Expression and pharmacological characterization of the human μ‐opioid receptor in the methylotrophic yeast Pichia pastoris

FEBS Letters

Milon, Alain¹  Sidobre, Stéphane¹  Talmont, Franck¹  Demange, Pascal¹  Emorine, Laurent J.¹ 

[1] Institut de Pharmacologie et de Biologie Structurale, CNRS-UPR 9062, 205, route de Narbonne, 31077 Toulouse Cedex, France

关键词: μ-Opioid receptor;    Heterologous expression;    Pichia pastoris;    Membrane receptor;    Methylotrophic yeast;    HuMOR;    human μ-opioid receptor;    AOX1;    gene for alcohol oxidase-1;    STE2;    gene for α-mating factor receptor;    MD;    minimal dextrose;    MM;    minimal methanol;    BMGY;    buffered minimal glycerol-complex medium;    BMMY;    buffered minimal methanol-complex medium;    PMSF;    phenylmethylsulfonyl fluoride;    5-HT;    5-hydroxytryptamine;    DPN;    diprenorphine;    DAGO;    [d-Ala2;    N-Me-Phe4;    Gly-ol5]enkephalin;    DTLET;    Tyr-d-Thr-Gly-Phe-Leu-Thr;    U-50;    488;    trans-(−)-3;    4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)cyclohexyl]benzene acetamide;    β-FNA;    β-funaltrexamine;    ICI-174;    846;    N;    N-diallyl-Tyr-Aib-Aib-Phe-Leu;    NorBNI;    nor-binaltorphimine;   

DOI  :  10.1016/0014-5793(96)00971-4

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

The human μ-opioid receptor cDNA from which the 32 amino-terminal codons were substituted by the Saccharomyces cerevisiae α-mating factor signal sequence has been expressed in the methylotrophic yeast Pichia pastoris using the host promoter of the alcohol oxidase-1 gene. Cell membranes exhibited specific and saturable binding of the opioid antagonist [³H]diprenorphine (K _d = 0.2 nM and B _max = 400 fmol/mg protein or 800 sites/cell). Competition studies with non-selective, and μ-, δ- and κ-selective opioid agonists and antagonists revealed a typical μ-opioid receptor binding profile, suggesting proper folding of the protein in yeast membranes.

【 授权许可】

Unknown