Volume 10, No 2, Jun 2000

ISSN: 1001-0602 
EISSN: 1748-7838 2018 
impact factor 17.848* 
(Clarivate Analytics, 2019)

Volume 10 Issue 2, June 2000: 93-102

ORIGINAL ARTICLES

Human μ-opioid receptor overexpressed in Sf9 insect cells functionally coupled to endogenous G_i/o proteins

WEI Qiang¹, De He ZHOU^*,1, Qing Xiang SHEN², Jie CHEN¹, Li Wei CHEN¹, Tie Lin WANG¹, Gang PEI², Zhi Qiang CHI¹

¹ Shanghai Institute of Materia Medica,
² Shanghai Institute of Cell Biology, Shanghai Academy of Life Sciences, Chinese Academy of Sciences, Shanghai 200031, China. Correspondence:

Human m-opioid receptor (HmOR) with a tag of six consecutive histidines at its carboxyl terminus had been expressed in recombinant baculovirus infected Sf9 insect cells. The maximal binding capacity for the [³H]
diprenorphine and [³H]ohmefentanyl (Ohm) were 9.1 ± 0.7 and 6.52 ± 0.23 nmol/g protein, respectively. The [³H] diprenorphine or [³H] Ohm binding to the receptor expressed in Sf9 cells was strongly inhibited by m-selective agonists [D-Ala², N-methyl-Phe⁴, glyol⁵]enkephalin (DAGO), Ohm, and morphine, but neither by d nor by k selective agonist. Na⁺ (100 m M) and GTP (50 m M) could reduce HmOR agonists etorphine and Ohm affinity binding to the overexpressed HmOR. m-selective agonists DAGO and Ohm effectively stimulated [³⁵S]GTPgS binding (EC₅₀ = 2.7 n M and 6.9 n M) and inhibited forskolin- stimulated cAMP accumulation (IC₅₀ = 0.9 n M and 0.3 n M). The agonist-dependent effects could be blocked by opioid antagonist naloxone or by pretreatment of cells with pertussis toxin (PTX). These results demonstrated that HmOR overexpressed in Sf9 insect cells functionally coupled to endogenous G_i/o proteins.

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