The spider toxin omega-Aga IIIA defines a high affinity site on neuronal high voltage-activated calcium channels.
The spider toxin omega-agatoxin IIIA (omega-Aga-IIIA) is a potent inhibitor of high voltage-activated calcium currents in the mammalian brain. To establish the biochemical parameters governing its action, we radiolabeled the toxin and examined its binding to native and recombinant calcium channels. In experiments with purified rat synaptosomal membranes, both kinetic and equilibrium data demonstrate one-to-one binding of omega-Aga-IIIA to a single population of high affinity sites, with K(d) = approximately 9 pm and B(max) = approximately 1.4 pmol/mg protein. Partial inhibition of omega-Aga-IIIA binding by omega-conotoxins GVIA, MVIIA, and MVIIC identifies N and P/Q channels as components of this population. omega-Aga-IIIA binds to recombinant alpha(1B) and alpha(1E) calcium channels with a similar high affinity (K(d) = approximately 5-9 pm) in apparent one-to-one fashion. Results from recombinant alpha(1B) binding experiments demonstrate virtually identical B(max) values for omega-Aga-IIIA and omega-conotoxin MVIIA, providing further evidence for a one-to-one stoichiometry of agatoxin binding to calcium channels. The combined evidence suggests that omega-Aga-IIIA defines a unique, high affinity binding site on N-, P/Q-, and R-type calcium channels.
Yan
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Environmental Toxicology Graduate Program and Departments of Entomology and Neuroscience, University of California, Riverside, California 92521, USA.
Adams
M E
ME
D060848:Agatoxins; D000818:Animals; D001921:Brain / Q000378:metabolism; D002121:Calcium Channel Blockers / Q000493:pharmacokinetics*; D020864:Calcium Channels, N-Type / Q000737:chemistry / Q000378:metabolism*; D020865:Calcium Channels, P-Type / Q000737:chemistry / Q000378:metabolism; D020907:Calcium Channels, Q-Type / Q000737:chemistry / Q000378:metabolism; D020908:Calcium Channels, R-Type / Q000737:chemistry / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D006801:Humans; D007425:Intracellular Membranes / Q000378:metabolism; D007457:Iodine Radioisotopes; D007700:Kinetics; D009474:Neurons / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins / Q000378:metabolism; D013111:Spider Venoms / Q000493:pharmacokinetics*; D013112:Spiders; D013574:Synaptosomes / Q000378:metabolism*; D014162:Transfection
D060848:Agatoxins; D002121:Calcium Channel Blockers; D020864:Calcium Channels, N-Type; D020865:Calcium Channels, P-Type; D020907:Calcium Channels, Q-Type; D020908:Calcium Channels, R-Type; D007457:Iodine Radioisotopes; D011994:Recombinant Proteins; D013111:Spider Venoms; C070132:omega-agatoxin III