Cyanobacteria, a group of photosynthetic prokaryotes, are attractive hosts for biotechnological applications. It is envisaged that future biorefineries will deploy engineered cyanobacteria for the conversion of carbon dioxide to useful chemicals via light-driven, endergonic reactions. Fast-growing, genetically amenable, and stress-tolerant cyanobacteria are desirable as chassis for such applications. The recently reported strains such as Synechococcus elongatus UTEX 2973 and PCC 11801 hold promise, but additional strains may be needed for the ongoing efforts of metabolic engineering. Here, we report a novel, fast-growing, and naturally transformable cyanobacterium, S. elongatus PCC 11802, that shares 97% genome identity with its closest neighbor S. elongatus PCC 11801. The new isolate has a doubling time of 2.8 h at 1% CO2, 1000 µmole photons.m−2.s−1 and grows faster under high CO2 and temperature compared to PCC 11801 thus making it an attractive host for outdoor cultivations and eventual applications in the biorefinery. Furthermore, S. elongatus PCC 11802 shows higher levels of key intermediate metabolites suggesting that this strain might be better suited for achieving high metabolic flux in engineered pathways. Importantly, metabolite profiles suggest that the key enzymes of the Calvin cycle are not repressed under elevated CO2 in the new isolate, unlike its closest neighbor.
Cyanobacteria
are emerging as hosts for various biotechnological
applications. The ability to engineer these photosynthetic prokaryotes
greatly depends on the availability of well-characterized promoters.
Inducer-free promoters of a range of activities may be desirable for
the eventual large-scale, outdoor cultivations. Further, several native
promoters of cyanobacteria are repressed by high carbon dioxide or
light, and it would be of interest to alter this property. We started
with PrbcL and PcpcB, the well-characterized
native promoters of the model cyanobacterium Synechococcus
elongatus PCC 7942, found upstream of the two abundantly
expressed genes, Ribulose-1,5-Bisphosphate Carboxylase/Oxygenase,
and phycocyanin β-1 subunit, respectively. The library of 48
promoters created via error-prone PCR of these 300-bp-long native
promoters showed 2 orders of magnitude dynamic range with activities
that were both lower and higher than those of the wild-type promoters.
A few mutants of the PrbcL showed greater strength than
PcpcB, which is widely considered a superstrong promoter.
A number of mutant promoters did not show repression by high CO2 or light, typically found for PrbcL and PcpcB, respectively. Further, the wild-type and mutant promoters
showed comparable activities in the fast-growing and stress-tolerant
strains S. elongatus PCC 11801 and PCC 11802, suggesting
that the library can be used in different cyanobacteria. Interestingly,
the majority of the promoters showed strong expression in E. coli, thus adding to the repertoire of inducer-free promoters
for this heterotrophic workhorse. Our results have implications in
the metabolic engineering of cyanobacteria and E. coli.
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