Characterization of acetylcholine receptor isolated from Torpedo californica electroplax through the use of an easily removable detergent, beta-D-octylglucopyranoside.
Non-ionic detergents used for the solubilization and purification of acetylcholine receptor from Torpedo californica electroplax may remain tightly bound to this protein. The presence of detergent greatly hinders spectrophotometric and hydrodynamic studies of the receptor protein. beta-D-Octylglucopyranoside, however, is found to be effective in solubilizing the receptor from electroplax membranes with minimal interference in the characterization of the protein. The acetylcholine receptor purified from either octylglucopyranoside- or Triton X-100-solubilized extracts exhibits identical amino acid compositions, alpha-Bungarotoxin and (+)-tubocurarine binding parameters, and subunit distributions in SDS-polyacrylamide gels. The use of octylglucopyranoside allows for the assignment of a molar absorptivity for the purified receptor at 280 nm of approx. 530000 M-1 . cm-1. Additionally, successful reconstitution of octylglucopyranoside-extracted acetylcholine receptor into functional membrane vesicles has recently been achieved (Gonzales-Ros, J.M., Paraschos, A. and Martinez-Carrion, M. (1980) Proc. Natl. Acad. Sci. U.S.A. 77, 1796--1799). Removal of octylglucopyranoside by dialysis does not alter the specific toxin and antagonist binding ability of the receptor or its solubility at low protein concentrations. Sedimentation profiles of the purified acetylcholine receptor in sucrose density gradients reveal several components. Sedimentation coefficients obtained for the slowest sedimenting species agree with previously reported molecular weight values. Additionally, the different sedimenting forms exhibit distinctive behavior in isoelectric focusing gels. Our results suggest that both the concentration and type of detergent greatly influence the physicochemical behavior of the receptor protein.
D000818:Animals; D003902:Detergents; D004557:Electric Organ / Q000378:metabolism*; D005399:Fishes; D005960:Glucosides; D007700:Kinetics; D046911:Macromolecular Substances; D008970:Molecular Weight; D011950:Receptors, Cholinergic / Q000302:isolation & purification* / Q000378:metabolism; D012995:Solubility
D003902:Detergents; D005960:Glucosides; D046911:Macromolecular Substances; D011950:Receptors, Cholinergic; C018619:octyl-beta-D-glucoside