title: Variations in receptor site-3 on rat brain and insect sodium channels highlighted by binding of a funnel-web spider delta-atracotoxin.
abstract: Delta-atracotoxins (delta-ACTXs) from Australian funnel-web spiders differ structurally from scorpion alpha-toxins (Sc(alpha)Tx) but similarly slow sodium current inactivation and compete for their binding to sodium channels at receptor site-3. Characterization of the binding of 125I-labelled delta-ACTX-Hv1a to various sodium channels reveals a decrease in affinity for depolarized (0 mV; Kd=6.5 +/- 1.4 nm) vs.polarized (-55 mV; Kd=0.6 +/- 0.2 nm) rat brain synaptosomes. The increased Kd under depolarized conditions correlates with a 4.3-fold reduction in the association rate and a 1.8-increase in the dissociation rate. In comparison, Sc(alpha)Tx binding affinity decreased 33-fold under depolarized conditions due to a 48-fold reduction in the association rate. The binding of 125I-labelled delta-ACTX-Hv1a to rat brain synaptosomes is inhibited competitively by classical Sc(alpha)Txs and allosterically by brevetoxin-1, similar to Sc(alpha)Tx binding. However, in contrast with classical Sc(alpha)Txs, 125I-labelled delta-ACTX-Hv1a binds with high affinity to cockroach Na+ channels (Kd=0.42 +/- 0.1 nm) and is displaced by the Sc(alpha)Tx, Lqh(alpha)IT, a well-defined ligand of insect sodium channel receptor site-3. However, delta-ACTX-Hv1a exhibits a surprisingly low binding affinity to locust sodium channels. Thus, unlike Sc(alpha)Txs, which are capable of differentiating between mammalian and insect sodium channels, delta-ACTXs differentiate between various insect sodium channels but bind with similar high affinity to rat brain and cockroach channels. Structural comparison of delta-ACTX-Hv1a to Sc(alpha)Txs suggests a similar putative bioactive surface but a 'slimmer' overall shape of the spider toxin. A slimmer shape may ease the interaction with the cockroach and mammalian receptor site-3 and facilitate its association with different conformations of the rat brain receptor, correlated with closed/open and slow-inactivated channel states.
authors.affiliation: CEA, Dèpartement d'Ingènierie et d'Etudes des Protèines, Gif-sur-Yvette, France.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Gilles
authors.forename: Greg
authors.initials: G
authors.lastname: Harrison
authors.forename: Izhar
authors.initials: I
authors.lastname: Karbat
authors.forename: Michael
authors.initials: M
authors.lastname: Gurevitz
authors.forename: Graham M
authors.initials: GM
authors.lastname: Nicholson
authors.forename: Dalia
authors.initials: D
authors.lastname: Gordon
mesh_terms: D000494:Allosteric Regulation; D000818:Animals; D001921:Brain / Q000378:metabolism*; D016764:Cell Polarity; D003058:Cockroaches; D006110:Grasshoppers; D007700:Kinetics; D008564:Membrane Potentials; D051381:Rats; D015222:Sodium Channels / Q000378:metabolism*; D013111:Spider Venoms / Q000378:metabolism*; D013574:Synaptosomes / Q000378:metabolism
chemical_list: D015222:Sodium Channels; D013111:Spider Venoms; C406870:delta-atracotoxin-HV1