Research
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Synthetic Biology underpins advances in the bioeconomy
Biological systems - including the simplest cells - exhibit a broad range of functions to thrive in their environment. Research in the Imperial College Centre for Synthetic Biology is focused on the possibility of engineering the underlying biochemical processes to solve many of the challenges facing society, from healthcare to sustainable energy. In particular, we model, analyse, design and build biological and biochemical systems in living cells and/or in cell extracts, both exploring and enhancing the engineering potential of biology.
As part of our research we develop novel methods to accelerate the celebrated Design-Build-Test-Learn synthetic biology cycle. As such research in the Centre for Synthetic Biology highly multi- and interdisciplinary covering computational modelling and machine learning approaches; automated platform development and genetic circuit engineering ; multi-cellular and multi-organismal interactions, including gene drive and genome engineering; metabolic engineering; in vitro/cell-free synthetic biology; engineered phages and directed evolution; and biomimetics, biomaterials and biological engineering.
Publications
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Journal articleKis Z, Shattock R, Shah N, et al., 2019,
Correction: Emerging technologies for low‐cost, rapid vaccine manufacture
, Biotechnology Journal, Vol: 14, Pages: 1-2, ISSN: 1860-6768 -
Journal articleNikolados E, Weisse A, Ceroni F, et al., 2019,
Growth defects and loss-of-function in synthetic gene circuits
, ACS Synthetic Biology, Vol: 8, Pages: 1231-1240, ISSN: 2161-5063Synthetic gene circuits perturb the physiology of their cellular host. The extra load on endogenous processes shifts the equilibrium of resource allocation in the host, leading to slow growth and reduced biosynthesis. Here we built integrated host-circuit models to quantify growth defects caused by synthetic gene circuits. Simulations reveal a complex relation between circuit output and cellular capacity for gene expression. For weak induction of heterologous genes, protein output can be increased at the expense of growth defects. Yet for stronger induction, cellular capacity reaches a tipping point, beyond which both gene expression and growth rate drop sharply. Extensive simulations across various growth conditions and large regions of the design space suggest that the critical capacity is a result of ribosomal scarcity. We studied the impact of growth defects on various gene circuits and transcriptional logic gates, which highlights the extent to which cellular burden can limit, shape, and even break down circuit function. Our approach offers a comprehensive framework to assess the impact of host-circuit interactions in silico, with wide-ranging implications for the design and optimization of bacterial gene circuits.
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Journal articleBrittain R, Jones N, Ouldridge T, 2019,
Biochemical Szilard engines for memory-limited inference
, New Journal of Physics, Vol: 21, ISSN: 1367-2630By designing and leveraging an explicit molecular realisation of a measurement-and-feedback-powered Szilard engine, we investigate the extraction of work from complex environments by minimal machines with finite capacity for memory and decision-making. Living systems perform inference to exploit complex structure, or correlations, in their environment, but the physical limits and underlying cost/benefit trade-offs involved in doing so remain unclear. To probe these questions, we consider a minimal model for a structured environment—a correlated sequence of molecules—and explore mechanisms based on extended Szilard engines for extracting the work stored in these non-equilibrium correlations. We consider systems limited to a single bit of memory making binary 'choices' at each step. We demonstrate that increasingly complex environments allow increasingly sophisticated inference strategies to extract more free energy than simpler alternatives, and argue that optimal design of such machines should also consider the free energy reserves required to ensure robustness against fluctuations due to mistakes.
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SoftwareQuast N, Ouldridge T, 2019,
Simulation of DNA tile self-assembly
Submitted in accompaniment with final year report of MEng Project. DNA tile self-assembly is the spontaneous assembly of nano-structures made from short single-stranded DNA sequences. Successful assembly occurs within a narrow parameter window. Thisproject presents a model with which DNA self-assembly is simulated. Simulations for different tem-perature, sequence binding specificity and DNA tile concentrations indicate that: the growth rateof assemblies from uniform strand solutions is linear and highly temperature dependent; the aver-age nucleation times of assembly increase exponentially with temperature; high binding strengthsof boundary strands improve the stability of the complete assembly; locally high concentrations ofstrands seed the growth of the assembly; and locally low strand concentrations spatially direct thegrowth of the assembly. The source code is written in C.
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Journal articleVarghese F, Kabasakal BV, Cotton CA, et al., 2019,
A low-potential terminal oxidase associated with the iron-only nitrogenase from the nitrogen-fixing bacterium Azotobacter vinelandii
, Journal of Biological Chemistry, Vol: 294, Pages: 9367-9376, ISSN: 0021-9258The biological route for nitrogen gas entering the biosphere is reduction to ammonia by the nitrogenase enzyme, which is inactivated by oxygen. Three types of nitrogenase exist, the least studied of which is the iron-only nitrogenase. The Anf3 protein in the bacterium Rhodobacter capsulatus is essential for diazotrophic (i.e. nitrogen-fixing) growth with the iron-only nitrogenase, but its enzymatic activity and function are unknown. Here, we biochemically and structurally characterize Anf3 from the model diazotrophic bacterium Azotobacter vinelandii. Determining the Anf3 crystal structure to atomic resolution, we observed that it is a dimeric flavocytochrome with an unusually close interaction between the heme and the flavin adenine dinucleotide cofactors. Measuring the reduction potentials by spectroelectrochemical redox titration, we observed values of -420 ± 10 mV and -330 ± 10 mV for the two FAD potentials and -340 ± 1 mV for the heme. We further show that Anf3 accepts electrons from spinach ferredoxin and that Anf3 consumes oxygen without generating superoxide or hydrogen peroxide. We predict that Anf3 protects the iron-only nitrogenase from oxygen inactivation by functioning as an oxidase in respiratory protection, with flavodoxin or ferredoxin as the physiological electron donors.
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Journal articleMadsen C, Goni Moreno A, Palchick Z P U, et al., 2019,
Synthetic Biology Open Language Visual (SBOL Visual) version 2.1
, Journal of Integrative Bioinformatics, Vol: 16, Pages: 1-78, ISSN: 1613-4516People who are engineering biological organisms often find it useful to communicate in diagrams, both about the structure of the nucleic acid sequences that they are engineering and about the functional relationships between sequence features and other molecular species . Some typical practices and conventions have begun to emerge for such diagrams. The Synthetic Biology Open Language Visual (SBOL Visual) has been developed as a standard for organizing and systematizing such conventions in order to produce a coherent language for expressing the structure and function of genetic designs. This document details version 2.1 of SBOL Visual, which builds on the prior SBOL Visual 2.0 standard by expanding diagram syntax to include methods for showing modular structure and mappings between elements of a system, interactions arrows that can split or join (with the glyph at the split or join indicating either superposition or a chemical process), and adding new glyphs for indicating genomic context (e.g., integration into a plasmid or genome) and for stop codons.
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Journal articleKotidis P, Demis P, Goey C, et al., 2019,
Constrained global sensitivity analysis for bioprocess design space identification
, Computers and Chemical Engineering, Vol: 125, Pages: 558-568, ISSN: 1873-4375The manufacture of protein-based therapeutics presents unique challenges due to limited control over the biotic phase. This typically gives rise to a wide range of protein structures of varying safety and in vivo efficacy. Herein we propose a computational methodology, enabled by the application of constrained Global Sensitivity Analysis, for efficiently exploring the operatingrange of process inputs in silico and identifying a design space that meets output constraints. The methodology was applied to an antibody-producing Chinese hamster ovary (CHO) cell culture system: we explored >8000 feeding strategies to identify a subset of manufacturing conditions that meet constraints on antibody titre and glycan distribution as an attribute of product quality. Our computational findings were then verified experimentally, confirming the applicability of this approach to a challenging production system. We envisage that this methodology can significantly expedite bioprocess development and increase operational flexibility.
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Journal articleDet-Udom R, Gilbert C, Liu L, et al., 2019,
Towards semi-synthetic microbial communities: Enhancing soy sauce fermentation properties in B. subtilis co-cultures
, Microbial Cell Factories, Vol: 18, ISSN: 1475-2859BackgroundMany fermented foods and beverages are produced through the action of complex microbial communities. Synthetic biology approaches offer the ability to genetically engineer these communities to improve the properties of these fermented foods. Soy sauce is a fermented condiment with a vast global market. Engineering members of the microbial communities responsible for soy sauce fermentation may therefore lead to the development of improved products. One important property is the colour of soy sauce, with recent evidence pointing to a consumer preference for more lightly-coloured soy sauce products for particular dishes.ResultsHere we show that a bacterial member of the natural soy sauce fermentation microbial community, Bacillus, can be engineered to reduce the ‘browning’ reaction during soy sauce production. We show that two approaches result in ‘de-browning’: engineered consumption of xylose, an important precursor in the browning reaction, and engineered degradation of melanoidins, the major brown pigments in soy sauce. Lastly, we show that these two strategies work synergistically using co-cultures to result in enhanced de-browning.ConclusionsOur results demonstrate the potential of using synthetic biology and metabolic engineering methods for fine-tuning the process of soy sauce fermentation and indeed for many other natural food and beverage fermentations for improved products.
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Journal articleEllis T, 2019,
What is synthetic genomics anyway?
, The Biochemist, Vol: 41, Pages: 6-9, ISSN: 0954-982XYou may have heard of synthetic genomics. This headline-grabbing, high-profile, big science topic is starting to emerge catalysed by the pioneering work of famous names in synthetic biology and biotechnology like George Church and Craig Venter. But what is synthetic genomics and what is it being used for? As a prominent researcher at a recent UK meeting said: “Is it just synthetic biology with bigger bits of DNA?” Well no, not quite…
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Journal articleRajakumar PD, Gower G, Suckling L, et al., 2019,
Rapid prototyping platform for Saccharomyces cerevisiae using computer-aided genetic design enabled by parallel software and workcell platform development
, Slas Technology, Vol: 24, Pages: 291-297, ISSN: 2472-6303Biofoundries have enabled the ability to automate the construction of genetic constructs using computer-aided design. In this study, we have developed the methodology required to abstract and automate the construction of yeast-compatible designs. We demonstrate the use of our in-house software tool, AMOS, to coordinate with design software, JMP, and robotic liquid handling platforms to successfully manage the construction of a library of 88 yeast expression plasmids. In this proof-of-principle study, we used three fluorescent genes as proxy for three enzyme coding sequences. Our platform has been designed to quickly iterate around a design cycle of four protein coding sequences per plasmid, with larger numbers possible with multiplexed genome integrations in Saccharomyces cerevisiae. This work highlights how developing scalable new biotechnology applications requires a close integration between software development, liquid handling robotics, and protocol development.
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Journal articleLawrence J, Chang S, Rodriguez LC, et al., 2019,
Students go through the gears at the iGEM competition for engineering biology
, The Biochemist, Vol: 41, Pages: 58-61, ISSN: 0954-982XThe annual International Genetically Engineered Machine (iGEM) competition, represents an exciting opportunity for students to experience first-hand the potential of synthetic biology approaches to solve real-world problems. In this article, an iGEM team based at Imperial College London share some of the highlights from their participation in the 2018 iGEM event, including sharing their work at the annual Jamboree in Boston, Massachusetts.
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Journal articleWitmer K, Fraschka SAK, Vlachou D, et al., 2019,
Epigenetic regulation underlying Plasmodium berghei gene expression during its developmental transition from host to vector
, bioRxiv, ISSN: 2045-2322ABSTRACT Epigenetic regulation of gene expression is an important attribute in the survival and adaptation of the malaria parasite Plasmodium in its human host. Our understanding of epigenetic regulation of gene expression in Plasmodium developmental stages beyond asexual replication in the mammalian host is sparse. We used chromatin immune-precipitation (ChIP) and RNA sequencing to create an epigenetic and transcriptomic map of the murine parasite Plasmodium berghei development from asexual blood stages to male and female gametocytes, and finally, to ookinetes. We show that heterochromatin 1 (HP1) almost exclusively associates with variantly expressed gene families at subtelomeric regions and remains stable across stages and various parasite lines. Variant expression based on heterochromatic silencing is observed only in very few genes. In contrast, the active histone mark histone 3 Lysine 9 acetylation (H3K9ac) is found between heterochromatin boundaries and occurs as a sharp peak around the start codon for ribosomal protein genes. H3K9ac occupancy positively correlates with gene transcripts in asexual blood stages, male gametocytes and ookinetes. Interestingly, H3K9ac occupancy does not correlate with transcript abundance in female gametocytes. Finally, we identify novel DNA motifs upstream of ookinete-specific genes thought to be involved in transcriptional activation upon fertilization.
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Journal articlePerin G, Jones PR, 2019,
Economic feasibility and long-term sustainability criteria on the path to enable a transition from fossil fuels to biofuels.
, Current Opinion in Biotechnology, Vol: 57, Pages: 175-182, ISSN: 0958-1669Currently the production of liquid biofuels relies on plant biomass, which in turn depends on the photosynthetic conversion of light and CO2 into chemical energy. As a consequence, the process is renewable on a far shorter time-scale than its fossil counterpart, thus rendering a potential to reduce the environmental impact of the transportation sector. However, the global economy is not intensively pursuing this route, as current generation biofuel production does not meet two key criteria: (1) economic feasibility and (2) long-term sustainability. Herein, we argue that microalgal systems are valuable alternatives to consider, although it is currently technologically immature and therefore not possible to reach criterion 1, nor evaluate criterion 2. In this review we discuss the major limiting factors for this technology and highlight how further research efforts could be deployed to concretize an industrial reality.
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Journal articleHillson N, Caddick M, Cai Y, et al., 2019,
Building a global alliance of biofoundries
, NATURE COMMUNICATIONS, Vol: 10, Pages: 1-4, ISSN: 2041-1723 -
Journal articleGang S, Sharma S, Saraf M, et al., 2019,
Analysis of Indole-3-acetic Acid (IAA) Production in Klebsiella by LC-MS/MS and the Salkowski Method
, BIO-PROTOCOL, Vol: 9 -
Conference paperKelwick R, Webb AJ, Wang Y, et al., 2019,
ISEV2019 Abstract Book. PT09.10: Protease biomarker detection using functionalised bioplastic-based biosensors
, ISEV 2019, Publisher: Co-Action Publishing, ISSN: 2001-3078 -
Journal articleShaw W, Yamauchi H, Mead J, et al., 2019,
Engineering a model cell for rational tuning of GPCR signaling
, Cell, Vol: 177, Pages: 782-796.e27, ISSN: 0092-8674G protein-coupled receptor (GPCR) signaling is the primary method eukaryotes use to respond to specific cues in their environment. However, the relationship between stimulus and response for each GPCR is difficult to predict due to diversity in natural signal transduction architecture and expression. Using genome engineering in yeast, we constructed an insulated, modular GPCR signal transduction system to study how the response to stimuli can be predictably tuned using synthetic tools. We delineated the contributions of a minimal set of key components via computational and experimental refactoring, identifying simple design principles for rationally tuning the dose response. Using five different GPCRs, we demonstrate how this enables cells and consortia to be engineered to respond to desired concentrations of peptides, metabolites, and hormones relevant to human health. This work enables rational tuning of cell sensing while providing a framework to guide reprogramming of GPCR-based signaling in other systems.
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Journal articleDebalke S, Habtewold T, Duchateau L, et al., 2019,
The effect of silencing immunity related genes on longevity in a naturally occurring Anopheles arabiensis mosquito population from southwest Ethiopia
, Parasites & Vectors, Vol: 12, ISSN: 1756-3305BackgroundVector control remains the most important tool to prevent malaria transmission. However, it is now severely constrained by the appearance of physiological and behavioral insecticide resistance. Therefore, the development of new vector control tools is warranted. Such tools could include immunization of blood hosts of vector mosquitoes with mosquito proteins involved in midgut homeostasis (anti-mosquito vaccines) or genetic engineering of mosquitoes that can drive population-wide knockout of genes producing such proteins to reduce mosquito lifespan and malaria transmission probability.MethodsTo achieve this, candidate genes related to midgut homeostasis regulation need to be assessed for their effect on mosquito survival. Here, different such candidate genes were silenced through dsRNA injection in the naturally occurring Anopheles arabiensis mosquitoes and the effect on mosquito survival was evaluated.ResultsSignificantly higher mortality rates were observed in the mosquitoes silenced for FN3D1 (AARA003032), FN3D3 (AARA007751) and GPRGr9 (AARA003963) genes as compared to the control group injected with dsRNA against a non-related bacterial gene (LacZ). This observed difference in mortality rate between the candidate genes and the control disappeared when gene-silenced mosquitoes were treated with antibiotic mixtures, suggesting that gut microbiota play a key role in the observed reduction of mosquito survival.ConclusionsWe demonstrated that interference with the expression of the FN3D1, FN3D3 or GPRGr9 genes causes a significant reduction of the longevity of An. arabiensis mosquito in the wild.
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Journal articleGoey CH, Bell D, Kontoravdi C, 2019,
CHO cell cultures in shake flasks and bioreactors present different host cell protein profiles in the supernatant
, Biochemical Engineering Journal, Vol: 144, Pages: 185-192, ISSN: 1369-703XSeveral studies on the impact of cell culture parameters on the profile of host cell protein (HCP) impurities have been carried out in shake flasks. Herein, we explore how transferable the findings and conclusions of such investigations are to lab-scale bioreactors. Experiments were performed in both systems in fed-batch mode under physiological temperature and with a shift to mild hypothermia and the impact on key upstream performance indicators was quantified. Under both temperatures, bioreactors produced a richer HCP pool despite the overall concentration being similar at both scales and temperatures. The number of different HCPs detected in bioreactor supernatants was four times higher than that in flasks under physiological temperature and more than eight times higher under mild hypothermia. The origin of HCPs was also altered from mostly naturally secreted proteins in flasks to mainly intracellular proteins in bioreactors at the lower temperature. Although the number of species correlated with apoptotic cell density in bioreactors, this was not the case in flasks. Even though the level of HCP impurities and mAb/HCP concentration ratio were similar under all four conditions with an average of approximately 330 μg HCP/mL culture and 0.3 mg HCP/mg IgG4, respectively, the fact that culture method significantly affects the number of species present in the supernatant can have implications for downstream processing steps.
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Journal articleStopnitzky E, Still S, Ouldridge TE, et al., 2019,
Physical limitations of work extraction from temporal correlations.
, Physical Review E, Vol: 99, ISSN: 1539-3755Recently proposed information-exploiting systems extract work from a single heat bath by using temporal correlations on an input tape. We study how enforcing time-continuous dynamics, which is necessary to ensure that the device is physically realizable, constrains possible designs and drastically diminishes efficiency. We show that these problems can be circumvented by means of applying an external, time-varying protocol, which turns the device from a "passive," free-running machine into an "actively" driven one.
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