14-PSIL: Plug and Play Photosynthesis for RuBisCO Independent Fuels

Lead Research Organisation: Imperial College London
Department Name: Life Sciences

Abstract

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Technical Summary

Solar energy is a sustainable resource exceeding human energy demands by >3 orders of magnitude. If this diffuse energy can be concentrated and stored, it has the capacity to provide for human energy needs. The biological transformation of light to chemical energy (photosynthesis) is limited by the rate of carbon reduction. The goal of this project is to engineer pathways for diverting energy from carbon reduction to alternative sinks.
Our strategy is to engineer an intercellular, plug and play platform that allows electrons and/or reduced chemicals to move from photosynthetic cells to engineered fuel production modules, bypassing the inefficient carbon-fixing catalyst RuBisCO. This will be achieved by increasing flux through natural electron dissipation pathways, creating electrical connections between cell types, and employing a soluble redox shuttle to transfer reducing equivalents between cells. This international and interdisciplinary project is building bridges between the US and UK scientific communities in critical areas of synthetic biology, photosynthesis, electro-chemistry, catalysis, and metabolic regulation.
The scientific goals are organized around 4 specific aims. 1 Characterize components and control flux through the natural photosynthetic pathways in the cyanobacteria Synechocystis. 2 Construct artificial systems to sink reducing equivalents from photosynthesis. 3 Develop artificial means to move reducing equivalents outside the cell. 4 Produce artificial fuel production modules that require only reducing equivalents and CO2.
The project represents a radical approach to surpass natural photosynthesis by engineering a modular division of labor through electrical/chemical connectivity. The aims are devised to generate transformative research for technological applications and enable the discovery of fundamental science. Our goal, therefore, is to establish a platform to open a vast new frontier to develop new modular photosynthetic technologies.

Planned Impact

The long-term goals beyond the immediate project are to develop a pipeline of synthetic biology knowledge, skills, and personnel to rapidly optimize host organisms for the carbon neutral bioeconomy. This pipeline will support the UK bioprocessing, chemical, and energy industries and communities by acting as a hub of technology and will build the 'human infrastructure' of trained professionals. It is anticipated that the knowledge and technology infrastructure will ultimately accelerate time to market for developed and emerging bioprocessing applications.

Who will benefit from this research?
The synthetic biology / carbon fuel industry, as well as human society as a whole and the environment, will be the major beneficiaries of this research at all levels from multi-nationals to SMEs and spin-out companies. In addition, UK/US HEIs, students and the general public will also be beneficiaries, not to mention the UK/USA-plc as a whole. Also, the successful completion of the scientific goals of this program will transform thinking about photosynthesis by creating independent modules for studying and optimizing the light and dark processes as well as portable biowires to establish functional contacts between distinct cell types. These modules, as well as the platform for testing them as a system, will be freely shared with other researchers i.e. an open source technology approach. The successful completion of the project objective will enable potential adoption of new routes to sustainable materials, allowing industrial and academic users to explore a range of renewable bio-products in a rapid and cost-effective manner.

How will they benefit from this research?
Industry: The synthetic biology / carbon fuel industry will benefit from the new technologies generated in this research since it will provide performance upgrades to photosynthetic devices, as well as establishing a new solar-fuel paradigm that is RuBisCO independent. In particular, the need to bypass biological limitations for carbon capture is required to ensure that photosynthetic fuels can become a commercial reality. These benefits will be of great interest to SMEs and spin-outs that supply synthetic biology components and develop niche applications that will directly utilize and develop some of the technology in other directions, developing niche high-value applications. For instance, the knowledge generated could have third-party applications in new solar-fuel-cell devices that exploit, at a fundamental level, the new opportunities that multi-level devices have. The race to produce a realistic solar-fuel-system is gathering pace and this proposal could ensure that the UK-USA becomes the focal point for this development rather than the far east, Russia, or South Africa.
Education and Training: The interactions between molecular and synthetic biologists, inorganic chemists, and metabolic engineers proposed in this grant will also yield great potential teaching and research benefits for the students and the universities. This is because we estimate >20 undergraduate, >4 ERASMUS, and >12 PhD students will get the chance to take part in research that crosses the interface of this project and it may also be possible to develop a research masters based on this area that will train the next generation of researchers and engineers for solar fuels and photosynthesis.
General Public: The general public will benefit from this research due to the increase in wealth that will be developed and from the public understanding and promotion of science activities and public lectures planned for the UK and US sites.

Publications

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Description We have developed genetic tools that make it easier to study cyanobacteria and to genetically modify them for applied purposes, such as developing strains which produce industrial chemicals from carbon dioxide. These include a rhamnose-inducible system for precise and temporal external control of gene expression in cyanobacteria, developed by introducing the rhamnose-inducible rhaBAD promoter of Escherichia coli into the model freshwater cyanobacterium Synechocystis sp. PCC 6803 and demonstrating it has superior properties to previously reported cyanobacterial inducible promoter systems, such as a non-toxic, photostable, non-metabolisable inducer, a linear response to inducer concentration and crucially no basal transcription in the absence of inducer. We also used a reporter gene system to characterise the properties of a panel of heterologous and synthetic intrinsic (Rho-independent) transcriptional terminators Synechocystis sp. PCC 6803. Terminators are potentially important for heterologous gene expression, but had previously been little studied.

The overall aim of this project was to engineer systems to divert electrons and/or reduced chemicals from their natural destinations/sinks, bypassing the inefficient carbon-fixing catalyst RuBisCO, ultimately towards fuel/chemical production. To this end, a series of genetically modified strains of Synechocystis sp. PCC 6803 were constructed. Challenges were encountered, but preliminary indications of the intended functionality were obtained at the end of the project, and have been continued after the end of the project.
Exploitation Route These findings contribute to the body of underpinning research and capabilities for genetic modification of microbial strains, particularly cyanobacteria, in order to produce useful chemicals. These are useful to academic researchers and industrial researchers performing such work. At the time of publication, we deposit plasmid DNA samples for each genetic tool and synthetic biology technology at Addgene (addgene.org) who then distribute these to researchers on request.
Sectors Manufacturing, including Industrial Biotechology

 
Description rhaBAD syn bio, syn chem, enzymatic synthesis collaboration 
Organisation Kagawa University
Country Japan 
Sector Academic/University 
PI Contribution We performed synthetic biology work and studied the effect on our constructed biological system of the synthetic sugars prepared by our partners.
Collaborator Contribution Our partners contributed synthetic chemistry and enzymatic synthesis work to prepare synthetic sugars.
Impact The disciplines involved are synthetic biology, synthetic chemistry, and enzymatic synthesis.
Start Year 2015
 
Description rhaBAD syn bio, syn chem, enzymatic synthesis collaboration 
Organisation University of Oxford
Department Department of Chemistry
Country United Kingdom 
Sector Academic/University 
PI Contribution We performed synthetic biology work and studied the effect on our constructed biological system of the synthetic sugars prepared by our partners.
Collaborator Contribution Our partners contributed synthetic chemistry and enzymatic synthesis work to prepare synthetic sugars.
Impact The disciplines involved are synthetic biology, synthetic chemistry, and enzymatic synthesis.
Start Year 2015
 
Description rhaBAD syn bio, syn chem, enzymatic synthesis collaboration 
Organisation University of Santiago de Compostela
Country Spain 
Sector Academic/University 
PI Contribution We performed synthetic biology work and studied the effect on our constructed biological system of the synthetic sugars prepared by our partners.
Collaborator Contribution Our partners contributed synthetic chemistry and enzymatic synthesis work to prepare synthetic sugars.
Impact The disciplines involved are synthetic biology, synthetic chemistry, and enzymatic synthesis.
Start Year 2015