mu Opioid receptor: role for the amino terminus as a determinant of ligand binding affinity.
The importance of the amino-terminal domain of the mu opioid receptor (MOR) as a component of the high affinity ligand-binding pocket was evaluated. A deletion mutant lacking 64 amino acids from the amino-terminus of MOR (DeltaN64) was constructed and expressed in HEK 293 cells. The affinities of bremazocine and cyclazocine were similar for the truncated and full-length MORs. Affinities of the mu receptor antagonist, naloxone, and the mu receptor agonist, morphine, were decreased 3.5-fold and 6-fold, respectively, for the truncated receptor relative to the wild-type MOR. Similarly, the affinities of the opioid peptide agonists, DAMGO (Tyr-D-Ala-Gly-MePhe-Gly-ol), beta-endorphin, and DADL (Tyr-D-Ala-Gly-Phe-D-Leu), for the DeltaN64 receptor were decreased from 3- to 8-fold as a result of the deletion. In contrast, the affinities of the alkaloid agonists, methadone and fentanyl, and the peptide agonists, endomorphin 1 and endomorphin 2, for the truncated receptor relative to MOR were reduced dramatically by 20- to 60-fold. MOR is glycosylated when expressed in HEK 293 cells; however, analysis of N-glycosidase F-treated membranes indicated that N-glycan chains within the amino-terminal domain of MOR do not contribute significantly to ligand affinities. These results indicate that amino acid residues within the amino-terminal domain of MOR play a crucial role in the composition of the binding pocket for a select group of agonists.
Chaturvedi
K
K
Department of Biochemistry and Molecular Biology, University of Medicine and Dentistry of New Jersey, New Jersey Medical School, 185 South Orange Avenue, Newark, NJ 07103, USA.
Shahrestanifar
M
M
Howells
R D
RD
D000470:Alkaloids / Q000378:metabolism; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000235:genetics; D002460:Cell Line; D006801:Humans; D006868:Hydrolysis; D008024:Ligands; D008969:Molecular Sequence Data; D010455:Peptides / Q000378:metabolism; D011134:Polysaccharides / Q000378:metabolism; D011869:Radioligand Assay; D017450:Receptors, Opioid, mu / Q000819:agonists / Q000235:genetics / Q000378:metabolism*; D017384:Sequence Deletion; D013329:Structure-Activity Relationship; D014162:Transfection
D000470:Alkaloids; D000596:Amino Acids; D008024:Ligands; D010455:Peptides; D011134:Polysaccharides; D017450:Receptors, Opioid, mu