THE HANDEDNESS OF THE SUBUNIT ARRANGEMENT OF THE NICOTINIC ACETYLCHOLINE-RECEPTOR FROM TORPEDO-CALIFORNICA

Cross-linking an alpha-neurotoxin with a known three-dimensional struc ture and with photoactivatable groups in known positions to native mem brane-bound acetylcholine receptor reveals its quaternary structure, i ncluding the handedness of its circular subunit arrangement. Photolabe lling with alpha-neurotoxin carrying the photoactivatable group at pos ition Lys46 is inhibited by the competitive antagonist (+)-tubocurarin e in a biphasic manner, indicating that it reacts with both alpha-subu nits that were shown to have different affinities for this antagonist [Neubig, R. R. & Cohen, J. B. (1979) Biochemistry 18, 5464-5475]. Lys4 6 is located on loop III of the neurotoxin. The other information nece ssary for the elucidation of the handedness was provided by the recent finding that the central loop of the toxin (loop II) is orientated to wards the central pore of the receptor, securing the overall orientati on of the bound toxin [Machold, J., Utkin, Y. N., Kirsch, D., Kaufmann , R., Tsetlin, V. & Hucho, F. (1995b) Proc. Natl Acad. Sci. USA 92, 72 82-7286]. Looking at the receptor from the synaptic side of the postsy naptic membrane, it was concluded that the clockwise subunit arrangeme nt is alpha(H)-gamma-alpha(L)-delta-beta (alpha(H) and alpha(L) are th e alpha-subunits binding (+)-tubocurarine with high and low affinity, respectively). Its mirror image alpha(L)-gamma-alpha(H)-beta-delta cou ld thus be excluded.