Tris+/Na+ permeability ratios of nicotinic acetylcholine receptors are reduced by mutations near the intracellular end of the M2 region

• Published in: The Journal of general physiology Vol. 99; no. 4; pp. 545 - 572
• Main Authors: COHEN, B. N, LABARCA, C, CZYZYK, L, DAVIDSON, N, LESTER, H. A
• Format: Journal Article
• Language: English
• Published: New York, NY Rockefeller University Press 01.04.1992; The Rockefeller University Press
• Subjects: Amino Acid Sequence; Animals; Biochemistry; Biological and medical sciences; Biological Transport - physiology; Cell Membrane Permeability - physiology; Cell receptors; Cell structures and functions; Cellular biology; DNA - analysis; DNA - genetics; Fundamental and applied biological sciences. Psychology; Genetics; Mathematics; Membrane Potentials - physiology; Mice; Molecular and cellular biology; Molecular Sequence Data; Monoamines receptors (catecholamine, serotonine, histamine, acetylcholine); Mutation - genetics; Mutation - physiology; Oocytes - chemistry; Oocytes - physiology; Oocytes - ultrastructure; Receptors, Nicotinic - analysis; Receptors, Nicotinic - genetics; Receptors, Nicotinic - physiology; Sodium - pharmacokinetics; Torpedo; Tromethamine - pharmacokinetics; Xenopus; Monovalent cation; Xenopus laevis; Rodentia; Amphibia; Torpedo californica; Salientia; Selectivity; Permeability; Vertebrata; Cholinergic receptor; Mammalia; Structure activity relation; Mouse; Pisces; Toad; Mutation; Molecular cloning; Localization; Nicotinic receptor; Oocyte
• ISSN: 0022-1295; 1540-7748
• DOI: 10.1085/jgp.99.4.545

Tris+/Na+ permeability ratios were measured from shifts in the biionic reversal potentials of the macroscopic ACh-induced currents for 3 wild-type (WT), 1 hybrid, 2 subunit-deficient, and 25 mutant nicotinic receptors expressed in Xenopus oocytes. At two positions near the putative intracellular end of M2, 2' (alpha Thr244, beta Gly255, gamma Thr253, delta Ser258) and -1', point mutations reduced the relative Tris+ permeability of the mouse receptor as much as threefold. Comparable mutations at several other positions had no effects on relative Tris+ permeability. Mutations in delta had a greater effect on relative Tris+ permeability than did comparable mutations in gamma; omission of the mouse delta subunit (delta 0 receptor) or replacement of mouse delta with Xenopus delta dramatically reduced relative Tris+ permeability. The WT mouse muscle receptor (alpha beta gamma delta) had a higher relative permeability to Tris+ than the wild-type Torpedo receptor. Analysis of the data show that (a) changes in the Tris+/Na+ permeability ratio produced by mutations correlate better with the hydrophobicity of the amino acid residues in M2 than with their volume; and (b) the mole-fraction dependence of the reversal potential in mixed Na+/Tris+ solutions is approximately consistent with the Goldman-Hodgkin-Katz voltage equation. The results suggest that the main ion selectivity filter for large monovalent cations in the ACh receptor channel is the region delimited by positions -1' and 2' near the intracellular end of the M2 helix.