Improved Feedstock Production Using Engineered Cyanobacterial for Mixotrophy Under Natural Light Conditions

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With the advent of global research towards the biological synthesis of value-added compounds and hydrocarbon molecules, photosynthetic organisms are attracting considerable interest in the scientific world. Cyanobacteria, prokaryotic photosynthetic organisms, are regarded as efficient platform cell factories for the production of these chemicals through genetic modifications.

This could be attributed to their natural competency to uptake extracellular DNA, polyploidy and use of genomic integration strategy or homologous recombination for genetic engineering. However, scale-up of the mutants generated through such modifications is dependent on quality as well as the quantity of the feedstock produced. Moreover, all scale-ups have to be performed under natural light conditions as the operational culture volumes are in million liters. Additionally, natural light is low input high output alternative over artificial light, in terms of both capital as well as operational expenditure. However, natural light exhibits diurnal as well as seasonal variation which could affect cell sustenance and cell growth.

Thus, reducing the sole dependency of cyanobacterial cells on light and CO2 by providing an alternative energy source could help improve biomass feedstock under sunlight. Providing both the energy sources simultaneously (light, CO2; and organic C), the phenomenon is known as mixotrophy, would help cells grow as well as withstand natural light variations.

We at DBT-ICT Centre for Energy Biosciences, Mumbai generated Synechococcus elongatus PCC 7942 mutant, as demonstrated by Dr. Atsumi at UC Davis, that could uptake extracellular glucose, which otherwise cannot. Essentially, we had projected two goals for the study, one was to construct the mutants for growing under natural light, and another was to establish physiological principles of cyanobacterial mixotrophy.

In a recent research article published in Biotechnology Progress, we found that when the glucose uptaking mutants were fed with 10g/L glucose in the growth medium BG-11, the biomass could improve 3-3.5 times over the one cultivated autotrophically i.e. without supplementation of any organic C, under natural light.

Further, we established three important physiological principles underlying natural light mixotrophy. Cyanobacteria display improved growth under natural light conditions (maximum photon intensity 1200 ± 100 µmol/m2/s) as compared to that under cool artificial light (55 ± 0.5 µmol/m2/s). Cyanobacteria when cultivated under mixotrophy, reduce their dependency on light thereby reducing the yield of photosynthetic pigments; essentially chlorophyll-a molecules per cell.

This helps in reducing photo-oxidative damage to the photosynthetic machinery of the cell. Nitrogen is a vital macronutrient required by the cell which could be provided to the cell in an inorganic nitrate form. Nitrate assimilation is generally coupled with photosynthesis as electrons generated during photophosphorylation are used for nitrate reduction to nitrite and ultimately ammonium. Thus under mixotrophy, with a decrease in photosynthetic reaction centers, there is a reduction in nitrate uptake rate by the cell.

Understanding the mixotrophic physio-chemical behavior of the cyanobacteria is necessary for improving high scale production of natural or engineered product from the cell. For instance, the coupling of hexose transporter gene with another gene for synthesis of the desired biomolecule could not only help in increasing the biomass but also supports improved titer of the desired product by the cell.

These findings are described in the article entitled Growth engineering of Synechococcus elongatus PCC 7942 for mixotrophy under natural light conditions for improved feedstock production, published in the journal Biotechnology Progress. This work was led by Dr. Reena Pandit and her Ph.D. student Aditya P. Sarnaik at DBT-ICT Centre for Energy Biosciences at Mumbai, India.

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