A challenge of the synthetic biology approach is to use our understanding of a system to recreate a biological function with specific properties. We have applied this framework to bacterial enhancers, combining a driver, transcription factor binding sites, and a poised polymerase to create synthetic modular enhancers. Our findings suggest that enhancer-based transcriptional control depends critically and quantitatively on DNA looping, leading to complex regulatory effects when the enhancer cassettes contain additional transcription factor binding sites for TetR, a bacterial transcription factor. We show through a systematic interplay of experiment and thermodynamic modeling that the level of gene expression can be modulated to convert a variable inducer concentration input into discrete or step-like output expression levels. Finally, using a different DNA-binding protein (TraR), we show that the regulatory output is not a particular feature of the specific DNA-binding protein used for the enhancer but a general property of synthetic bacterial enhancers.
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