Expression of the porcine beta2-adrenergic receptor in Chinese hamster ovary cells.
The gene for the porcine beta2-adrenergic receptor (pbeta2AR) was transfected into Chinese hamster ovary (CHO) cells for expression. Fourteen stable cell lines were obtained and exhibited receptor densities ranging from 12 to 2,371 fmol/mg membrane protein. The receptor density was not correlated with estimates of gene copy number obtained by Southern hybridization. The pbeta2AR in CHO cells exhibited saturable binding of [125I]CYP (Kd = 14.5 pM) and stereospecificity for (-)- and (+)-isoproterenol. The relative affinities for (-)-isoproterenol (ISO), (-)-epinephrine (EPI), and (-)-norepinephrine (NEPI) were ISO > EPI > NEPI, which are characteristic of beta2AR. The affinity values for these ligands were similar to those in other species. Binding of ISO, EPI, and NE revealed two affinity states of the betaAR; the high-affinity state was eliminated by adding Gpp(NH)p, a nonhydrolyzable GTP analogue. Binding of the antagonist propranolol modeled to only one affinity state, and Gpp(NH)p did not affect binding. Multiple affinity states are characteristic of agonist-induced coupling of betaAR with G-proteins, and the data suggest that the cloned pbetaAR is functionally competent. Data confirm that the pbeta2AR is the pig version of beta2AR. Stable CHO cell lines will be useful for characterization of pbeta2AR and screening and designing potential drugs that may be used to enhance pig production.
Liang
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W
Department of Animal Sciences, Purdue University, West Lafayette, IN 47907, USA.
Bidwell
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CA
Collodi
P R
PR
Mills
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SE
D000818:Animals; D001667:Binding, Competitive; D016466:CHO Cells; D006224:Cricetinae; D020120:Iodocyanopindolol / Q000378:metabolism; D018343:Receptors, Adrenergic, beta-2 / Q000096:biosynthesis* / Q000235:genetics; D013552:Swine / Q000378:metabolism*; D014162:Transfection
D018343:Receptors, Adrenergic, beta-2; D020120:Iodocyanopindolol