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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.

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  • Journal article
    Lv X, Wu Y, Tian R, Gu Y, Liu Y, Li J, Du G, Ledesma-Amaro R, Liu Let al., 2020,

    Synthetic metabolic channel by functional membrane microdomains for compartmentalized flux control

    , Metabolic Engineering, Vol: 59, Pages: 106-118, ISSN: 1096-7176

    The anchoring of metabolic pathway enzymes to spatial scaffolds can significantly improve their reaction efficiency. Here, we successfully constructed a multi-enzyme complex assembly system able to enhance bioproduction in bacteria by using the endogenous spatial scaffolds─functional membrane microdomains (FMMs). First, using VA-TIRFM and SPT analysis, we reveal that FMMs possess high temporal and spatial stability at the plasma membrane and can be used as endogenous spatial scaffolds to organize enzyme pathways. Then, taking the synthesis of N-acetylglucosamine (GlcNAc) in Bacillus subtilis as a proof-of-concept demonstration, we found that anchoring of various enzymes required for GlcNAc synthesis onto FMMs to obtain the FMMs-multi-enzyme complex system resulted in a significant increase in GlcNAc titer and an effectively alleviate in cell lysis at the later stage of fermentation compared to that in control strains expressing the related enzymes in the cytoplasm. Combining with metabolic model and kinetics analysis, the existence of a constructed substrate channel that maximizes the reaction efficiency is verified. In summary, we propose a novel metabolic pathway assembly model which allowed improved titers and compartmentalized flux control with high spatial resolution in bacterial metabolism.

  • Journal article
    Wilkinson MD, Lai H-E, Freemont PS, Baum Jet al., 2020,

    A biosynthetic platform for antimalarial drug discovery

    , Antimicrobial Agents and Chemotherapy, Vol: 64, Pages: 1-9, ISSN: 0066-4804

    Advances in synthetic biology have enabled production of a variety of compounds using bacteria as a vehicle for complex compound biosynthesis. Violacein, a naturally occurring indole pigment with antibiotic properties, can be biosynthetically engineered in Escherichia coli expressing its non-native synthesis pathway. To explore whether this synthetic biosynthesis platform could be used for drug discovery, here we have screened bacterially-derived violacein against the main causative agent of human malaria, Plasmodium falciparum. We show the antiparasitic activity of bacterially-derived violacein against the P. falciparum 3D7 laboratory reference strain as well as drug-sensitive and resistant patient isolates, confirming the potential utility of this drug as an antimalarial. We then screen a biosynthetic series of violacein derivatives against P. falciparum growth. The demonstrated varied activity of each derivative against asexual parasite growth points to potential for further development of violacein as an antimalarial. Towards defining its mode of action, we show that biosynthetic violacein affects the parasite actin cytoskeleton, resulting in an accumulation of actin signal that is independent of actin polymerization. This activity points to a target that modulates actin behaviour in the cell either in terms of its regulation or its folding. More broadly, our data show that bacterial synthetic biosynthesis could become a suitable platform for antimalarial drug discovery with potential applications in future high-throughput drug screening with otherwise chemically-intractable natural products.

  • Journal article
    Debalke S, Habtewold T, Christophides GK, Duchateau Let al., 2020,

    Stability of the effect of silencing fibronectin type III domain-protein 1 (FN3D1) gene on Anopheles arabiensis reared under different breeding site conditions

    , Parasites and Vectors, Vol: 13, Pages: 1-9, ISSN: 1756-3305

    BackgroundMalaria vector mosquitoes acquire midgut microbiota primarily from their habitat. The homeostasis of these microbial communities plays an essential role in the mosquito longevity, the most essential factor in the mosquito vectorial capacity. Our recent study revealed that silencing genes involved in regulation of the midgut homeostasis including FN3D1, FN3D3 and GPRGr9 reduced the survival of female adult Anopheles arabiensis mosquitoes. In the present study, we investigate the stability of the gene silencing efficiency of mosquitoes reared in three different breeding conditions representing distinct larval habitat types: town brick pits in Jimma, flood pools in the rural land of Asendabo and roadside pools in Wolkite.MethodsFirst-instar larvae of An. arabiensis mosquitoes were reared separately using water collected from the three breeding sites. The resulting adult females were micro-injected with dsRNA targeting the FN3D1 gene (AARA003032) and their survival was monitored. Control mosquitoes were injected with dsRNA Lacz. In addition, the load of midgut microbiota of these mosquitoes was determined using flow cytometry.ResultsSurvival of naïve adult female mosquitoes differed between the three sites. Mosquitoes reared using water collected from brick pits and flood pools survived longer than mosquitoes reared using water collected from roadside. However, the FN3D1 gene silencing effect on survival did not differ between the three sites.ConclusionsThe present study revealed that the efficacy of FN3D1 gene silencing is not affected by variation in the larval habitat. Thus, silencing this gene has potential for application throughout sub-Saharan Africa.

  • Journal article
    Moore SJ, Lai H-E, Kelwick RJR, Chee SM, Bell DJ, Polizzi KM, Freemont PSet al., 2020,

    Correction to EcoFlex: a multifunctional MoClo kit for E. coli synthetic biology.

    , ACS Synthetic Biology, ISSN: 2161-5063

    It has been brought to our attention that the original article contains a typographical error within Figure 1B, part ii. One of the 4-bp overhangs reads “GGAC” and should instead be “GTAC”, as is consistent throughout the original manuscript and deposited AddGene sequences.

  • Journal article
    Misirli G, Beal J, Gorochowski TE, Stan G-B, Wipat A, Myers CJet al., 2020,

    SBOL visual 2 ontology

    , ACS Synthetic Biology, Vol: 9, Pages: 972-977, ISSN: 2161-5063

    Standardizing the visual representation of genetic parts and circuits is essential for unambiguously creating and interpreting genetic designs. To this end, an increasing number of tools are adopting well-defined glyphs from the Synthetic Biology Open Language (SBOL) Visual standard to represent various genetic parts and their relationships. However, the implementation and maintenance of the relationships between biological elements or concepts and their associated glyphs has up to now been left up to tool developers. We address this need with the SBOL Visual 2 Ontology, a machine-accessible resource that provides rules for mapping from genetic parts, molecules, and interactions between them, to agreed SBOL Visual glyphs. This resource, together with a web service, can be used as a library to simplify the development of visualization tools, as a stand-alone resource to computationally search for suitable glyphs, and to help facilitate integration with existing biological ontologies and standards in synthetic biology.

  • Journal article
    Witmer K, Fraschka S, Vlachou D, Bartfai R, Christophides Get al., 2020,

    An epigenetic map of malaria parasite development from host to vector

    , Scientific Reports, Vol: 10, ISSN: 2045-2322

    The malaria parasite replicates asexually in the red blood cells of its vertebrate host employing epigenetic mechanisms to regulate gene expression in response to changes in its environment. We used chromatin immunoprecipitation followed by sequencing in conjunction with RNA sequencing to create an epigenomic and transcriptomic map of the developmental transition from asexual blood stages to male and female gametocytes and to ookinetes in the rodent malaria parasite Plasmodium berghei. Across the developmental stages examined, heterochromatin protein 1 associates with variantly expressed gene families localised at subtelomeric regions and variant gene expression based on heterochromatic silencing is observed only in some genes. Conversely, the euchromatin mark histone 3 lysine 9 acetylation (H3K9ac) is abundant in non-heterochromatic regions across all developmental stages. H3K9ac presents a distinct pattern of enrichment around the start codon of ribosomal protein genes in all stages but male gametocytes. Additionally, H3K9ac occupancy positively correlates with transcript abundance in all stages but female gametocytes suggesting that transcription in this stage is independent of H3K9ac levels. This finding together with known mRNA repression in female gametocytes suggests a multilayered mechanism operating in female gametocytes in preparation for fertilization and zygote development, coinciding with parasite transition from host to vector.

  • Journal article
    Evans SW, Beal J, Berger K, Bleijs DA, Cagnetti A, Ceroni F, Epstein GL, Garcia-Reyero N, Gillum DR, Harkess G, Hillson NJ, Hogervorst PAM, Jordan JL, Lacroix G, Moritz R, ÓhÉigeartaigh SS, Palmer MJ, van Passel MWJet al., 2020,

    Embrace experimentation in biosecurity governance

    , Science, Vol: 368, Pages: 138-140, ISSN: 0036-8075
  • Journal article
    Redhai S, Pilgrim C, Gaspar P, van Giesen L, Lopes T, Riabinina O, Grenier T, Milona A, Chanana B, Swadling J, Wang Y-F, Dahalan F, Yuan M, Wilsch-Brauninger M, Lin W-H, Dennison N, Capriotti P, Lawniczak M, Baines R, Warnecke T, Windbichler N, Leulier F, Bellono N, Miguel-Aliaga Iet al., 2020,

    An intestinal zinc sensor regulates food intake and developmental growth

    , Nature, Vol: 580, Pages: 263-268, ISSN: 0028-0836

    In cells, organs and whole organisms, nutrient sensing is key to maintaining homeostasis and adapting to a fluctuating environment1. In many animals, nutrient sensors are found within the enteroendocrine cells of the digestive system; however, less is known about nutrient sensing in their cellular siblings, the absorptive enterocytes1. Here we use a genetic screen in Drosophila melanogaster to identify Hodor, an ionotropic receptor in enterocytes that sustains larval development, particularly in nutrient-scarce conditions. Experiments in Xenopus oocytes and flies indicate that Hodor is a pH-sensitive, zinc-gated chloride channel that mediates a previously unrecognized dietary preference for zinc. Hodor controls systemic growth from a subset of enterocytes—interstitial cells—by promoting food intake and insulin/IGF signalling. Although Hodor sustains gut luminal acidity and restrains microbial loads, its effect on systemic growth results from the modulation of Tor signalling and lysosomal homeostasis within interstitial cells. Hodor-like genes are insect-specific, and may represent targets for the control of disease vectors. Indeed, CRISPR–Cas9 genome editing revealed that the single hodor orthologue in Anopheles gambiae is an essential gene. Our findings highlight the need to consider the instructive contributions of metals—and, more generally, micronutrients—to energy homeostasis.

  • Journal article
    Trinugroho JP, Beckova M, Shao S, Yu J, Zhao Z, Murray JW, Sobotka R, Komenda J, Nixon PJet al., 2020,

    Chlorophyll f synthesis by a super-rogue photosystem II complex (vol 6, pg 238, 2020)

    , NATURE PLANTS, Vol: 6, Pages: 427-427, ISSN: 2055-026X
  • Journal article
    James SL, Marshall JM, Christophides GK, Okumu FO, Nolan Tet al., 2020,

    Toward the definition of efficacy and safety criteria for advancing gene drive-modified mosquitoes to field testing

    , Vector-Borne and Zoonotic Diseases, Vol: 20, Pages: 237-251, ISSN: 1530-3667

    Mosquitoes containing gene drive systems are being developed as complementary tools to prevent transmission of malaria and other mosquito-borne diseases. As with any new tool, decision makers and other stakeholders will need to balance risks (safety) and benefits (efficacy) when considering the rationale for testing and deploying gene drive-modified mosquito products. Developers will benefit from standards for judging whether an investigational gene drive product meets acceptability criteria for advancing to field trials. Such standards may be formalized as preferred product characteristics and target product profiles, which describe the desired attributes of the product category and of a particular product, respectively. This report summarizes discussions from two scientific workshops aimed at identifying efficacy and safety characteristics that must be minimally met for an investigational gene drive-modified mosquito product to be deemed viable to move from contained testing to field release and the data that will be needed to support an application for first field release.

  • Journal article
    Thaore V, Tsourapas D, Shah N, Kontoravdi Cet al., 2020,

    Techno-Economic Assessment of Cell-Free Synthesis of Monoclonal Antibodies Using CHO Cell Extracts

    , PROCESSES, Vol: 8
  • Journal article
    Vivek A, Bolognesi G, Elani Y, 2020,

    Fusing artificial cell compartments and lipid domains using optical traps: a tool to modulate membrane composition and phase behaviour

    , Micromachines, Vol: 11, ISSN: 2072-666X

    New technologies for manipulating biomembranes have vast potential to aid the understanding of biological phenomena, and as tools to sculpt novel artificial cell architectures for synthetic biology. The manipulation and fusion of vesicles using optical traps is amongst the most promising due to the level of spatiotemporal control it affords. Herein, we conduct a suite of feasibility studies to show the potential of optical trapping technologies to (i) modulate the lipid composition of a vesicle by delivering new membrane material through fusion events and (ii) manipulate and controllably fuse coexisting membrane domains for the first time. We also outline some noteworthy morphologies and transitions that the vesicle undergoes during fusion, which gives us insight into the mechanisms at play. These results will guide future exploitation of laser-assisted membrane manipulation methods and feed into a technology roadmap for this emerging technology.

  • Journal article
    Ukegbu CV, Giorgalli M, Tapanelli S, Rona LDP, Jaye A, Wyer C, Angrisano F, Christophides G, Vlachou Det al., 2020,

    PIMMS43 is required for malaria parasite immune evasion and sporogonic development in the mosquito vector

    , Proceedings of the National Academy of Sciences of USA, Vol: 117, Pages: 7363-7373, ISSN: 0027-8424

    After being ingested by a female Anopheles mosquito during a bloodmeal on an infected host, and before they can reach the mosquito salivary glands to be transmitted to a new host, Plasmodium parasites must establish an infection of the mosquito midgut in the form of oocysts. To achieve this, they must first survive a series of robust innate immune responses that take place prior to, during, and immediately after ookinete traversal of the midgut epithelium. Understanding how parasites may evade these responses could highlight new ways to block malaria transmission. We show that an ookinete and sporozoite surface protein designated as PIMMS43 (Plasmodium Infection of the Mosquito Midgut Screen 43) is required for parasite evasion of the Anopheles coluzzii complement-like response. Disruption of PIMMS43 in the rodent malaria parasite Plasmodium berghei triggers robust complement activation and ookinete elimination upon mosquito midgut traversal. Silencing components of the complement-like system through RNAi largely restores ookinete-to-oocyst transition but oocysts remain small in size and produce a very small number of sporozoites that additionally are not infectious, indicating that PIMMS43 is also essential for sporogonic development in the oocyst. Antibodies that bind PIMMS43 interfere with parasite immune evasion when ingested with the infectious blood meal and significantly reduce the prevalence and intensity of infection. PIMMS43 genetic structure across African Plasmodium falciparum populations indicates allelic adaptation to sympatric vector populations. These data add to our understanding of mosquito–parasite interactions and identify PIMMS43 as a target of malaria transmission blocking.

  • Journal article
    Ledesma-Amaro R, Nikel PI, Ceroni F, 2020,

    Editorial: synthetic biology-guided metabolic engineering

    , Frontiers in Bioengineering and Biotechnology, Vol: 8, ISSN: 2296-4185
  • Journal article
    Zielonka D, Witkowski G, Puch EA, Lesniczak M, Mazur-Michalek I, Isalan M, Mielcarek Met al., 2020,

    Prevalence of non-psychiatric comorbidities in pre-symptomatic and symptomatic Huntington's disease gene carriers in Poland

    , Frontiers in Medicine, Vol: 7, ISSN: 2296-858X

    Huntington's disease (HD) is monogenic neurodegenerative disorder caused by CAG expansions within the Huntingtin gene (Htt); it has a prevalence of 1 in 10,000 worldwide and is invariably fatal. Typically, healthy individuals have fewer than 35 CAG repeats, while the CAG expansions range from 36 to ~200 in HD patients. The hallmark of HD is neurodegeneration, especially in the striatal nuclei, basal ganglia and cerebral cortex, leading to neurological symptoms that involve motor, cognitive, and psychiatric events. However, HD is a complex disorder that may also affect peripheral organs, so it is possible that HD patients could be affected by comorbidities. Hence, we investigated the prevalence of comorbid conditions in HD patients (pre-symptomatic and symptomatic groups) and compared the frequency of those conditions to a control group. Our groups represent 65% of HD gene carriers registered in Poland. We identified 8 clusters of comorbid conditions in both HD groups, namely: musculoskeletal, allergies, cardiovascular, neurological, gastrointestinal, thyroid, psychiatric, and ophthalmologic. We found that HD patients have a significantly higher percentage of co-existing conditions in comparison to the control group. Among the 8 clusters of diseases, musculoskeletal, psychiatric, and cardiovascular events were significantly more frequent in both pre- and symptomatic HD patients, while neurological and gastrointestinal clusters showed significantly higher occurrence in the HD symptomatic group. A greater recognition of comorbidity in HD might help to better understand health outcomes and improve clinical management.

  • Journal article
    Trinugroho J, Bečková M, Shao S, Yu J, Zhao Z, Murray JW, Sobotka R, Komenda J, Nixon PJet al., 2020,

    Chlorophyll f synthesis by a super-rogue photosystem II complex

    , Nature Plants, Vol: 6, Pages: 238-244, ISSN: 2055-0278

    Certain cyanobacteria synthesize chlorophyll molecules (Chl d and Chl f) that absorb in the far-red region of the solar spectrum, thereby extending the spectral range of photosynthetically active radiation1,2. The synthesis and introduction of these far-red chlorophylls into the photosynthetic apparatus of plants might improve the efficiency of oxygenic photosynthesis, especially in far-red enriched environments, such as in the lower regions of the canopy3. Production of Chl f requires the ChlF subunit, also known as PsbA4 (ref. 4) or super-rogue D1 (ref. 5), a paralogue of the D1 subunit of photosystem II (PSII) which, together with D2, bind cofactors involved in the light-driven oxidation of water. Current ideas suggest that ChlF oxidizes Chl a to Chl f in a homodimeric ChlF reaction centre (RC) complex and represents a missing link in the evolution of the heterodimeric D1/D2 RC of PSII (refs. 4,6). However, unambiguous biochemical support for this proposal is lacking. Here, we show that ChlF can substitute for D1 to form modified PSII complexes capable of producing Chl f. Remarkably, mutation of just two residues in D1 converts oxygen-evolving PSII into a Chl f synthase. Overall, we have identified a new class of PSII complex, which we term ‘super-rogue’ PSII, with an unexpected role in pigment biosynthesis rather than water oxidation.

  • Journal article
    McCarty NS, Graham AE, Studená L, Ledesma-Amaro Ret al., 2020,

    Multiplexed CRISPR technologies for gene editing and transcriptional regulation

    , Nature Communications, Vol: 11, ISSN: 2041-1723

    Multiplexed CRISPR technologies, in which numerous gRNAs or Cas enzymes are expressed at once, have facilitated powerful biological engineering applications, vastly enhancing the scope and efficiencies of genetic editing and transcriptional regulation. In this review, we discuss multiplexed CRISPR technologies and describe methods for the assembly, expression and processing of synthetic guide RNA arrays in vivo. Applications that benefit from multiplexed CRISPR technologies, including cellular recorders, genetic circuits, biosensors, combinatorial genetic perturbations, large-scale genome engineering and the rewiring of metabolic pathways, are highlighted. We also offer a glimpse of emerging challenges and emphasize experimental considerations for future studies.

  • Journal article
    Fasulo B, Meccariello A, Morgan M, Borufka C, Papathanos PA, Windbichler Net al., 2020,

    A fly model establishes distinct mechanisms for synthetic CRISPR/Cas9 sex distorters

    , PLOS GENETICS, Vol: 16, ISSN: 1553-7404
  • Journal article
    Rodgers FH, Cai JA, Pitaluga AN, Mengin-Lecreulx D, Gendrin M, Christophides GKet al., 2020,

    Functional analysis of the three major PGRPLC isoforms in the midgut of the malaria mosquito Anopheles coluzzii

    , INSECT BIOCHEMISTRY AND MOLECULAR BIOLOGY, Vol: 118, ISSN: 0965-1748
  • Journal article
    Papathanasiou MM, Kontoravdi C, 2020,

    Engineering challenges in therapeutic protein product and process design

    , Current Opinion in Chemical Engineering, Vol: 27, Pages: 81-88, ISSN: 2211-3398

    Biologics represent the fastest growing sector of the pharmaceutical industry, yet their manufacture lags significantly behind that of small molecule drugs. This paper discusses the main product-related and process-related challenges during the development and production of therapeutic proteins, with particular focus on product heterogeneity and process monitoring and analytics. Emphasis is placed on novel contributions from the field of computational research that aim to enable the application of model-based process control strategies or are working towards the development of a digital twin of bioprocesses. Lastly, we review promising developments in the paradigm shift from batch to continuous processing.

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