TY - JOUR
T1 - The serotonin binding site of human and murine 5-HT 2B receptors. Molecular modeling and site-directed mutagenesis
AU - Manivet, Philippe
AU - Schneider, Benôt
AU - Smith, Jeremy Christopher
AU - Choi, Doo Sup
AU - Maroteaux, Luc
AU - Kellermann, Odile
AU - Launay, Jean Marie
PY - 2002/5/10
Y1 - 2002/5/10
N2 - Bacteriorhodopsin and rhodopsin crystal structures were used as templates to build structural models of the mouse and human serotonin (5-HT)-2B receptors (5-HT 2BRs). Serotonin was docked to the receptors, and the amino acids predicted to participate to its binding were subjected to mutagenesis. 5-HT binding affinity and 5-HT-induced inositol triphosphate production were measured in LMTK - cells transfected with either wild-type or mutated receptor genes. According to these measurements, the bacteriorhodopsin-based models of the 5-HT 2BRs appear more confident than the rhodopsin-based ones. Residues belonging to the transmembrane domains 3 and 6, i.e. Asp 3.32, Ser 3.36, Phe 6.52, and Asn 6-55, make direct contacts with 5-HT. In addition, Trp 3.28, Phe 3.35, Phe 6.52, and Phe 7.38 form an aromatic box surrounding 5-HT. The specificity of human and mouse 5-HT 2BRs may be reflected by different rearrangements of the aromatic network upon 5-HT binding. Two amino acids close to Pro 5.50 in the human transmembrane domain 5 sequence were permuted to introduce a "mouse-like" sequence. This change was enough to confer the human 5-HT 2BR properties similar to those of the mouse. Taken together, the computed models and the site-directed mutagenesis experiments give a structural explanation to (i) the different 5-HT pK D values measured with the human and mouse 5-HT 2BRs (7.9 and 5.8, respectively) and (ii) the specificity of 5-HT binding to 5-HT 2BRs as compared with other serotonergic G-protein coupled receptors.
AB - Bacteriorhodopsin and rhodopsin crystal structures were used as templates to build structural models of the mouse and human serotonin (5-HT)-2B receptors (5-HT 2BRs). Serotonin was docked to the receptors, and the amino acids predicted to participate to its binding were subjected to mutagenesis. 5-HT binding affinity and 5-HT-induced inositol triphosphate production were measured in LMTK - cells transfected with either wild-type or mutated receptor genes. According to these measurements, the bacteriorhodopsin-based models of the 5-HT 2BRs appear more confident than the rhodopsin-based ones. Residues belonging to the transmembrane domains 3 and 6, i.e. Asp 3.32, Ser 3.36, Phe 6.52, and Asn 6-55, make direct contacts with 5-HT. In addition, Trp 3.28, Phe 3.35, Phe 6.52, and Phe 7.38 form an aromatic box surrounding 5-HT. The specificity of human and mouse 5-HT 2BRs may be reflected by different rearrangements of the aromatic network upon 5-HT binding. Two amino acids close to Pro 5.50 in the human transmembrane domain 5 sequence were permuted to introduce a "mouse-like" sequence. This change was enough to confer the human 5-HT 2BR properties similar to those of the mouse. Taken together, the computed models and the site-directed mutagenesis experiments give a structural explanation to (i) the different 5-HT pK D values measured with the human and mouse 5-HT 2BRs (7.9 and 5.8, respectively) and (ii) the specificity of 5-HT binding to 5-HT 2BRs as compared with other serotonergic G-protein coupled receptors.
UR - http://www.scopus.com/inward/record.url?scp=0037053372&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0037053372&partnerID=8YFLogxK
U2 - 10.1074/jbc.M200195200
DO - 10.1074/jbc.M200195200
M3 - Article
C2 - 11859080
AN - SCOPUS:0037053372
SN - 0021-9258
VL - 277
SP - 17170
EP - 17178
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 19
ER -