Synthetic Chemical Inducers and Genetic Decoupling Enable Orthogonal Control of the rhaBAD Promoter

• DOI: 10.1021/acssynbio.6b00030
• Publication Date: 2016-06-01T02:09:46Z
• Authors: Ciarán L. Kelly; Zilei Liu; Akihide Yoshihara; Sarah F. Jenkinson; Mark R. Wormald; Jose Otero; Amalia Estévez; Atsushi Kato; Mikkel H. S. Marq...; George W. J. Fleet; Ramón J. Estévez; Ken Izumori; John T. Heap
• Keywords: Inducer; Synthetic Biology

External control of gene expression is crucial in synthetic biology and biotechnology research applications, commonly achieved using inducible promoter systems. The E. coli rhamnose-inducible rhaBAD has properties superior to more used systems, but marred by transient caused degradation the native inducer, l-rhamnose. To address this problem, 35 analogues l-rhamnose were screened for induction promoter, no strong inducers identified. In configuration, an inducer must bind activate two transcriptional activators, RhaR RhaS. Therefore, system was reconfigured decouple from rhaSR regulatory cascade so that candidate need only terminal transcription factor Rescreening compounds modified revealed several promising inducers. These characterized further determine strength, kinetics, concentration-dependence induction; whether as a carbon source coli; modality (distribution) among populations cells. l-Mannose found be most useful orthogonal providing even greater range than l-rhamnose, crucially, allowing sustained instead induction. findings key limitation suggest it may now suitable many applications.