13) yielded peptides that only partially blocked single-channel current. These derivatives, and others with simultaneous substitution of a second residue, were used to elucidate the structural basis of the toxin's blocking action. The charge at residue-13 was the most striking determinant. A positive charge was necessary, though not sufficient, for complete block. Blocking efficacy increased with increasing residue-13 side chain size, regardless of charge, suggesting a steric contribution to inhibition. Charges grouped on one side of the toxin molecule at positions 2, 12, and 14 had a weaker influence, whereas residue-16, on the opposite face of the toxin, was more influential. Most directly interpreted, the data suggest that one side of the toxin is masked by close apposition to a binding surface on the pore, whereas the other side, bearing Lys-16, is exposed to an aqueous cavity accessible to entering ions. Strong charge-dependent effects emanate from this toxin surface. In the native toxin, Arg-13 probably presents a strategically placed electrostatic barrier rather than effecting a complete steric occlusion of the pore. This differs from other well-described channel inhibitors such as the charybdotoxin family of potassium channel blockers and the sodium channel-blocking guanidinium toxins (tetrodotoxin and saxitoxin), which appear to occlude the narrow part of the pore.
authors.affiliation: Department of Physiology and Biophysics, University of Calgary, 3330 Hospital Drive NW, Calgary, Alberta, Canada T2N 4N1.
authors.forename: Kwokyin
authors.initials: K
authors.lastname: Hui
authors.forename: Gregory
authors.initials: G
authors.lastname: Lipkind
authors.forename: Harry A
authors.initials: HA
authors.lastname: Fozzard
authors.forename: Robert J
authors.initials: RJ
authors.lastname: French
mesh_terms: D000595:Amino Acid Sequence; D002121:Calcium Channel Blockers / Q000737:chemistry* / Q000494:pharmacology*; D020916:Conotoxins / Q000737:chemistry* / Q000494:pharmacology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D018482:Muscle, Skeletal / Q000187:drug effects* / Q000502:physiology; D026941:Sodium Channel Blockers; D015222:Sodium Channels / Q000502:physiology; D055672:Static Electricity; D013237:Stereoisomerism
chemical_list: D002121:Calcium Channel Blockers; D020916:Conotoxins; D026941:Sodium Channel Blockers; D015222:Sodium Channels