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Common Determinants of Single Channel Conductance within the Large Cytoplasmic Loop of 5-Hydroxytryptamine Type 3 and α4β2 Nicotinic Acetylcholine Receptors
- Source :
- Journal of Biological Chemistry. 281:8062-8071
- Publication Year :
- 2006
- Publisher :
- Elsevier BV, 2006.
-
Abstract
- Homomeric 5-hydroxytryptamine type 3A receptors (5-HT3ARs) have a single channel conductance (gamma) below the resolution of single channel recording (966 +/- 75 fS, estimated by variance analysis). By contrast, heteromeric 5-HT3A/B and nicotinic acetylcholine receptors (nAChRs) have picosiemen range gamma values. In this study, single channel recordings revealed that replacement of cytoplasmic membrane-associated (MA) helix arginine 432 (-4'), 436 (0'), and 440 (4') residues by 5-HT3B (-4'Gln, 0'Asp, and 4'Ala) residues increases gamma to 36.5 +/- 1.0 pS. The 0' residue makes the most substantial contribution to gamma of the 5-HT3AR. Replacement of 0'Arg by aspartate, glutamate (alpha7 nAChR subunit MA 0'), or glutamine (beta2 subunit MA 0') increases gamma to the resolvable range (6 pS). By contrast, replacement of 0'Arg by phenylalanine (alpha4 subunit MA 0') reduced gamma to 416 +/- 107 fS. In reciprocal experiments with alpha4beta2 nAChRs (gamma = 31.3 +/- 0.8 pS), replacement of MA 0' residues by arginine in alpha4beta2(Q443R) and alpha4(F588R)beta2 reduced gamma slightly. By contrast, the gamma of double mutant alpha4(F588R)beta2(Q443R) was halved. The MA -4' and 4' residues also influenced gamma of 5-HT3ARs. Replacement of nAChR alpha4 or beta2 MA 4' residues by arginine made current density negligible. By contrast, replacement of both -4' residues by arginine produced functional nAChRs with substantially reduced gamma (11.4 +/- 0.5 pS). Homology models of the 5-HT3A and alpha4beta2 nAChRs against Torpedo nAChR revealed MA -4', 0', and 4' residues within five intracellular portals. This locus may be a common determinant of ion conduction throughout the Cys loop receptor family.
- Subjects :
- Models, Molecular
Cytoplasm
Serotonin
RM
DNA, Complementary
Patch-Clamp Techniques
Time Factors
Arginine
Protein Conformation
Stereochemistry
Protein subunit
Molecular Sequence Data
Receptors, Nicotinic
Torpedo
Transfection
Biochemistry
Cell Line
law.invention
law
Animals
Humans
Homomeric
Amino Acid Sequence
Cysteine
Receptor
Molecular Biology
QC
Acetylcholine receptor
Ions
Models, Statistical
Sequence Homology, Amino Acid
Chemistry
Cell Membrane
Glutamate receptor
DNA
Cell Biology
QP
Protein Structure, Tertiary
Rats
Electrophysiology
Nicotinic agonist
Mutation
Calcium
Subjects
Details
- ISSN :
- 00219258
- Volume :
- 281
- Database :
- OpenAIRE
- Journal :
- Journal of Biological Chemistry
- Accession number :
- edsair.doi.dedup.....8f0732f1c5d807239750bc7709d55d15