Biosensors are important components of many synthetic biology and metabolic engineering applications. Here, we report a second generation of Saccharomyces cerevisiae digoxigenin and progesterone biosensors based on destabilized dimeric ligand-binding domains that undergo ligand-induced stabilization. The biosensors, comprising one ligand-binding domain monomer fused to a DNA-binding domain and another fused to a transcriptional activation domain, activate reporter gene expression in response to steroid binding and receptor dimerization. The introduction of a destabilizing mutation to the dimer interface increased biosensor dynamic range by an order of magnitude. Computational redesign of the dimer interface and functional selections were used to create heterodimeric pairs with further improved dynamic range. A heterodimeric biosensor built from the digoxigenin and progesterone ligand-binding domains functioned as a synthetic “AND”-gate, with 20-fold stronger response to the two ligands in combination than to either one alone. We also identified mutations that increase the sensitivity or selectivity of the biosensors to chemically similar ligands. These dimerizing biosensors provide additional flexibility for the construction of logic gates and other applications.

中文翻译：

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来自二聚体配体结合域的工程生物传感器

生物传感器是许多合成生物学和代谢工程应用的重要组成部分。在这里，我们报告第二代酿酒酵母地高辛配基和孕激素生物传感器基于经过配体诱导的稳定化的不稳定二聚体配体结合域。该生物传感器包含一个与DNA结合域融合的配体结合域单体和另一个与转录激活域融合的生物传感器，它们响应类固醇结合和受体二聚化而激活报告基因的表达。将不稳定的突变引入二聚体界面将生物传感器动态范围增加了一个数量级。二聚体界面的计算重新设计和功能选择被用于创建具有进一步改善的动态范围的异二聚体对。由洋地黄毒苷和孕酮配体结合域构建的异二聚体生物传感器可作为合成的“与”门，对两种配体的响应要比对任何一个配体的响应强20倍。我们还确定了增加生物传感器对化学相似配体的敏感性或选择性的突变。这些二聚化生物传感器为逻辑门和其他应用的构建提供了额外的灵活性。