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Mapping the interaction site for a ?-scorpion toxin in the pore module of domain III of voltage-gated Na(+) channels.

ABSTRACT: Activation of voltage-gated sodium (Na(v)) channels initiates and propagates action potentials in electrically excitable cells. ?-Scorpion toxins, including toxin IV from Centruroides suffusus suffusus (CssIV), enhance activation of Na(V) channels. CssIV stabilizes the voltage sensor in domain II in its activated state via a voltage-sensor trapping mechanism. Amino acid residues required for the action of CssIV have been identified in the S1-S2 and S3-S4 extracellular loops of domain II. The extracellular loops of domain III are also involved in toxin action, but individual amino acid residues have not been identified. We used site-directed mutagenesis and voltage clamp recording to investigate amino acid residues of domain III that are involved in CssIV action. In the IIISS2-S6 loop, five substitutions at four positions altered voltage-sensor trapping by CssIV(E15A). Three substitutions (E1438A, D1445A, and D1445Y) markedly decreased voltage-sensor trapping, whereas the other two substitutions (N1436G and L1439A) increased voltage-sensor trapping. These bidirectional effects suggest that residues in IIISS2-S6 make both positive and negative interactions with CssIV. N1436G enhanced voltage-sensor trapping via increased binding affinity to the resting state, whereas L1439A increased voltage-sensor trapping efficacy. Based on these results, a three-dimensional model of the toxin-channel interaction was developed using the Rosetta modeling method. These data provide additional molecular insight into the voltage-sensor trapping mechanism of toxin action and define a three-point interaction site for ?-scorpion toxins on Na(V) channels. Binding of ?- and ?-scorpion toxins to two distinct, pseudo-symmetrically organized receptor sites on Na(V) channels acts synergistically to modify channel gating and paralyze prey.

SUBMITTER: Zhang JZ

PROVIDER: S-EPMC3436316 | biostudies-literature | 2012 Aug

REPOSITORIES: biostudies-literature

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Mapping the interaction site for a β-scorpion toxin in the pore module of domain III of voltage-gated Na(+) channels.

Zhang Joel Z JZ Yarov-Yarovoy Vladimir V Scheuer Todd T Karbat Izhar I Cohen Lior L Gordon Dalia D Gurevitz Michael M Catterall William A WA

The Journal of biological chemistry 20120702 36

Activation of voltage-gated sodium (Na(v)) channels initiates and propagates action potentials in electrically excitable cells. β-Scorpion toxins, including toxin IV from Centruroides suffusus suffusus (CssIV), enhance activation of Na(V) channels. CssIV stabilizes the voltage sensor in domain II in its activated state via a voltage-sensor trapping mechanism. Amino acid residues required for the action of CssIV have been identified in the S1-S2 and S3-S4 extracellular loops of domain II. The ext ...[more]

PMID: 22761417

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Project description:Voltage-gated sodium channel NaV1.8 regulates transmission of pain signals to the brain. While NaV1.8 has the potential to serve as a drug target, the molecular mechanisms that shape NaV1.8 gating are not completely understood, particularly mechanisms that couple activation to inactivation. Interactions between toxin producing animals and their predators provide a novel approach for investigating NaV structure-function relationships. Arizona bark scorpions produce Na+ channel toxins that initiate pain signaling. However, in predatory grasshopper mice, toxins inhibit NaV1.8 currents and block pain signals. A screen of synthetic peptide toxins predicted from bark scorpion venom showed that peptide NaTx36 inhibited Na+ current recorded from a recombinant grasshopper mouse NaV1.8 channel (OtNaV1.8). Toxin NaTx36 hyperpolarized OtNaV1.8 activation, steady-state fast inactivation, and slow inactivation. Mutagenesis revealed that the first gating charge in the domain I (DI) S4 voltage sensor and an acidic amino acid (E) in the DII SS2 - S6 pore loop are critical for the inhibitory effects of NaTx36. Computational modeling showed that a DI S1 - S2 asparagine (N) stabilizes the NaTx36 - OtNaV1.8 complex while residues in the DI S3 - S4 linker and S4 voltage sensor form electrostatic interactions that allow a toxin glutamine (Q) to contact the first S4 gating charge. Surprisingly, the models predicted that NaTx36 contacts amino acids in the DII S5 - SS1 pore loop instead of the SS2 - S6 loop; the DII SS2 - S6 loop motif (QVSE) alters the conformation of the DII S5 - SS1 pore loop, enhancing allosteric interactions between toxin and the DII S5 - SS1 pore loop. Few toxins have been identified that modify NaV1.8 gating. Moreover, few toxins have been described that modify sodium channel gating via the DI S4 voltage sensor. Thus, NaTx36 and OtNaV1.8 provide tools for investigating the structure-activity relationship between channel activation and inactivation gating, and the connection to alternative pain phenotypes.

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Dataset Information

Mapping the interaction site for a ?-scorpion toxin in the pore module of domain III of voltage-gated Na(+) channels.

Publications

Mapping the interaction site for a β-scorpion toxin in the pore module of domain III of voltage-gated Na(+) channels.

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