Synthetic Biology and the Biofuel Revolution
In November 2011, NNFCC – the UK’s National Centre for Biorenewable Energy, Fuels and Materials –published a report urging for a substantial increase in investment into research on new types of bio-fuel for energy generation.
Why is there such an urgent need to develop new fuel sources? Records show that the UK alone consumes 200 million tonnes of fuel a year, with a large chunk derived from non-renewable fossil fuels, including petrol and diesel. The rising costs of these fuels, and dwindling natural reserves, makes it imperative that we find alternative energy solutions. Fossil fuels also have a significant environmental impact, producing greenhouse gases like carbon dioxide, which contribute to global warming. There is a massive drive by the UK Government to replace 10% of the fossil fuel consumption with alternative sources by 2020, and reduce 80% of the greenhouse emissions by 2050.
Biofuels are considered the next-generation fuels, as they have the potential to be “carbon-neutral”. The rate at which carbon is trapped during photosynthesis by plants is almost the same as the rate of its release into the atmosphere, when plant-derived biofuels are burnt to release energy. Achieving a truly carbon-neutral output does present some challenges, however, as a considerable part of the carbon in plants can be trapped as indigestible material, like cellulose and lignin. First generation biofuels, like corn-based ethanol, use the edible and easily digestible leafy parts of the plant to produce biofuel, leading to a competition for food versus fuel.
Engineered E. coli cells produce biodiesel from sugar (seen here as oil droplets surrounding the cell). Image credit: Marcin Zemla and Manfred Auer, JBEI Source: http://newscenter.lbl.gov/news-releases/2010/01/27/microbes-produce-biofuels/
Another challenge lies in creating ‘drop-in’ fuels – fuels that can directly be added to current engines without any further modifications. While bioethanol can be added to petrol-based engines, it is a lot less ‘energy-dense’ than petrol, and can also cause corrosion of engine parts and transport pipelines. Similarly, biodiesel composition can vary from batch to batch, and not all of it meets the standards required for current diesel engines. Thus neither of these fuels is an ideal replacement for use in jet engines or freight lorries, both of which are responsible for the majority of greenhouse emissions into the atmosphere.
These concerns have led to investment in an advanced generation of biofuels, based on synthetic biology. The goal of the scientists in this field is to create longer, more ‘energy-dense’ alcohols than ethanol, and to produce alternative synthetic fuels that will more closely resemble the traditional fuels, like petrol and diesel, which can be directly dropped into existing engines. While microorganisms like yeast readily produces alcohol, and algae produces oil as a by-product of photosynthesis, the challenge lies in getting these organisms to produce the fuels on the right ‘type’—for example, the right carbon chain length for petrol—and in volumes large enough to compete with, and meet current fuel needs.
Biodiesel, the most common replacement for fossil-fuel derived diesel, is a fatty ester derived from a combination of fatty acid (oils) and alcohol. Bacteria like E. coli can produce fatty acids as part of their metabolic pathway, but are unable to process it further to ester.
Engineered E. coli cells produce biodiesel from sugar (seen here as oil droplets surrounding the cell). Image credit: Marcin Zemla and Manfred Auer, JBEI Source: http://newscenter.lbl.gov/news-releases/2010/01/27/microbes-produce-biofuels/
Accumulation of fatty acids in the cell also leads to inhibition of this synthesis pathway. Purification of fatty acids from microbial cells, and chemical transesterification (reacting the fatty acid with alcohol outside the cell) can add significantly to the production costs.
One of the first synthetic biologists to address the need for alternative synthetic fuels is Jay Keasling, a Professor of Chemical Engineering, based at the University of California, Berkeley. Keasling and his colleagues have constructed an entirely novel metabolic pathway for biodiesel synthesis, by stitching together genes from ten different organisms onto a common chassis. Starting with simple sugars as the raw material, each of these genes catalyses a step in a multistep reaction, with the product of one reaction acting as a raw material for the next, ultimately resulting in the production of biodiesel.
To construct the chassis for this pathway, they first modified the genes of E. coli to manufacture fatty acids in large quantities. Next, they introduced a set of genes from the bacterium Zymomonas mobilis, which turns the chemical intermediates of respiration into ethanol. The gene from the yeast Saccharomyces cervisiae produces an enzyme that links the fatty acid and alcohol chains to form fatty esters (biodiesel). The final gene is the jigsaw is the xylanase gene from Bacteroides ovatus, which produces an enzyme capable of breaking down hemicellulose, a difficult-to-digest component of plant cells, into simple sugars that can be utilised as a precursor for both fatty acid and alcohol synthesis. The chassis containing the modified genes, called a ‘plasmid’, is inserted into E. coli cells. When supplied with 2% xylan, a hemicellulose carbon source, the cells are able to churn out 11.6 mg biodiesel per litre of growth medium, a yield of the order of magnitude required for industrial scale-up.
Designing a synthetic biology chassis for the production of biodiesel from sugar.Four key genes are involved – the xsa gene which digests hemicellulose found in plant cell walls; the TES-ACL-FAR genes which produce modified fatty acids, the pdc and adhB genes to produce alcohol, and the AT gene which links the fatty acid and alcohol together to form biodiesel.These genes are stitched together as a ‘plasmid’, which is then inserted into E. coli cells.Image adapted from: Steen et al., Nature, 2010, doi:10.1038/nature08721; and Steven K. Ritter, Chemical and Engineering News, 2008, http://tinyurl.com/bvf5oce.
In addition to producing biodiesel as a consolidated metabolic pathway in E. coli, Keasling also showed that it was possible to manipulate the length of the carbon chain in the biodiesel produced by the microorganisms. This is important for industrial applications, as petrol is made up of short chain hydrocarbons (a molecule made up entirely of carbon and hydrogen), around 8 carbon atoms in length, whereas diesel is primarily made up of longer hydrocarbon chains, 14 to 18 carbon atoms long. Keasling’s bacteria secrete the ‘drop-in’ biodiesel directly into the medium, where, being insoluble with water, it floats to the top and can be siphoned off. This, along with the ability to use previously hard-to-digest plant material, results in a saving of 80% in purification costs compared to conventional methods of purifying biodiesel.
Generating liquid sunshine, using bacterial co-cultures. The first bacterium converts the energy from light into chemical energy in the form of glucose. The second bacterium transfers the energy from glucose into the carbon-hydrogen bonds of biodiesel. Both bacterial strains are modified via synthetic biology to optimise their gene output.Image adapted from: Wackett Lab, University of Minnesota. http://biohydrocarbon.umn.edu/microbe.shtml
Other scientists are engaged in harnessing energy from the Sun for the production of energy-efficient, carbon-neutral biofuels. A group led by Professor Larry Wackett, of the University of Minnesota, is utilising a co-culture of two bacteria to achieve this goal—one bacterial species turns the energy from the Sun into chemical energy, and the other uses this chemical energy to produce fuel.
The first bacterium, Synechococcus, is a single-celled ‘green’ bacterium, commonly found in the marine environment, which possesses the pigments necessary to carry out photosynthesis, where the energy from the Sun is used to ‘fix’ atmospheric CO2 into organic molecules, including glucose. These sugars can be used an energy source for respiration by the bacterium Shewanella, which turns the chemical by-products of respiration into hydrocarbons-based oils, very like conventional petroleum-based fuels. The project has received substantial funding from the U.S. Department of Energy, based on its potential to transform ‘free’ energy from sunlight into useful petroleum-like fuels on an industrial scale.
Craig Venter, and his company Synthetic Genomics, have also turned their attention to the photosynthetic mechanisms in algae to produce biofuels. Unlike bacterial co-cultures, algae are capable of both photosynthesis, and converting the photosynthetic intermediates into lipid-based substances that can be turned into fuel. Algae will grow rapidly when supplied with mineral nutrients, water and sunlight, thus they eliminate the need for valuable land space. His company has to date received close to $300 million in funding from the oil giantExxonMobil, to optimise the production of biofuels from algae.
In both these processes outlined above, the CO2, which currently poses an environmental challenge, will be recycled to provide more energy. Venter calls this CO2 ‘the renewable feedstock of the future’. The algae or bacteria can be fed CO2 obtained from any source, including industrial plants and power stations. Though the carbon trapped in CO2 will eventually be discharged into the environment, each carbon molecule would have effectively been recycled twice for energy, thus paving the way for a truly carbon-neutral, and even a a carbon-negative, process.
Research into advanced biofuels is set to have significant environmental impact – the NNFCC estimates that replacing 4% of the fossil fuel consumed each year with advanced biofuels will result in a net saving of 3.2 million tonnes of CO2—equivalent to taking a million cars off the road!
This short video below, presented by Professor Keasling outlines the challenges facing biofuel production today, and how synthetic biologists are trying to address this problem.
Editor’s Note: Statistical amendments have been made to this article since its publication.
Hi Peter - synthetic biology might not be able to meet current fuel demands, but every car or jet engine that runs on biofuel reduces the one-way build-up of CO2 in the atmosphere. Not only do you end up recycling waste plant material, but also the carbon released from combustion of the biofuel - something that just cannot be achieved with fossil fuels at our current rate of growth. Deriving energy from algae is even more attractive, as they can grow in fresh or saline water (which represents 70% of our surface area), have a much faster growth cycle and a shorter harvesting cycle compared to plants (thus 'fixing' more CO2), and produce more fuel gram-per-gram as compared to plant material. People like Jay Keasling are very aware that the global demand for fuel is much higher than our current ability to meet it, and have assimilated this aim while designing synthetic biofuel pathways. What I describe here are only the initial results of the study - since this publication Keasling has improved the yield another 30-40% and has moved on from bacteria to yeast, which can produce larger quantities of biofuel - not anywhere near demand, but one step closer to the goal.
One correction 'UK alone consumes 200 tonnes of fuel' should read 'UK alone consumes 212 million tonnes' (DECC figs for 2010). This figure alone demonstrates why none of the biofuel solutions will ever be useful or in fact more than a rounding error in the overall fuel use. hey simply cannot scale up to the enormous quantities used. For example about 0.25 million litres of used vegetable oils are converted into bio diesel in UK per year which sounds like a lot until you work out that this is 208 tonnes or about a millionth of what we use. The only way forward is to reduce the usage. More localisation, less travel, better insulation and draught proofing, less meat more vegetables etc etc.
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