Nicotinic acetylcholine receptor from foetal human skeletal muscle

Nicotinic acetylcholine receptor protein has been purified from foetal and adult skeletal muscle by extraction in non-ionic detergent followed by purification on immobilised α-toxin. Purified foetal and adult receptors focused as single, sharp peaks whether directly labelled with (125)I or indirectly labelled with (125)I-α-bungarotoxin. Polyacrylamide gel electrophoresis of the purified foetal and adult receptors each showed four major protein bands with Mr 44,000, 51,000, 58,000 and 66,000; only that with Mr 44,000 was, in each case, labelled with the affinity reagent, 4-(N-maleimido) [(3)H]benzyltrimethylammonium. When the four major subunits, obtained by polyacrylamide gel electrophoresis, were labelled with (125)I-ConA, markedly different patterns of radioactivity were shown by the foetal and adult receptors, the band at 44,000 being less heavily labelled in the foetal case. Foetal and adult receptors behaved similarly with respect to inhibition by ConA of binding of (125)I-α-bungarotoxin; inhibition in both cases reaching a maximum of 70%. Foetal and adult receptors each showed single (125)I-α-bungarotoxin binding species in sucrose density gradient centrifugation with S20w = 8.5Sand 9.5S, respectively, although the former peak was broader, possibly reflecting the relative instability of the purified foetal receptor. Our finding of marked differences in the glycosylation of foetal and adult human acetylcholine receptors suggest that, in otherwise very similar proteins, the carbohydrate moieties could determine the known differences in location and stability of the two receptor types.