Editorial pubs.acs.org/synthbio
Synthetic Biology in Asia: New Kids on the Block
F
Furthermore, Px can be modified with various functional groups such as fluorescent moieties and biotin to provide a sitespecific DNA labeling for detection or immobilization, thus providing an expanded genetic alphabet system. A control of protein expression can also be harnessed to optimize enzyme activity especially if complex folding and formation are required. Multimeric proteins for enzymatic activity may require a correct stoichiometry for complex formation and functionality during in vitro reconstitution after purification. In Qin et al. (DOI: 10.1021/acssynbio.5b00291), the expression of multicomponent protein complex that leads to efficient assembly, hence enzymatic activity was demonstrated by their expression platform, EcoExpress. The design aids optimal gene assembly by having individual monomer with user-defined epitope-tag into individual expression vectors, which is followed by one-pot assembly of a single vector to express the entire protein complex for copurification. EcoExpress is a toolbox that provides a platform to investigate protein complexes in various different microbial systems. Even with controlled expression to optimize enzyme activity, screening enzyme activity in high throughput manner for rapid selection is always highly sought after to accommodate the rapid development of the field. In Matsumoto et al. (DOI: 10.1021/acssynbio.6b00194), metabolic enzyme ligation was harnessed for the redirection of metabolic flux through metabolic channeling, demonstrating that sortase A-mediated enzyme ligation improves acetate-producing flux and acetate production in E. coli. In Kim et al. (DOI: 10.1021/ acssynbio.5b00287), an advanced version of genetic enzyme screening system (GESS) using a phenotypic reporter and cell−cell communication was shown to improve the efficiency of the screening technique, which can aid in screening of novel microbes with target enzymes of interest. With ever expanding genetic tools, the biological production of value-added chemicals is becoming more and more economically feasible. In 2004, the U.S. Department of Energy (DOE) has published a list of sugar-based chemical building blocks, which can be subsequently converted to a number of value-added chemicals.3 Over the past decade, numerous metabolic engineering studies across the world including Asia have been conducted to produce these building blocks using microbial hosts. Furthermore, driven by large and widespread feedstocks and government incentives, Southeast Asia aims to become a hub for biobased materials and chemicals. In lieu of this movement, this special issue contains four pieces of work that produce the top listed building blocks respectively (succinate, 3-hydroxypropionic acid (3-HP) and secondary chemical, poly-3-hydroxybutyrate). In Li et al. (DOI: 10.1021/acssynbio.6b00052), E. coli was reprogrammed to overproduce succinate from acetate as the sole carbon source through an extensive metabolic engineering. Although high concentration of acetate had negative effect on
rom the advent of recombinant DNA technology to the creation of programmable genetic circuits and artificial genomes, the unraveling of the secrets of biology and technological advancements made in the field of synthetic biology has been astonishing. Given the relatively short history of the field, the influence of synthetic biology on various research disciplines is unrivaled. While the foundational movements and efforts have been initiated and centered mostly in the United States and the Europe, the following waves of contributions and drive toward expanding the field of synthetic biology have taken worldwide efforts. In particular, with a growing recognition of the potential of synthetic biology to contribute to the biobased economy, a number of countries in Asia have established national initiatives on synthetic biology, promoting technological innovations and economic growths through synthetic biology. For instance, China has greatly invested in synthetic biology driven by a coordinated national strategy under state sponsored programs.1 This effort was reflected in the growing number of journal publications, where 10% of the papers published on synthetic biology globally in 2013 were reportedly contributed by China.2 Other countries in Asia such as Japan and Korea have rapidly developed vibrant synthetic biology communities, underpinned by national institutes on synthetic biology, and generated research outputs that could significantly contribute to the advancements in the field. Singapore has also made a noteworthy progress in building core research communities and capabilities in synthetic biology, attested by the establishment of a biofoundry and a national consortium for synthetic biology. Certainly, synthetic biology in Asia is poised to play a greater role at the global stage. This special issue of ACS Synthetic Biology celebrates these accomplishments by highlighting recent advancements made in synthetic biology in Asia that range from the development of foundational genetic tools to the biosynthesis of industrially relevant chemicals. Foundational genetic tools that enable a defined control of gene or protein expression are important key elements in synthetic biology and have been a focal point of synthetic biology efforts in Asia as well. In Saeki et al. (DOI: 10.1021/ acssynbio.5b00230), an inducible transcriptional factor BetI was systematically engineered to generate a library of genetic switches that show various inducible and repressible promoter systems in Escherichia coli. This new set of Bet-ON and BetOFF systems was shown to be compatible with other inducible expression systems such as tet to work as two-input Boolean logic gates. In expanding the genetic codes available to impose a defined control over the functions of genetic circuits, synthetic xenobiology is very much on the horizon. In Okamoto et al. (DOI: 10.1021/acssynbio.5b00253), a hydrophobic base pair between 7-(2-thienyl)-iidazo [4,5-b] pyridine (Ds) and 2-nitro4-propynylpyrrole (Px) was developed based on shapecomplementarity between pairing bases. The unnatural Ds− Px pair retained a high fidelity in polymerase chain reaction (PCR) with 3′-exonuclease-proficient DNA polymerase. © 2016 American Chemical Society
Received: November 2, 2016 Published: November 18, 2016 1182
DOI: 10.1021/acssynbio.6b00327 ACS Synth. Biol. 2016, 5, 1182−1183
ACS Synthetic Biology
Editorial
control functionalities. In Wang et al. (DOI: 10.1021/ acssynbio.5b00180), synthetic microRNA cluster was developed for genome editing driven by multiplex RNA interference. By identifying miRNA precursor architecture for precise miRNA maturation and integration efficiency, a highly efficient multiplex gene knockdown was achieved. The method provides a powerful modular genetic tool that enables complex functional genomic studies to identify multidimensional gene interactions at the post-transcriptional level. Further, in Lee et al. (DOI: 10.1021/acssynbio.5b00249), a rational engineering strategy was devised to identify specific functional genes (Igf bp4 and AqpI) that are altered in suspension adaptation by high-throughput RNA sequencing, which was subsequently followed by validating the effect through gene deletion mediated by DNA-free RNA-guided Cas9 (CRISPR associated protein 9). This integrative tool can enable the elucidation of molecular mechanisms underlying the effect of disrupting target genes on cellular adaptation, which can be used for rational genome engineering of CHO cells for desirable phenotypes. A key inspiration of the creation of a special issue on synthetic biology in Asia was to commemorate the achievements and advancements made so far by the Asian synthetic biology communities. The work featured in this issue certainly exemplifies the scientific contributions that Asia has made to the field of synthetic biology. Given increasing public and private support for synthetic biology and rapidly growing synthetic biology communities in Asia, many now foresee that Asia will take on a greater role at the global stage in advancing synthetic biology for the benefit of humanity.
succinate conversion, the highest succinate concentration (61.71 mM) was achieved in a resting cell conversion system in minimal medium. Since acetate is a nontraditional carbon source that is readily available in industrial wastewater and lignocellulosic hydrolysate and yet less costly than glucose, this work demonstrates a potentially economical method for production of highly valued succinate. 3-Hydroxypropionic acid (HP), which is ranked third in the list by DOE, was produced by two independent groups based on two different approaches. In Lim et al. (DOI: 10.1021/ acssynbio.5b00303), rebalancing the biochemical pathway via 5′UTR modification and fine-tuning the expression ratio of two enzymes required to optimally facilitate a 2-step reaction conversion of glycerol to 3-HP, significantly improved the productivity in E. coli. Upon optimization of the fed-batch cultivation, production of 40.51g/L 3-HP was achieved. Impressively, the work showed an unprecedented production yield of 0.97g/g glycerol. In Song et al. (DOI: 10.1021/ acssynbio.6b00007), metabolic pathways were rerouted to increase the level of β-alanine and malonic semialdehyde (MSA) from glucose. MSA was then utilized to produce 3-HP and malonic acid (MA) via redox reaction mediated by semialdehyde dehydrogenase (yneI) and MSA reductase (ydfg), respectively. This approach has shown 31.3 g/L 3-HP and 3.6 g/L malonic acid production from glucose by fed-batch cultivation. Poly-3-hydroxybutyrate (PHB) Bioplastic is the most common type of polyhydroxyalkanoates produced by bacterial fermentation processes in large quantity. However, relatively high production cost and complex purification processes are the limitations faced for commercialization. In addressing this issue, Chen and his colleagues (DOI: 10.1021/acssynbio.6b00083) took the approach of harnessing in-silico RBS library design and high-throughput screening based on visual on-plate selection to improve microbial production of poly-3-hydroxybutyrate (PHB). As PHB exists as solid granules inside the cells, the group further improved their design by introducing an extracellular production of soluble and high value-added chemical, such as 5-aminolevulinic acid (ALA) for coproduction (DOI: 10.1021/acssynbio.6b00105). This not only lowers the production cost, but also supports multiple productions from one fermentation process, which addresses the two limitations faced in commercialization. This special issue also highlights programmable genetic sensors engineered for the detection of an environment contaminant and a human pathogen. In Chong and Ching (DOI: 10.1021/acssynbio.6b00061), a colorimetric whole-cell biosensor was developed for the organophosphorus compound parathion, a potent pesticide and pollutant, by sensing its hydrolysis product, 4-nitrophenol. This work also demonstrates that a simple visual detection method can lead to a detection limit as low as 10 μM. Furthermore, in Holowko et al. (DOI: 10.1021/acssynbio.6b00079), a whole-cell biosensor was engineered for the detection of the water-borne human pathogen Vibrio cholerae by harnessing the quorum sensing mechanism of the pathogen. In addition, in silico modeling and simulation enabled tightly and optimally regulated expression that results in a highly sensitive V. cholera detection. These two studies laid the solid groundwork for the development of versatile, sustainable, low-cost and operationally stable wholecell biosensors for various real-life applications. Lastly, this special issue features two pieces of seminal work on mammalian synthetic biology that demonstrate genetic
Matthew Wook Chang*
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Department of Biochemistry, Yong Loo Lin School of Medicine, and NUS Synthetic Biology for Clinical and Technological Innovations (SynCTI), National University of Singapore, 119077 Singapore
AUTHOR INFORMATION
Corresponding Author
*E-mail:
[email protected]. Notes
Views expressed in this editorial are those of the author and not necessarily the views of the ACS.
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REFERENCES
(1) Pei, L., Schmidt, M., and Wei, W. (2011) Synthetic biology: An emerging research field in China. Biotechnol. Adv. 29 (6−3), 804−814. (2) Committee on Science, Technology, and Law; Policy and Global Affairs; Board on Life Sciences; Division on Earth and Life Sciences; National Academy of Engineering; National Research Council. In Positioning Synthetic Biology to Meet the Challenges of the 21st Century: Summary Report of a Six Academies Symposium Series, National Academies Press (US), Washington, D.C., Aug 5, 2013, p 3, Strategies for Advancing Synthetic Biolgy. (3) Werpy, T. A., Holladay, J. E., and White, J. F. Top Value Added Chemicals From Biomass: I. Results of Screening for Potential Candidates from Sugars and Synthesis Gas, Pacific Northwest National Laboratory (PNNL), Richland, WA, 2004.
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DOI: 10.1021/acssynbio.6b00327 ACS Synth. Biol. 2016, 5, 1182−1183
