The arrangement of the subunits of the acetylcholine receptor of Torpedo californica.
The monomeric form of the acetylcholine receptor from torpedo is composed of five, membrane-spanning chains with the stoichiometry alpha 2 beta gamma delta. The native receptor is predominantly a dimer cross-linked by a disulfide bridge between delta chains. We reduced native dimer to monomer and generated a different dimer by diamide-induced disulfide formation specifically between beta chains. Purified beta-beta cross-linked dimer, when adsorbed to a carbon film and negatively stained, appears in the electron microscope as two contiguous disks, frequently with central, stain-filled pits; i.e. it looks like native receptor in situ viewed normal to the plane of the membrane. We take the region of closest approach of the disks to mark the portions of the beta chains involved in the cross-link. In addition, we tagged the acetylcholine binding sites (one on each alpha chain) for electron microscopic identification, using a complex of monobiotinylated cobratoxin and avidin. Based on the locations of avidins bound to the beta-beta cross-linked dimer, the two toxin binding sites/monomer appear to be separated on the average by 110 degrees, as measured between lines from the center of the monomer to the centers of the avidins. One toxin binding site appears close to the beta-beta cross-link and the other close to the end of the monomer opposite to the cross-link; these locations are similar to the locations of the toxin binding sites relative to the delta-delta cross-link in native dimer. On the assumptions that the chains are compact units and are arranged in a unique order around the central pit, we interpret these results as indicating that the alpha chains are not contiguous and that neither the beta chain nor the delta chain lies between them. Therefore, the arrangement of the chains most easily reconciled with our assumptions and observations is alpha gamma alpha beta delta.
D000818:Animals; D001360:Avidin / Q000378:metabolism; D004557:Electric Organ / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D046911:Macromolecular Substances; D008970:Molecular Weight; D011487:Protein Conformation; D011950:Receptors, Cholinergic / Q000302:isolation & purification* / Q000378:metabolism; D014101:Torpedo
D046911:Macromolecular Substances; D011950:Receptors, Cholinergic; D001360:Avidin