Role of negatively charged amino acids in beta 4 F-loop in activation and desensitization of alpha 3 beta 4 rat neuronal nicotinic receptors
Language English Country Netherlands Media print-electronic
Document type Journal Article, Research Support, Non-U.S. Gov't
PubMed
18249185
DOI
10.1016/j.bbamem.2008.01.010
PII: S0005-2736(08)00038-2
Knihovny.cz E-resources
- MeSH
- Amino Acids chemistry metabolism MeSH
- Bridged Bicyclo Compounds, Heterocyclic pharmacology MeSH
- Chlorocebus aethiops MeSH
- COS Cells MeSH
- Kinetics MeSH
- Rats MeSH
- Molecular Sequence Data MeSH
- Mutation genetics MeSH
- Neurons metabolism MeSH
- Nicotine pharmacology MeSH
- Receptors, Nicotinic chemistry metabolism MeSH
- Nicotinic Agonists pharmacology MeSH
- Protein Subunits chemistry metabolism MeSH
- Pyridines pharmacology MeSH
- Protein Structure, Secondary MeSH
- Amino Acid Sequence MeSH
- Sequence Alignment MeSH
- Tubocurarine pharmacology MeSH
- Structure-Activity Relationship MeSH
- Animals MeSH
- Check Tag
- Rats MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Names of Substances
- Amino Acids MeSH
- Bridged Bicyclo Compounds, Heterocyclic MeSH
- epibatidine MeSH Browser
- Nicotine MeSH
- Receptors, Nicotinic MeSH
- Nicotinic Agonists MeSH
- Protein Subunits MeSH
- Pyridines MeSH
- Tubocurarine MeSH
The role of negatively charged amino acids in the F-loop of the beta 4 subunit in channel activation and desensitization was studied using the patch-clamp technique. The selected amino acids were changed to their neutral analogs via point mutations. Whole-cell currents were recorded in COS cells transiently transfected with the alpha 3 beta 4 nicotinic acetylcholine receptor. The application of acetylcholine (ACh), nicotine (Nic), cytisine (Cyt), carbamylcholine (CCh) and epibatidine (Epi) to cells clamped at -40 mV produced inward currents which displayed biphasic desensitization. The EC50 of Epi and Nic were increased by a factor of 3-6 due to mutations D191N or D192N. Only Epi remained an agonist in the double-mutated receptors with EC50 increased 17-fold. The interaction of the receptors with the competitive antagonist (+)tubocurarine (TC) was weakened almost 3-fold in the double-mutated receptors. The mutations increased the proportion of the slower desensitization component and increased the response plateau, resulting in decreased receptor desensitization. The double mutation substantially accelerated the return from long-term desensitization induced by Epi.
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