Alkylation of free sulfhydryls fortifies electroplax subsynaptic structures.
The cysteine-rich 43,000-dalton peripheral membrane protein, nu 1, is localized at the cytoplasmic face of electroplax and muscle cholinergic synapses, where it is thought to play an important role in the endplate supramolecular structure. The peripheral membrane protein properties of nu 1 are inferred by its removal from nicotinic cholinergic membranes by the action of mild alkali or lithium diiodosalicylate. An interesting property of nu 1 is its high concentration of free sulfhydryl groups, whose exact role in synaptic structure is still largely unknown. Alkylation of free sulfhydryls with N-ethylmaleimide (3 mM) has a profound effect on the association of nu 1 with synaptic membranes, rendering nu 1 unextractable by pH 11 treatment or by lithium diiodosalicylate and, concomitantly, decreasing nu 1's electrophoretic mobility on sodium dodecyl sulfate-polyacrylamide gels. Iodoacetamide and iodoacetate have similar effects, but at concentrations 10- to 100-fold higher than required for N-ethylmaleimide. Furthermore, sulfhydryl modification also stabilizes the association of nicotinic receptor subunits with the detergent-insoluble cytoskeleton. N-Ethylmaleimide treatment increases the fraction of insoluble receptor molecules on extraction with Triton X-100, sodium cholate, or octylglucoside. These results suggest an important role of sulfhydryl groups in the structural stability of the postsynaptic cholinergic membrane.
D000478:Alkylation; D000818:Animals; D002458:Cell Fractionation / Q000379:methods; D002462:Cell Membrane / Q000187:drug effects / Q000648:ultrastructure; D003902:Detergents; D004492:Edetic Acid / Q000494:pharmacology; D004533:Egtazic Acid / Q000494:pharmacology; D004557:Electric Organ / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure*; D005033:Ethylmaleimide / Q000494:pharmacology*; D008565:Membrane Proteins / Q000378:metabolism*; D008970:Molecular Weight; D011978:Receptors, Nicotinic / Q000187:drug effects / Q000378:metabolism; D013438:Sulfhydryl Compounds / Q000378:metabolism*; D013569:Synapses / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D014101:Torpedo
D003902:Detergents; D008565:Membrane Proteins; D011978:Receptors, Nicotinic; D013438:Sulfhydryl Compounds; D004533:Egtazic Acid; D004492:Edetic Acid; D005033:Ethylmaleimide