2 research outputs found
On the Use of Metabolic Control Analysis in the Optimization of Cyanobacterial Biosolar Cell Factories
Oxygenic photosynthesis will have
a key role in a sustainable future. It is therefore significant that
this process can be engineered in organisms such as cyanobacteria
to construct cell factories that catalyze the (sun)light-driven conversion
of CO<sub>2</sub> and water into products like ethanol, butanol, or
other biofuels or lactic acid, a bioplastic precursor, and oxygen
as a byproduct. It is of key importance to optimize such cell factories
to maximal efficiency. This holds for their light-harvesting capabilities
under, for example, circadian illumination in large-scale photobioreactors.
However, this also holds for the “dark” reactions of
photosynthesis, that is, the conversion of CO<sub>2</sub>, NADPH,
and ATP into a product. Here, we present an analysis, based on metabolic
control theory, to estimate the optimal capacity for product formation
with which such cyanobacterial cell factories have to be equipped.
Engineered l-lactic acid producing <i>Synechocystis</i> sp. PCC6803 strains are used to identify the relation between production
rate and enzymatic capacity. The analysis shows that the engineered
cell factories for l-lactic acid are fully limited by the
metabolic capacity of the product-forming pathway. We attribute this
to the fact that currently available promoter systems in cyanobacteria
lack the genetic capacity to a provide sufficient expression in single-gene
doses
Nonhierarchical Flux Regulation Exposes the Fitness Burden Associated with Lactate Production in <i>Synechocystis</i> sp. PCC6803
Cyanobacteria are
mostly engineered to be sustainable cell-factories
by genetic manipulations alone. Here, by modulating the concentration
of allosteric effectors, we focus on increasing product formation
without further burdening the cells with increased expression of enzymes.
Resorting to a novel 96-well microplate cultivation system for cyanobacteria,
and using lactate-producing strains of <i>Synechocystis</i> PCC6803 expressing different l-lactate dehydrogenases (LDH),
we titrated the effect of 2,5-anhydro-mannitol supplementation. The
latter acts in cells as a nonmetabolizable analogue of fructose 1,6-bisphosphate,
a known allosteric regulator of one of the tested LDHs. In this strain
(SAA023), we achieved over 2-fold increase of lactate productivity.
Furthermore, we observed that as carbon is increasingly deviated during
growth toward product formation, there is an increased fixation rate
in the population of spontaneous mutants harboring an impaired production
pathway. This is a challenge in the development of green cell factories,
which may be countered by the incorporation in biotechnological processes
of strategies such as the one pioneered here