Recombinant expression of aryl hydrocarbon receptor for quantitative ligand-binding analysis

Ming Qi Fan, Alex R Bell, David R Bell, Sally Clode, Alwyn Fernandes, Paul MD Foster, Jeffrey R Fry, Tao Jiang, George Loizou, Alan MacNicoll, Brian G Miller, Martin Rose, O Shaikh-Omar, Lang Tran, Shaun White

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Recombinant expression of the aryl hydrocarbon receptor (AhR) yields small amounts of ligand-binding-competent AhR. Therefore, Spodoptera frugiperda (Sf9) cells and baculovirus have been evaluated for high-level and functional expression of AhR. Rat and human AhR were expressed as soluble protein in significant amounts. Expression of ligand-binding-competent AhR was sensitive to the protein concentration of Sf9 extract, and coexpression of the chaperone p23 failed to affect the yield of functional ligand-binding AhR. The expression system yielded high levels of functional protein, with the ligand-binding capacity (Bmax) typically 20-fold higher than that obtained with rat liver cytosol. Quantitative estimates of the ligand-binding affinity of human and rat AhR were obtained; the Kd for recombinant rat AhR was indistinguishable from that of native rat AhR, thereby validating the expression system as a faithful model for native AhR. The human AhR bound TCDD with significantly lower affinity than the rat AhR. These findings demonstrate high-level expression of ligand-binding-competent AhR, and sufficient AhR for quantitative analysis of ligand binding.
Original languageUndefined
Pages (from-to)279-287
Number of pages9
JournalAnalytical Biochemistry
Issue number2
Publication statusPublished - 2009

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