alpha subunit compositions of Kv1.1-containing K+ channel subtypes fractionated from rat brain using dendrotoxins.
K+ channels from the Kv1 subfamily contain four alpha-subunits and the combinations (from Kv1.1-1.6) determine susceptibility to dendrotoxin (DTX) homologues. The subunit composition of certain subtypes in rat brain was investigated using DTXk which only interacts with Kv1.1-containing channels and alphaDTX (and its closely related homologue DTXi) that binds preferentially to Kv1. 2-possessing homo- or hetero-oligomers. Covalent attachment of [125I]DTXk bound to channels in synaptic membranes unveiled subunits of Mr = 78 000 and 96 000. Immunoprecipitation of these solubilized and dissociated cross-linked proteins with IgG specific for each of the alpha-subunits identified Kv1.1, 1.2 and 1.4; this led to assemblies of Kv1.1/1.2 and 1.1/1.4 being established. Kv1. 2-enriched channels, purified from rat brain by chromatography on immobilized DTXi, contained Kv1.1, 1.2 and 1.6 confirming one of the above-noted pairs and indicating an additional Kv1.1-containing oligomer (Kv1.1/1.2/1.6); the notable lack of Kv1.4 excludes a Kv1. 1/1.2/1.4 combination. On the other hand, channels with Kv1.1 as a constituent, isolated using DTXk, possessed Kv1.4 in addition to those found in the DTXi-purified oligomers; this provides convergent support for the occurrence of the three combinations established above but adds a possible fourth (Kv1.1/1.4/1.6), though this was not confirmed. Moreover, sequential purification on DTXi and DTXk resins yielded channels containing only Kv1.1/1.2 but with an apparent predominance of Kv1.1, reaffirming the latter multimer.
Wang
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Department of Biochemistry, Imperial College, London, UK.
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Dolly
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D000818:Animals; D001923:Brain Chemistry; D002846:Chromatography, Affinity; D003432:Cross-Linking Reagents; D004546:Elapid Venoms / Q000378:metabolism; D007700:Kinetics; D051662:Kv1.1 Potassium Channel; D008970:Molecular Weight; D010455:Peptides / Q000378:metabolism; D015221:Potassium Channels / Q000737:chemistry* / Q000302:isolation & purification* / Q000378:metabolism; D024642:Potassium Channels, Voltage-Gated; D011233:Precipitin Tests; D011487:Protein Conformation; D051381:Rats; D013570:Synaptic Membranes / Q000737:chemistry
D003432:Cross-Linking Reagents; D004546:Elapid Venoms; D010455:Peptides; D015221:Potassium Channels; D024642:Potassium Channels, Voltage-Gated; C081506:dendrotoxin K; D051662:Kv1.1 Potassium Channel; C020273:dendrotoxin