Synthetic Biology: Hope or Hype?
Products like beer, wine and yoghurt would not be possible without microorganisms. For thousands of years humans have exploited these 'cell-factories' in industry for the production of many commercial goods. Increased knowledge of microbes, such as Baker's yeast Saccharomyces cerevisiae and the bacterium Escherichia coli, has allowed scientists to manipulate these organisms for use in the biotechnology industry. Advancements in DNA sequencing (technology used to analyse DNA), the ease of gene synthesis and our knowledge of genetics has opened the doors to a whole new age of (bio)synthesis, which has impacted a diverse range of industries including pharmaceuticals, biomedicine, fine chemicals, biofuels and biopolymers. The advancements in these areas have paved the way for synthetic biology.
Synthetic biology is the design of new biological systems using DNA or redesign of existing systems through altering DNA of various components within a host organism. Because DNA is a universal language across all life, taking a line of code from one species to another can in theory have the same effect. Synthetic biology requires the use of DNA manipulation in order to programme organisms to make products they don't make in nature. Hence, these organisms become living factories that produce commercially valuable products or therapeutics using cheap and renewable biomass as feedstock. This article serves to introduce the principles of synthetic biology and discuss its diverse applications: present and future.
Although a promising area of research with the potential of a huge impact on our economy, synthetic biology is still in its infancy. However, the potential benefits of the newly emerging synthetic biology technologies has been recognised by governments, with the UK and USA both designating this research as a national priority in funding investment. For example, in the United Kingdom, it has been identified as one of the eight great British technologies of the future. Through the use of biological systems, synthetic biology promises a greener, cheaper and more efficient alternative to the traditional methods currently employed in the industry, such as extraction from natural sources and chemical synthesis. The latter often requires harsh environmental conditions, such as the use of strong acids and bases, high temperatures and pressures, expensive metal catalysts and large number of reaction and purification steps. Obvious implications of synthetic biology for the UK economy and biomedical and biotechnology industries has resulted in recent major funding allocations for the set-up of several specialised synthetic biology centers across the country in Manchester, Nottingham, Cambridge and Bristol as well as another two yet centers to be announced.
DNA is the software encoding the characteristics and processes of every cell. This means the function and identity of each cell within an organism is determined by the genes that are active or turned on in the cell. As such, synthetic biologists use DNA-level engineering to 'reprogramme' host organisms to produce new compound(s). You could reengineer the bacteria E. coli to make menthol (mint flavor) by synthesizing the mint plant DNA responsible for menthol production and delivering it into E. coli. This is illustrated in Figure 1, below. To programme the cells, functional DNA assemblies and circuits are constructed using individual components (e.g. elements that control when a gene is turned on and to what extent it is on) known as BioBrick parts, just like using Lego(TM) building blocks. Each component serves a function that affects the eventual output (product, cell behaviour, signal etc.)
Figure 1 - DNA constructs containing genes of interest are assembled and delivered into the host organisms such as E-coli. These pathways are designed to sense environmental conditions to actuate responses.
Although far from perfect, E. coli and S. cerevisiae are the prevalent choice of host-organisms, as genetic manipulation is relatively easy. Organisms such as cyanobacteria that have minimal feedstock requirements are ideal alternative factories for scaling-up production, however, these organisms are not as well known on a genetic level. When adding new genes into an organism, the challenge is to avoid disrupting the other natural processes vital to its survival. Further challenges are the engineered organism's survival against evolutionary pressures, circumventing toxicity to host organisms and balancing organism's energy requirements. Scaling up the processes from lab bench to industrial scale application presents further challenges (2-4). To ensure that engineered pathways do not disturb the host organism's own biochemical pathways, research is being conducted into engineering of minimal 'chassis' host organism, containing only the bare minimum genome required for its survival. John Craig Venter Institute has already successfully made synthetic life, dubbed Synthia. This institute is currently working to synthesise a 'skeleton' organism named Mycoplasma laboratorium using only a minimal set of genes from Mycoplasma genitalium (found to have only 382 essential genes through gene deletion and inactivation studies) (5, 6).
Applications in the Pharmaceutical industry:
There is extensive synthetic biology research activity in the area of terpenoid production. Terpenoids are the largest and most diverse class of metabolite-compounds made and used in reactions that keep organisms alive- found extensively in nature. These compounds include herbivore-repellents, pollinator-attractants, phototoxins and photoreceptive agents such as carotenoids and sterols (7, 8). The drug Paclitaxel is a terpenoid extracted from the bark of the pacific yew trees and is used in chemotherapy treatment. The native trees take 200 years to grow to only 40ft of height. The bark from one-40ft tree yields 0.5g of taxol, and as 3g of this drug is required for a single treatment; the demand of this drug exponentially exceeds supply (9). Furthermore, the distillation and fractionation steps required to extract these compounds from cells are costly, making the treatment very expensive. A taxol precursor, taxadiene, has been produced successfully at around 1 g/L using a synthetic biology approach (10). This is an improvement upon native sources, however further optimization of the product yield is needed for industrial scale production. This would result in increased availability of such drugs at lower costs, relieving the financial strain of such treatments on healthcare institutions like the UK's National Health Service (NHS).
Another prominent example is the anti-malarial drug, artemisinin. Artemisinin is extracted from a sweet wormwood plant called Artimisinua annua. The Keasling laboratory in Berkeley, California have designed and implemented an industrial scale production of artemisinin using the synthetic biology approach through funding from Bill and Melinda Gates foundation. Artemisinin is hailed as a synthetic biology success since the lab demonstrated production of a significant amount - 25g/L - of the artemisinin precursor called artemisinic acid (11). Pharma-giant Sanofi has licensed the engineered S. cerevisae strain for industrial scale semi-synthetic production of artemisinic acid, which is then chemically converted to artesunate for use in combination therapy for malaria. In developing countries, malaria affects up to 200 million people every year, claiming the lives of 650,000 (mostly children) every year. To make this drug available at lower costs, the Keasling lab agreed to royalty-free licensing of the engineered strain to Sanofi. Sanofi use the semi-synthetic route to produce artemisinin using no profit, no-loss production model to maintain a low cost for the treatment (12). Therefore, synthetic biology has the potential to make a huge impact on the pharmaceutical industry in revolutionising the way drugs of the future are made.
Multi-drug resistance is among one of the biggest challenges of the 21st Century. By 2050, drug resistant infections could cost the global economy up to $100 trillion, set to claim the lives of 10 million people every year - killing more people than cancer (13). This is of a huge concern because the overuse of antibiotics leads to an acceleration of levels of drug-resistant pathogens. Synthetic Biology enables the design of new treatment methods to target bacterial biofilms, potentiation of current antibiotics and engineering of new antibiotics. Multi-drug resistance can be targeted through the design of circuitry that switch off bacterial resistance, engineering designer phages (organisms that kill bacteria) that destroy biofilms and reprogramming probiotic bacteria that can fight pathogens (14, 15).
Fine Chemicals for use in cosmetics, flavourings and fragrances:
Monoterpenoids (a class of terpenoids) include compounds such as limonene, which is a precursor to menthols and carvone. Limonene naturally found in citrus peels can is used in almost all cosmetics and toiletries. Menthols are valuable to the flavouring, fragrance and cosmetic industry. There is an annual turnover of 7000 tonnes of menthol, resulting in the net revenue of US $300 million which makes these compounds commercially valuable (16).
Menthol constitutes 0.5% of the dry-weight of mint leaves and the proportion of the different types of menthol can vary greatly between harvests depending on environmental conditions. Processes that are used to extract menthol from leaves are very costly (8, 16). Chemical synthesis routes require the use of expensive catalysts and several purification steps for the separation of a single type of menthol (7).
Natural biochemical pathways for the production of such commercially valuable compounds can often be found in nature. These pathways usually occur at mild conditions - those needed for a mint plant to grow. Furthermore, biochemical engineering can be used to produce such pure compounds in high quantities. This makes synthetic biology a greener, more efficient and cheaper alternative to chemical synthesis for the production of fine chemicals (17). It can revolutionise our chemical industries by enabling the move away from petrochemicals.
Through putting the biosynthetic pathway of mint that is responsible for menthol production into E. coli and fine-tuning it, synthetic biologists may be able to produce pure menthol at greater concentrations than are available from natural sources, at minimal expenses. Limonene is the precursor to menthols and its anti-microbial properties make it toxic to E. coli; thus, the organism removes this compound as soon as it is produced. Hence, there is less of it available to be converted to menthol. Synthetic biologists face such problems when engineering organisms to make products that it does not produce in nature. Large-scale production of menthols through this route is thus commercially desirable for the fragrance, flavour and cosmetics industry (7, 17).
Other applications in the field of Biomedicine and Biofuels:
Synthetic biology can be used to engineer biological based sensors called biosensors that can sense, analyse and activate under in-vivo conditions i.e. conditions within living organisms such as hallmarks of cancer, inflammatory responses and changes resultant from bacterial infections. One such example of an engineered biosensor is the University of Cambridge iGEM team's project "E.chromi", which won the 2009 iGEM competition (18). The team engineered E. coli to express different pigments dependent on the in vivo conditions (18). This has the potential to be used for diagnostic purposes for a range of gastrointestinal diseases. Hence, synthetic biology opens the door to a completely new class of diagnostics. Another example is a protein from tissue necrotising organism (organism that destroys the human cells and tissues) Streptococcus pyogenes. The protein has two parts that search for each other and bind to each other with extremely strong covalent bonds. Research will be conducted to engineer this protein for the detection of circulating cancer cells and serve to diagnose the spread of cancer cells.
Another key area of synthetic biology research is biofuels. Extensive research has been conducted for the production of second and third generation biofuels that have high-energy content and are compatible with the current engine infrastructure. Higher alcohols and fatty-acid derived fuels and hydrocarbons are under focus. However, there are numerous hurdles to be addressed such as host-toxicity. Further exciting developments are coming forth in the production of bioplastics via synthetic biology approaches to challenge the dominance of naptha-based plastics.
Synthetic biology is a burgeoning area of research with a promising future. The development of this exciting technology in the UK is at forefront as evident by the extensive funding plan implemented by the UK. As synthetic biology is an emerging research area, it is difficult to predict the ways in which it will have the most impact on the sectors discussed above. Industry interest is of paramount importance to the development in this area, which is likely to occur if the technologies resulting from synthetic biology compete in terms of costs and efficiency with the existing technologies employed by the industry. The continued research interest in this area promises significant advancements and revolution for a diverse range of industries in the coming years as well as huge impact on national and international economies. Despite the many potential benefits we can reap from synthetic biology research, we must assess the risks associated with making and manipulating organisms. An example would be terrorists making bioweapons constructing dangerous, virulent organisms such as small pox or Spanish flu. There may be the rare risk of accidental production of something dangerous that the designer is unable to control. Furthermore, recent publications on production of opiates using synthetic biology have raised concerns over the "DIY" approach being abused by illegal-users of opiates. Therefore, careful consideration needs to be given to account for responsible research and innovation.