J. Machold et al., THE HANDEDNESS OF THE SUBUNIT ARRANGEMENT OF THE NICOTINIC ACETYLCHOLINE-RECEPTOR FROM TORPEDO-CALIFORNICA, European journal of biochemistry, 234(2), 1995, pp. 427-430
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.