Human μ—opioid receptor overexpressed in Sf9 insect cells functionally coupled to endogenous Gi／0 proteins-中国Betway体育网页登陆

摘要 Humanμ-opioidreceptor(HμOR)withatagofsixconsecutivehistidinesatitscarboxylterminushasbeenexpressedinrecombinantbaculovirusinfectedSf9insectcells.Themaximalbindingcapacityforthe[^3H]diprenorphineand[^3H]ohmefentanyl(Ohm)were9.1±0.7and6.52±0.23nmol/gprotein,respectively.The[^3H]diprenorphineor[^3H]OhmbindingtothereceptorexpressedinSf9cellswasstronglyinhibitedbyμ-selectiveagonists[D-Ala^2,N-methyl-Phe^4,glyol^5]enkephalin(DAGO),Ohm,andmorphine,butneitherbyδnorbyκselectiveagonist.Na^+(100mM)andGTP(50μM)couldreduceHμORagonistsetorphineandOhmaffinitybindingtotheoverexpressedHμOR.μ-selectiveagonistsDAGOandOhmeffectivelystimulated[^35S]GTPγSbinding(EC50=2.7nMand6.9nM)andinhibitedforskolin-stimulatedcAMPaccumulation(IC50=0.9nMand0.3nM).Theagonist-dependenteffectscouldbeblockedbyopioidantagonistnaloxoneorbypretreatmentofcellswithpertussistoxin(PTX).TheseresultsdemonstratedthatHμORoverexpressedinSf9insectcellsfunctionallycoupledtoendogenousCi/oproteins.

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Human μ—opioid receptor overexpressed in Sf9 insect cells functionally coupled to endogenous Gi／0 proteins

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