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Revision as of 06:40, 7 December 2018
Coup Dy’état
A coup d’état is the sudden overthrow of a government through illegal force by a small group.
It is in this spirit that we offer Coup Dy’état, a novel biomanufacturing plaform which we hope has the potential to disrupt both conventional biomanufacturing and the synthetic dye industry.
Presenting, the Eco-friendly Biomanufacturing of Flavonoid Dyes in Escherichia coli via Computer-mediated Optogenetic Regulation.
The Problem
Water pollution is a key problem caused by the textile dyeing and dye manufacturing industry. This pollution is the result of the irresponsible disposal of industrial effluents, which is compounded by the persistence of synthetic dyes in the environment.
Photo: Iqbal Ksumadireza, The Straits Times, 2018
What is Happening
To illustrate the severity of the problem, we need only look at the Citarum River in Indonesia. It is not an uncommon sight to see the water rapidly changing colours due to the high level of toxic dye effluents pumped into the river daily by the synthetic dye industry. As a result, there is virtually no life left in the river [1]. Some 30 million residents who rely on the river as their only water source and livelihood are experiencing adverse skin conditions and increased exposure to infectious diseases [2].
The Citarum River is far from being an isolated case. China's Pearl River, Buriganga River in Bangladesh, and let's not forget the Bagmati River in India. In Dhaka, Bangladesh, 719 factories and textile mills produce close to 200 metric tons of wastewater per ton of fabric per year [4]. According to the World Bank, on the global scale, the textile dyeing and dyestuff production industry is the second most pollutive industry after oil, and generates a fifth of the world’s water pollution. It also consumes the most water apart from agriculture [5].
Treating the effluent is expensive, and legislation has been ineffective. In recent years, some efforts have been put in to revive natural dyes as a more sustainable alternative to synthetic dyes [6], but with little success. This is because traditional methods of producing natural dyes, primarily through plant extraction, face many constraints. The dyes produced are often inconsistent in quality. Furthermore, it is land and labour intensive, thus competing with agricultural demands.
Our Motivation
Our team strongly believes that the current approach of producing synthetic dyes is unsustainable. The world urgently needs a more sustainable and eco-friendly solution. We think that synthetic biology has the potential to increase the viability of natural dyes as an alternative to synthetic dyes. We want our dyes to be biodegradable and non-toxic, and for the manufacturing process to be cost-effective, involve less chemicals and be more environmentally-friendly. Ultimately, our product must also remain appealing to fashion designers and consumers.
The Challenges
However, biomanufacturing suffers from various constraints which prevent us from unlocking its full potential. Cost is a major limitation, as expensive chemical inducers and feedstock are often required [7].
Besides that, cells can only be maintained for so long in the bioreactor before they lose their biological activity or accumulate too many mutations for them to continue serving their intended purposes. Hence, frequent replacing of fermentation batches is necessary, which also diminishes the cost-effectiveness of biomanufacturing. The culprit is cellular stress, which is often left unregulated.
Our Solution
To address the challenges, our team developed a computer-mediated optogenetic regulation system as a technological platform for future biomanufacturing. The expression of enzymes involved in the biosynthetic pathway can be rapidly regulated by light. Burden-based cellular stress is monitored and regulated using custom hardware.
Firstly, to eliminate the use of expensive chemical inducers, we use optogenetic tools to facilitate dynamic gene regulation. This enables us to have precise and reversible induction of enzyme expression. We achieved this by designing an optogenetic circuit based on the blue light repressible system. The behaviour of this system was also characterized by self-designed hardware tools.
We also engineered our cells to promote utilization of xylose, a major constituent of lignocellulosic waste. This allows us to make use of this under-utilized waste, which represents a much more sustainable feedstock than conventional sugar sources.
Moreover, the monitoring of cellular stress is critical to prolong the productivity of cells in the bioreactor. We addressed this by introducing a stress-sensing fluorescence reporter, which provides us a way to quantify the level of burden experienced by cells when expressing recombinant proteins. This paves the way for stress regulation and ensuing optimization of protein expression.
Lastly, to enhance feedback operation and optimization of the biomanufacturing process, we developed a computer-aided system involving sensor modules. This allows automatic regulation of gene expression by activating or deactivating light according to feedback collected by the sensors.
To demonstrate this approach, we produced Luteolin, a natural yellow dye. Please view the animation below for an overview of how the system works.
References
[2] Asian Development Bank; The World Bank. Downstream Impacts of Water Pollution in the Upper Citarum River, West Java, Indonesia. Technical Paper. Manila: Asian Development Bank, 2013. Document. https://www.adb.org/sites/default/files/publication/154493/citarum-river-downstream-impacts-water-pollution.pdf
[3] Visvanathan, C. "3Rs for Water Security in Asia and the Pacific." United Nations Centre for Regional Development. 2015. 17. Document. http://www.uncrd.or.jp/content/documents/2660Pre-Final-BG-Plenary%20Session-6_FINAL.pdf
[4] Chequer, F. M., de Oliveira, G. A., Ferraz, E. R., Cardoso, J. C., Zanoni, M. V., & de Oliveira, D. P. (2013). Textile Dyes: Dyeing Process and Environmental Impact. IntechOpen, 151-176.
[5] Asian Development Bank; The World Bank. (2013). Downstream Impacts of Water Pollution in the Upper Citarum River, West Java, Indonesia. Manila: Asian Development Bank. Retrieved from https://www.adb.org/sites/default/files/publication/154493/citarum-river-downstream-impacts-water-pollution.pdf
[6] Křížová, Hana. (2015). Natural dyes: their past, present, future and sustainability. 59-71.
[7] Choi, W. (2018, Oct 11). Cost of Feedstock in Bio-based manufacturing. (A. Leow, Interviewer)
[8] Gasser, B., Saloheimo, M., Rinas, U., Dragosits, M., Rodríguez-Carmona, E., Baumann, K., ... & Porro, D. (2008). Protein folding and conformational stress in microbial cells producing recombinant proteins: a host comparative overview. Microbial cell factories, 7(1), 11.
[9] Ceroni, F., Boo, A., Furini, S., Gorochowski, T. E., Borkowski, O., Ladak, Y. N., ... & Ellis, T. (2018). Burden-driven feedback control of gene expression. Nature methods, 15(5), 387.
[10] Bäcklund et al. Microbial Cell Factories 2011, 10:35 http://www.microbialcellfactories.com/content/10/1/35