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P Pappone

Publications and source records attributed to P Pappone.

3 recordsLinked to original sources
PubMed1994

Mutations in the M4 domain of Torpedo californica acetylcholine receptor dramatically alter ion channel function.

Site-directed mutagenesis was used to mutate alpha Cys418 and beta Cys447 in the M4 domain of Torpedo californica acetylcholine receptor expressed in Xenopus laevis oocytes. The M4 region is a transmembrane domain thought to be located at the lipid-protein interface. By whole-cell voltage clamp analysis, mutation of both alpha subunits to alpha Trp418 increased maximal channel activity approximately threefold, increased the desensitization rate compared with wild-type receptor, and shifted the EC50 for acetylcholine from 32 microM to 13 microM. Patch measurements of single-channel currents revealed that the alpha Trp418 increased channel open times approximately 28-fold at 13 degrees C with no effect on channel conductance. All of our measured functional changes in the alpha Trp418 mutant are consistent with a simple kinetic model of the acetylcholine receptor in which only the channel closing rate is altered by the mutation. Our results show that changes in protein structure at the putative lipid-protein interface can dramatically affect receptor function.

Acetylcholine↗
PubMed1991

Effects of pH on acetylcholine receptor function.

We have examined the effects of changing extracellular pH on the function of nicotinic acetylcholine receptors from Torpedo californica using ion flux and electrophysiological methods. Agonist-induced cation efflux from vesicles containing purified, reconstituted receptors showed a monotonic dependence on external hydrogen ion concentration with maximal fluxes at alkaline pH and no agonist-induced efflux at pH's less than approximately 5. A similar pH dependence was measured for the peak agonist-activated membrane currents measured in microelectrode voltage-clamped Xenopus oocytes induced to express Torpedo receptor through mRNA injection. Half-maximal inhibition occurred at a similar pH in both systems, in the range of pH 6.5-7.0. Single-channel currents from Torpedo ACh receptors measured in patch-clamp recordings were also reduced in amplitude at acid pH with an apparent pKa for block of less than 5. Measurements of channel kinetics had a more complicated dependence on pH. The mean channel open time determined from patch-clamp measurements was maximal at neutral pH and decreased at both acid and alkaline pH's. Thus, both channel permeability properties and channel gating properties are affected by the extracellular pH.

Animals↗