Functional role of the spatial proximity of Asp114(2.50) in TMH 2 and Asn332(7.49) in TMH 7 of the μ opioid receptor

FEBS Letters

Weinstein, Harel²  Liu-Chen, Lee-Yuan¹  de Riel, J.Kim³  Li, Jian-Guo¹  Xu, Wei¹  Chen, Chonguang¹  Ozdener, Fatih¹ 

[1] Department of Pharmacology, Temple University School of Medicine, 3420 N. Broad St., Philadelphia, PA 19140, USA;Department of Physiology and Biophysics, Mount Sinai School of Medicine, New York, NY, USA;Fels Institute for Molecular Biology and Cancer Research, Temple University School of Medicine, Philadelphia, PA, USA

关键词: μ Opioid receptor;    Structure-activity relationship;    Site-directed mutagenesis;    CHO cells;    Chinese hamster ovary cells;    CTAP;    D-Phe-Cys-Tyr-D-Trp-Arg-Thr-Pen-Thr-NH2;    DAMGO;    Try-D-Ala-Gly-N-Me-Phe-Gly-ol;    GDP;    guanosine diphosphate;    GnRH receptor;    gonadotropin-releasing hormone receptor;    GTPγS;    guanosine-5′-O-(3-thio)triphosphate;    HA;    hemagglutinin;    5-HT receptor;    5-hydroxytrytamine receptor;    TEL buffer;    50 mM Tris-HCl buffer containing 1 mM EGTA and 4 μM leupeptin (pH 7.4);    TMH;    transmembrane helix;   

DOI  :  10.1016/S0014-5793(99)00316-6

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

We examined whether a proposed spatial proximity between Asp114(2.50) and Asn332(7.49) affected the functional properties of the μ opioid receptor. The D114(2.50)N mutant had reduced binding affinities for morphine, DAMGO and CTAP, but not for naloxone and [³H]diprenorphine; this mutation also abolished agonist-induced increase in [³⁵S]GTPγS binding. The N332(7.49)D mutation eliminated detectable binding of either [³H]diprenorphine or [³H]DAMGO. The combined D114(2.50)N-N332(7.49)D mutation restored high affinity binding for [³H]diprenorphine, CTAP and naloxone, and restored partially the binding affinities, potencies and efficacies of morphine and DAMGO. Thus, reciprocal mutations of Asp114(2.50) and Asn332(7.49) compensate for the detrimental effects of the single mutations, indicating that the residues are adjacent in space and that their chemical functionalities are important for ligand binding and receptor activation.

【 授权许可】

Unknown