Article: article from journal or magazin.
Hydroxylated residues influence desensitization behaviour of recombinant alpha3 glycine receptor channels.
Journal of Neurochemistry
Publication types: Journal Article ; Research Support, Non-U.S. Gov'tPublication Status: ppublish
The human glycine receptor subunit alpha3 exists in two splice variants (alpha3K/L), with alpha3L bearing an additional segment of 15 amino acids within the cytoplasmic TM3-4 loop. Homomeric alpha3K glycine receptors show faster desensitization than alpha3L receptors. Ion channel properties were compared of alpha3L, alpha3K, and of the triple mutant alpha3LDeltaOH = alpha3L(T358A/Y367F/S370A), where hydroxyl functions of the spliced insert had been removed by site-directed mutagenesis. Upon recombinant expression in HEK 293 cells, patch-clamp recording experiments revealed that removal of hydroxyl functions primarily affected receptor desensitization. The fraction of non-desensitizing current was 68 +/- 13% for alpha3L, 21 +/- 13% for alpha3K, and 48 +/- 16% for alpha3LDeltaOH. Desensitization time constants at saturating glycine concentration were 8.4 +/- 2.8 s, 1.9 +/- 2.3 s, and 2.8 +/- 0.4 s, for alpha3L, alpha3K, and the triple mutant alpha3LDeltaOH, respectively. In contrast, single-channel and whole-cell properties were similar for all three constructs. Thus, ion channel activation, desensitization, and conductance properties are independently controlled by distinct structural elements. Hydroxyl functions within the M3-4 loop of the glycine receptor alpha3 subunit are crucial, but not exclusive, determinants of receptor desensitization.
Alternative Splicing, Biological Transport/physiology, Blotting, Western, Cell Line, Dose-Response Relationship, Drug, Electrophysiology, Glycine/pharmacokinetics, Humans, Hydroxylation, Kidney/cytology, Kidney/metabolism, Mutagenesis, Site-Directed, Patch-Clamp Techniques, Protein Subunits, Receptors, Glycine/genetics, Receptors, Glycine/metabolism, Structure-Activity Relationship, Transfection
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