Solubilisation and binding characteristics of a recombinant beta2-adrenergic receptor expressed in the membrane of Escherichia coli for the multianalyte detection of beta-agonists and antagonists residues in food-producing animals.
The number of substances with beta-agonistic activity, illegally introduced in meat production or in sports doping as anabolic or beta-blocking agents is increasing. Analytical methods suited for their multianalyte detection are thus necessary. In this perspective, receptor assays were developed. The research activities undertaken in this study describe the solubilisation of a recombinant human beta(2)-adrenergic receptor produced in the inner membrane of genetically modified Escherichia coli, using the detergent n-dodecyl-beta-d-maltoside. Its potential to detect the presence of beta-agonists or beta-blockers in biological samples was evaluated. The solubilised beta(2)-adrenergic receptor retained its binding affinity in a radio-receptor assay based on the competition for the binding to receptors between a ligand (beta-agonist or antagonist) and the radioligand [(125)I]iodocyanopindolol. The IC(50) values ranged from 5+/-1 x 10(-8) M (clenbuterol) to 8+/-2 x 10(-6) M (isoxsuprine) for the beta-agonists tested and from 1.5+/-0.2 x 10(-10) M (carazolol) to 1.2+/-0.2 x 10(-5) M (metoprolol) for the beta-blockers tested. It was shown to have a lower limit of detection than a radio-receptor assay using the solubilised beta(2)-adrenoceptor expressed in a mammalian cell line. The solubilised recombinant human beta(2)-adrenoreceptor expressed in E. coli would be a useful tool to develop non radioactive multianalyte screening methods.
Danyi
Sophie
S
Department of Food Sciences, Laboratory of Analysis, Faculty of Veterinary Medicine, University of Liège, Sart-Tilman, Liège, Belgium. sdanyi@ulg.ac.be <sdanyi@ulg.ac.be>
Degand
Guy
G
Duez
Colette
C
Granier
Benoît
B
Maghuin-Rogister
Guy
G
Scippo
Marie-Louise
ML
D000318:Adrenergic beta-Agonists / Q000032:analysis*; D000319:Adrenergic beta-Antagonists / Q000032:analysis*; D000818:Animals; D004926:Escherichia coli / Q000235:genetics; D011485:Protein Binding; D018343:Receptors, Adrenergic, beta-2 / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012015:Reference Standards; D012995:Solubility
D000318:Adrenergic beta-Agonists; D000319:Adrenergic beta-Antagonists; D018343:Receptors, Adrenergic, beta-2; D011994:Recombinant Proteins