A bacterial biosensor of endocrine modulators.
The nuclear hormone receptors comprise one of the largest classes of protein targets for drug discovery, as their function has been linked to a variety of serious diseases, including several forms of cancer. Identifying novel compounds with the ability to modulate the function of these targets could lead to the development of effective therapeutics. In vivo sensors of ligand binding have emerged as tools that can greatly accelerate the lead identification process, allowing new drugs to be discovered more rapidly and cheaply. In this work, a novel sensor of nuclear hormone binding has been developed in Escherichia coli by constructing a fusion of the ligand-binding domain of the human estrogen receptor with a thymidylate synthase enzyme (TS). Expression of this fusion protein in TS-deficient bacterial cells resulted in growth phenotypes that were dependent on the presence of estrogen. Subsequent replacement of the estrogen receptor with the ligand-binding domain of the human thyroid hormone receptor led to specific thyroid hormone-enhanced growth that was insensitive to estrogen. This biosensor was then challenged with a small library of estrogen and thyroid hormone analogues, and it was observed that levels of cell growth correlate well with ligand-binding affinity. Remarkably, this simple biosensor was able to discriminate between agonistic and antagonistic activities, as combinations of estrogen agonists had an additive impact on cell growth, whereas known estrogen antagonists were found to neutralize agonist effects. This system constitutes a technique for facile selection of lead compounds with potential medical applications.
Skretas
Georgios
G
Department of Chemical Engineering, Princeton University, Engineering Quadrangle, Olden St., Princeton, NJ 08544, USA.
Wood
David W
DW
D015374:Biosensing Techniques; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D047628:Estrogen Receptor alpha / Q000378:metabolism*; D006801:Humans; D015202:Protein Engineering; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism*; D013940:Thymidylate Synthase / Q000235:genetics / Q000378:metabolism*
D047628:Estrogen Receptor alpha; D011993:Recombinant Fusion Proteins; D013940:Thymidylate Synthase