Differential labeling of the alpha and beta 1 subunits of the sodium channel by photoreactive derivatives of scorpion toxin.
The separation of two photoreactive derivatives of the alpha-scorpion toxin from Leiurus quinquestriatus is described. When the two photoreactive derivatives were photolyzed separately in the presence of brain membranes containing voltage-sensitive sodium channels, one labeled the alpha subunit preferentially while the other labeled beta 1 more intensely than alpha. Batrachotoxin enhanced the efficiency of covalent labeling by the photoreactive derivatives of scorpion toxin. In all the labeling experiments, the specific incorporation of radioactive scorpion toxin was eliminated by an excess of nonradioactive scorpion toxin. The alpha polypeptide labeled in synaptosomes by photoreactive scorpion toxin was demonstrated by immunological techniques to be the same large polypeptide identified in sodium channels purified by their saxitoxin binding activity. The alpha and beta 1 subunits were detected by rapid photoaffinity labeling of a freshly prepared brain homogenate in the presence of a mixture of nine protease inhibitors, indicating that they are components of the sodium channel in intact brain tissue. The association of the covalently labeled polypeptides with the membrane was investigated by treatment of labeled synaptosomes with various agents known to remove proteins only indirectly attached to the lipid bilayer via a membrane-bound protein. In all cases, both the alpha and the beta 1 polypeptides remained in the membrane fraction following extraction. This confirms earlier proposals that the alpha polypeptide has a portion of its mass embedded within the lipid bilayer and suggests that the beta 1 polypeptide does as well.
D000345:Affinity Labels / Q000378:metabolism*; D000818:Animals; D001496:Batrachotoxins / Q000494:pharmacology; D001921:Brain / Q000648:ultrastructure; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D007163:Immunosorbent Techniques; D007473:Ion Channels / Q000378:metabolism*; D010777:Photochemistry; D051381:Rats; D011919:Rats, Inbred Strains; D012604:Scorpion Venoms / Q000302:isolation & purification / Q000378:metabolism*; D012964:Sodium / Q000378:metabolism*; D013570:Synaptic Membranes / Q000378:metabolism; D013574:Synaptosomes / Q000378:metabolism
D000345:Affinity Labels; D001496:Batrachotoxins; D007473:Ion Channels; D012604:Scorpion Venoms; D012964:Sodium