title: Nonequivalence of alpha-bungarotoxin binding sites in the native nicotinic receptor molecule.
abstract: In the native, membrane-bound form of the nicotinic acetylcholine receptor (M-AcChR) the two sites for the cholinergic antagonist alpha-bungarotoxin (alpha-BGT) have different binding properties. One site has high affinity, and the M-AcChR/alpha-BGT complexes thus formed dissociate very slowly, similar to the complexes formed with detergent-solubilized AcChR (S-AcChR). The second site has much lower affinity (KD approximately 59 +/- 35 nM) and forms quickly reversible complexes. The nondenaturing detergent Triton X-100 is known to solubilize the AcChR in a form unable, upon binding of cholinergic ligands, to open the ion channel and to become desensitized. Solubilization of the AcChR in Triton X-100 affects the binding properties of this second site and converts it to a high-affinity, slowly reversible site. Prolonged incubation of M-AcChR at 4 degrees C converts the low-affinity site to a high-affinity site similar to those observed in the presence of Triton X-100. Although the two sites have similar properties when the AcChR is solubilized in Triton X-100, their nonequivalence can be demonstrated by the effect on alpha-BGT binding of concanavalin A, which strongly reduces the association rate of one site only. The Bmax of alpha-BGT to either Triton-solubilized AcChR or M-AcChR is not affected by the presence of concanavalin A. Occupancy of the high-affinity, slowly reversible site in M-AcChR inhibits the Triton X-100 induced conversion to irreversibility of the second site. At difference with alpha-BGT, the long alpha-neurotoxin from Naja naja siamensis venom (alpha-NTX) binds with high affinity and in a very slowly reversible fashion to two sites in the M-AcChR (Conti-Tronconi & Raftery, 1986). We confirm here that Triton-solubilized AcChR or M-AcChR binds in a very slowly reversible fashion the same amount of alpha-NTX.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Biochemistry, College of Biological Sciences, University of Minnesota, St. Paul 55108.
authors.forename: B M
authors.initials: BM
authors.lastname: Conti-Tronconi
authors.forename: F
authors.initials: F
authors.lastname: Tang
authors.forename: S
authors.initials: S
authors.lastname: Walgrave
authors.forename: W
authors.initials: W
authors.lastname: Gallagher
mesh_terms: D000818:Animals; D001665:Binding Sites; D002038:Bungarotoxins / Q000378:metabolism*; D002498:Centrifugation / Q000379:methods; D003039:Cobra Neurotoxin Proteins / Q000378:metabolism; D003208:Concanavalin A / Q000494:pharmacology; D007457:Iodine Radioisotopes; D007700:Kinetics; D008566:Membranes / Q000648:ultrastructure; D017830:Octoxynol; D011092:Polyethylene Glycols; D011950:Receptors, Cholinergic / Q000378:metabolism; D011978:Receptors, Nicotinic / Q000378:metabolism*; D014101:Torpedo
chemical_list: D002038:Bungarotoxins; D003039:Cobra Neurotoxin Proteins; D007457:Iodine Radioisotopes; D011950:Receptors, Cholinergic; D011978:Receptors, Nicotinic; D003208:Concanavalin A; D011092:Polyethylene Glycols; C025154:alpha-cobratoxin; D017830:Octoxynol