Abstract
As a novel chemical production platform, controllable and inducible modules in Synechococcus elongatus plus the ability of working in diurnal conditions are necessary. To the endeavors, inducible promoters, such as PTrc, have been refined from Escherichia coli, but the inducer isopropyl-β-D-thiogalactoside may cause several side-effects. Meanwhile, to promote the efficiency, photomixotrophic cultivation has been applied in S. elongatus with the additional organic carbon sources. In this study, we developed L-arabinose based modules consisted of both the PBAD inducible promoter and the metabolism of L-arabinose in S. elongatus, since L-arabinose is an ideal heterologous feedstock for its availability and economic and environmental benefits. As expected, we achieved homogeneous and linear expression of the exogenous reporter through the PBAD promoter, and the biomass increased in diurnal light condition via introducing L-arabinose metabolism pathway. Moreover, the combined AraBAD based toolkit containing both the PBAD inducible module and the L-arabinose metabolism module could obtain gene expression and metabolic robustness improvement in S. elongatus. With the only additive L-arabinose, the novel strategy may generate a win-win scenario for both regulation and metabolism for autotrophic bio-production platforms.
Highlights
The obligate photoautotrophic organism S. elongatus, has been engineered to produce various chemicals at higher titers and productivities relative to other cyanobacteria[4,19,20]
To expand synthetic biology toolboxes and to enable S. elongatus to accumulate biomass in diurnal condition, the current study proposed an applicable AraBAD based toolkit in S. elongatus including both the inducible module driven by PBAD promoter and L-arabinose metabolic pathway
We first compared the PTrc and PBAD promoters in S. elongatus using a modified GFP28 as a reporter because of a stronger fluorescence than eGFP (Fig. S1). Both PTrc-mtgfp and PBAD-mtgfp fragments were integrated into the chromosome at neutral site I (NSI)[29] in S. elongatus
Summary
The obligate photoautotrophic organism S. elongatus, has been engineered to produce various chemicals at higher titers and productivities relative to other cyanobacteria[4,19,20]. Natural sunlight is freely available, so it would be more feasible and commercial to utilize S. elongatus in natural conditions, where light and dark cycle every 12 h To overcome this innate physiology, several studies explored the potentials of transplanting heterotrophic carbon metabolic pathways to enable sugar utilization. To expand synthetic biology toolboxes and to enable S. elongatus to accumulate biomass in diurnal condition, the current study proposed an applicable AraBAD based toolkit in S. elongatus including both the inducible module driven by PBAD promoter and L-arabinose metabolic pathway. In this case, L-arabinose was the inducer and a carbon source to regulate genetic modules and promote the metabolic robustness. We believe that our strategy provides a preferable genetic toolkit for S. elongatus and generates a win-win scenario for both gene expression and metabolic robustness in autotrophic bio-production platforms
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