TY - JOUR
T1 - Identification of residues mediating inhibition of glycine receptors by protons
AU - Chen, Zhenglan
AU - Huang, Renqi
N1 - Funding Information:
This research was supported by American Heart Association Texas Affiliate (RQH), and Faculty Research Grant from University of North Texas Health Science Center (RQH). We thank Dr Glenn Dillon for helpful comments on the manuscript, and Dr Heinrich Betz for providing the wild type glycine α1 and β subunit cDNA.
PY - 2007/6
Y1 - 2007/6
N2 - We previously identified H109 of the glycine α1 subunit as a putative proton binding site. In the present studies, we explored additional proton binding site(s) as well as the mechanism underlying modulation of glycine receptors by protons. Whole-cell glycine currents were recorded from HEK 293 cells transiently expressing wild type or mutant glycine receptors. Individual mutation of 3 of 4 remaining extracellular histidine residue into alanine (i.e., α1 H107A, H215A or H419A), reduced the receptor sensitivity to protons to a varying extent. In contrast, mutation of α1 H201A did not affect proton sensitivity. Double, triple or quadruple histidine mutation of these residues caused a further reduction of proton sensitivity, suggesting multiple binding sites for proton action on glycine receptors. Furthermore, the substitution T133A, which mediates Zn2+ inhibition, virtually abolished the proton effect on peak amplitude and current kinetics of glycine response. Replacement of T with S on position 133 partially restored receptor sensitivity to protons, suggesting the hydroxyl group of residue T133 is essential for proton-mediated modulation. In heteromeric α1β receptors, mutations β H132A and S156A, which correspond to H109 and T133 of the α1 subunit, respectively, also affected proton inhibition. In conclusion, multiple extracellular histidine residues (H107, H109, H215 and H419) and threonine residues of the α1 and β Zn2+ coordination sites are critical for modulation of the glycine receptor by protons.
AB - We previously identified H109 of the glycine α1 subunit as a putative proton binding site. In the present studies, we explored additional proton binding site(s) as well as the mechanism underlying modulation of glycine receptors by protons. Whole-cell glycine currents were recorded from HEK 293 cells transiently expressing wild type or mutant glycine receptors. Individual mutation of 3 of 4 remaining extracellular histidine residue into alanine (i.e., α1 H107A, H215A or H419A), reduced the receptor sensitivity to protons to a varying extent. In contrast, mutation of α1 H201A did not affect proton sensitivity. Double, triple or quadruple histidine mutation of these residues caused a further reduction of proton sensitivity, suggesting multiple binding sites for proton action on glycine receptors. Furthermore, the substitution T133A, which mediates Zn2+ inhibition, virtually abolished the proton effect on peak amplitude and current kinetics of glycine response. Replacement of T with S on position 133 partially restored receptor sensitivity to protons, suggesting the hydroxyl group of residue T133 is essential for proton-mediated modulation. In heteromeric α1β receptors, mutations β H132A and S156A, which correspond to H109 and T133 of the α1 subunit, respectively, also affected proton inhibition. In conclusion, multiple extracellular histidine residues (H107, H109, H215 and H419) and threonine residues of the α1 and β Zn2+ coordination sites are critical for modulation of the glycine receptor by protons.
KW - Copper
KW - Glycine receptor
KW - Histidine
KW - Protons
KW - Threonine
KW - Zinc
KW - pH
UR - http://www.scopus.com/inward/record.url?scp=34249711369&partnerID=8YFLogxK
U2 - 10.1016/j.neuropharm.2007.03.005
DO - 10.1016/j.neuropharm.2007.03.005
M3 - Article
C2 - 17459427
AN - SCOPUS:34249711369
SN - 0028-3908
VL - 52
SP - 1606
EP - 1615
JO - Neuropharmacology
JF - Neuropharmacology
IS - 8
ER -