Synthetic Biology Research Articles

Effective removal of Pb from industrial wastewater: A new approach to remove Pb from wastewater based on engineered yeast

The use of synthetic biology to construct engineered strains has provided new perspectives for addressing Pb contamination; however, the large-scale treatment of contaminants is still limited by high operating costs and technological constraints. This study introduces a novel technique for applying engineered yeast in the removal of heavy metals, offering a solution to the cost and process scale challenges associated with utilizing engineered yeast. Hydrogen sulfide-producing engineered yeast strains were constructed based on existing strategies by knocking out the gene encoding the O-acetyl-L-homoserine mercapturic enzyme, which plays a role in sulfate assimilation. To facilitate the transition of engineered yeast from laboratory settings to industrial applications while reducing operating costs and addressing process scale-up issues, we proposes a new operational technology for engineered yeast based on their mechanistic understanding and a response surface optimization approach. The development and application of low-cost engineered media provide important guidance for utilizing engineered yeast to tackle Pb-contaminated wastewater and for the production of PbS crystalline nanomaterials. The industrial culture system was designed using economical materials and, through the response surface methodology, achieved removal rates of 99.02 ± 0.06 % and 80.95 ± 9.68 % of Pb²⁺ from Pb acid electrolyte and industrial Pb wastewater, respectively. This study presents a new technological solution for cost control and process scale-up based on the bioregulatory mechanisms of engineered yeast, laying the groundwork for their industrial application. Furthermore, it offers essential parameters and theoretical support for the industrial applications of engineered yeast in Pb wastewater treatment.

Impact of Polyploidy on Crop Improvement and Plant Breeding Strategies: A Review

Polyploidy, the condition of possessing multiple sets of chromosomes, is a widespread phenomenon in plants that has had a profound impact on crop evolution and improvement. This review explores the role of polyploidy in plant breeding strategies, emphasizing its contributions to enhancing agronomic traits, overcoming genetic barriers, and expanding genetic diversity. Polyploid crops, such as wheat, cotton, and Brassica species, exhibit superior yield, increased biomass, and enhanced stress tolerance compared to their diploid counterparts. Polyploidy can arise naturally or be induced artificially, providing breeders with opportunities to create novel crop varieties through synthetic polyploidization. The complexity of polyploid genomes poses significant challenges, including fertility issues, meiotic instability, and difficulties in genome management. Advances in genomic and biotechnological tools, such as genomic selection and CRISPR-based genome editing, are beginning to address these obstacles, allowing for precise manipulation of polyploid genomes and improved breeding outcomes. Additionally, polyploidy plays a crucial role in overcoming reproductive barriers in interspecific hybrids, facilitating the transfer of desirable traits between species that would otherwise be genetically incompatible. The potential of polyploidy for developing climate-resilient crops is particularly noteworthy, as polyploid plants often exhibit greater tolerance to drought, salinity, and extreme temperatures. This makes polyploid breeding a promising approach for addressing the challenges of climate change and ensuring food security. Future research should focus on understanding the genetic and epigenetic mechanisms underlying polyploid genome stability and trait expression, as well as integrating advanced computational tools to predict and manipulate polyploid gene function. Emerging areas such as synthetic biology and multi-omics integration will further enhance our ability to engineer polyploid crops with complex trait architectures. Polyploidy remains a powerful and versatile tool for plant breeders, offering immense potential for crop improvement, genetic innovation, and the development of resilient agricultural systems. As the understanding and technological capabilities in polyploid breeding continue to advance, polyploid crops will play a central role in sustainable agricultural development and global food production.

De Novo Production of the Bioactive Phenylpropanoid Artepillin C Using Membrane-Bound Prenyltransferase in Komagataella phaffii.

Artepillin C is a diprenylated phenylpropanoid with various pharmacological benefits for human health. Its natural occurrence is limited to a few Asteraceae plants, such as Baccharis species, necessitating a stable supply through synthetic biology. In Saccharomyces cerevisiae, the utilization of aromatic substrates within the cell was limited, resulting in very low production of artepillin C. In this study, we used AcPT1, a p-coumaric acid (p-CA)-specific diprenyltransferase, in Komagataella phaffii to produce artepillin C. Detailed studies revealed that the critical bottleneck in K. phaffii was the supply of prenyl diphosphates, not phenylpropanoid flux. By enhancing the prenyl substrate pathway through overexpression of isopentenyl diphosphate isomerase and a truncated HMG-CoA reductase, we achieved a strong increase in artepillin C production. A major part of artepillin C was accumulated in yeast cells. One of the advantages of K. phaffii is its superior growth and ability to achieve high cell density cultivation compared to that of S. cerevisiae. Therefore, fed-batch cultivation with glycerol was performed. As a result, the dry cell weight (DCW) reached 61.0 g/L, and the intracellular amount of de novo produced artepillin C reached 187.3 μg/DCW. Analysis of intermediates revealed that the supply of p-CA constituted a bottleneck in artepillin C production in the engineered strain. By enhancing the p-CA supply, the intracellular accumulation of artepillin C reached 1200 μg/DCW even in batch cultivation. Moreover, the total intra- and extracellular amounts of artepillin C reached 12.5 mg/L, marking the highest de novo synthesis amount of artepillin C reported thus far, even under batch cultivation conditions.

Oriented triplex DNA as a synthetic receptor for transmembrane signal transduction

Signal transduction across biological membranes enables cells to detect and respond to diverse chemical or physical signals, and replicating these complex biological processes through synthetic methods is of significant interest in synthetic biology. Here we present an artificial signal transduction system using oriented cholesterol-tagged triplex DNA (TD) as synthetic receptors to transmit and amplify signals across lipid bilayer membranes through H+-mediated TD conformational transitions from duplex to triplex. An auxiliary sequence, complementary to the third strand of the TD, ensures a controlled and preferred outward orientation of cholesterol-tagged TD on membranes. Upon external H+ stimuli, the conformational change triggers the translocation of the third strand from the outer to the inner membrane leaflet, resulting in effective transmembrane signal transduction. This mechanism enables fluorescence resonance energy transfer (FRET), selective photocleavage, catalytic signal amplification, and logic gate modulation within vesicles. Our findings demonstrate that these TD-based receptors mimic the functional dynamics of natural G protein-coupled receptors (GPCRs), providing a foundation for advanced applications in biosensing, cell signaling modulation, and targeted drug delivery systems.

Bioengineered therapeutic systems for improving antitumor immunity

Abstract Immunotherapy, a monumental advancement in antitumor therapy, still yields limited clinical benefits owing to its unguaranteed efficacy and safety. Therapeutic systems derived from cellular, bacterial, and viral sources possess inherent properties that are conducive to antitumor immunotherapy. However, crude biomimetic systems have restricted functionality and may produce undesired toxicity. With the advances in biotechnology, various toolkits are available to add or subtract certain properties of living organisms to create flexible therapeutic platforms. This review elaborates on the creation of bioengineered systems by gene editing, synthetic biology, and surface engineering to enhance immunotherapy. The modifying strategies on the systems are discussed, including equipment of navigation and recognition systems to improve therapeutic precision, introduction of controllable components to control the duration and intensity of treatment, addition of immunomodulatory components to amplify immune activation, and removal of toxicity factors to ensure biosafety. Finally, we summarize the advantages of bioengineered immunotherapeutic systems and possible directions for their clinical translation.

Beyond the blood: expanding CAR T cell therapy to solid tumors.

Chimeric antigen receptor (CAR) T cell therapy stands as a transformative advancement in immunotherapy, triumphing against hematological malignancies and, increasingly, autoimmune disorders. After a decade of relatively modest results for solid tumors, recent clinical trials and patient reports have also started to yield promising outcomes in glioblastoma and other challenging solid tumor entities. This Perspective seeks to explore the reasons behind these latest achievements and discusses how they can be sustained and expanded through different strategies involving CAR engineering and synthetic biology. Furthermore, we critically analyze how these breakthroughs can be leveraged to maintain momentum and broaden the therapeutic impact of CAR T cells across a variety of solid tumor landscapes.

Synthetic translational coupling element for multiplexed signal processing and cellular control.

Repurposing natural systems to develop customized functions in biological systems is one of the main thrusts of synthetic biology. Translational coupling is a common phenomenon in diverse polycistronic operons for efficient allocation of limited genetic space and cellular resources. These beneficial features of translation coupling can provide exciting opportunities for creating novel synthetic biological devices. Here, we introduce a modular synthetic translational coupling element (synTCE) and integrate this design with de novo designed riboregulators,toehold switches. A systematic exploration of sequence domain variants for synTCEs led to the identification of critical design considerations for improving the system performance. Next, this design approach was seamlessly integrated into logic computations and applied to construct multi-output transcripts with well-defined stoichiometric control. This module was further applied to signaling cascades for combined signal transduction and multi-input/multi-output synthetic devices. Further, the synTCEs can precisely manipulate the N-terminal ends of output proteins, facilitating effective protein localization and cellular population control. Therefore, the synTCEs could enhance computational capability and applicability of riboregulators for reprogramming biological systems, leading to future applications in synthetic biology, metabolic engineering and biotechnology.

Superior Photostability of the Unnatural Base 6-Amino-5-nitropyridin-2-ol: A Case Study Using Ultrafast Broadband Fluorescence, Transient Absorption, and Theoretical Computation.

6-Amino-5-nitropyridin-2-ol (Z), a nitroaromatic compound and a base for Hachimoji nucleic acids, holds significant potential in expanding the genetic alphabet, as well as in synthetic biology and biotechnology. Despite its promising applications, the spectral characterization and photoinduced properties of Z have remained largely unexplored until now. This study presents a comprehensive investigation into its excited state dynamics in various solvents, utilizing state-of-the-art ultrafast broadband time-resolved fluorescence and transient absorption spectroscopy, complemented by computational methods. The acquired results provide direct experimental evidence that, upon photoexcitation, Z emits prompt fluorescence from a nearly planar structure in its excited state, independent of solvent properties. This state deactivates nonradiatively within sub-picoseconds through internal conversion with a unitary yield, primarily mediated by the rotation of the nitro group. This unusually rapid deactivation pathway entirely excludes the involvement of long-lived nπ* states, triplet states, and photoproducts, which are commonly observed in most nitroaromatic compounds and natural DNA and RNA bases. Our findings underscore that Z, as an unnatural base, exhibits superior photostability compared to canonical natural bases. This provides valuable insights into the photodynamics of nitroaromatic compounds, which is beneficial for strategic substitution design in environmental and biological applications.

Substrate-Multiplexed Assessment of Aromatic Prenyltransferase Activity.

An increasingly effective strategy to identify synthetically useful enzymes is to sample the diversity already present in Nature. Here, we construct and assay a panel of phylogenetically diverse aromatic prenyltransferases (PTs). These enzymes catalyze a variety of C-C bond forming reactions in natural product biosynthesis and are emerging as tools for synthetic chemistry and biology. Homolog screening was further empowered through substrate-multiplexed screening, which provides direct information on enzyme specificity. We perform a head-to-head assessment of the model members of the PT family and further identify homologs with divergent sequences that rival these superb enzymes. This effort revealed the first bacterial O-Tyr PT and, together, provide valuable benchmarking for future synthetic applications of PTs.

Modifications of Prenyl Side Chains in Natural Product Biosynthesis.

In recent years, there has been a growing interest in understanding the enzymatic machinery responsible for the modifications of prenyl side chains and elucidating their roles in natural product biosynthesis. This interest stems from the pivotal role such modifications play in shaping the structural and functional diversity of natural products, as well as from their potential applications to synthetic biology and drug discovery. In addition to contributing to the diversity and complexity of natural products, unique modifications of prenyl side chains are represented by several novel biosynthetic mechanisms. Representative unique examples of epoxidation, dehydrogenation, oxidation of methyl groups to carboxyl groups, unusual C-C bond cleavage and oxidative cyclization are summarized and discussed. By revealing the intriguing chemistry and enzymology behind these transformations, this comprehensive and comparative review will guide future efforts in the discovery, characterization and application of modifications of prenyl side chains in natural product biosynthesis.

Efficient DNA-free co-targeting of nuclear genes in Chlamydomonas reinhardtii

Chlamydomonas reinhardtii, a model organism for unicellular green microalgae, is widely used in basic and applied research. Nonetheless, proceeding towards synthetic biology requires a full set of manipulation techniques for inserting, removing, or editing genes. Despite recent advancements in CRISPR/Cas9, still significant limitations in producing gene knock-outs are standing, including (i) unsatisfactory genome editing (GE) efficiency and (ii) uncontrolled DNA random insertion of antibiotic resistance markers. Thus, obtaining efficient gene targeting without using marker genes is instrumental in developing a pipeline for efficient engineering of strains for biotechnological applications. We developed an efficient DNA-free gene disruption strategy, relying on phenotypical identification of mutants, to (i) precisely determine its efficiency compared to marker-relying approaches and (ii) establish a new DNA-free editing tool. This study found that classical CRISPR Cas9-based GE for gene disruption in Chlamydomonas reinhardtii is mainly limited by DNA integration. With respect to previous results achieved on synchronized cell populations, we succeeded in increasing the GE efficiency of single gene targeting by about 200 times and up to 270 times by applying phosphate starvation. Moreover, we determined the efficiency of multiplex simultaneous gene disruption by using an additional gene target whose knock-out did not lead to a visible phenotype, achieving a co-targeting efficiency of 22%. These results expand the toolset of GE techniques and, additionally, lead the way to future strategies to generate complex genotypes or to functionally investigate gene families. Furthermore, the approach provides new perspectives on how GE can be applied to (non-) model microalgae species, targeting groups of candidate genes of high interest for basic research and biotechnological applications.

Modifications of Prenyl Side Chains in Natural Product Biosynthesis

AbstractIn recent years, there has been a growing interest in understanding the enzymatic machinery responsible for the modifications of prenyl side chains and elucidating their roles in natural product biosynthesis. This interest stems from the pivotal role such modifications play in shaping the structural and functional diversity of natural products, as well as from their potential applications to synthetic biology and drug discovery. In addition to contributing to the diversity and complexity of natural products, unique modifications of prenyl side chains are represented by several novel biosynthetic mechanisms. Representative unique examples of epoxidation, dehydrogenation, oxidation of methyl groups to carboxyl groups, unusual C−C bond cleavage and oxidative cyclization are summarized and discussed. By revealing the intriguing chemistry and enzymology behind these transformations, this comprehensive and comparative review will guide future efforts in the discovery, characterization and application of modifications of prenyl side chains in natural product biosynthesis.

Synthetic Biology in T-cell Engineering Research

Synthetic biology has emerged as a transformative discipline, enabling precise genetic and functional reprogramming of cellular systems. In T-cell engineering, it offers groundbreaking potential to revolutionize immunotherapy by endowing T cells with enhanced specificity, adaptability, and resilience against complex diseases such as cancer and autoimmune disorders. By integrating advanced genome-editing tools like CRISPR-Cas9 with modular synthetic constructs, researchers can design T cells with bespoke functionalities, such as tunable antigen recognition, controlled cytokine release, and resistance to immunosuppressive tumor microenvironments. This approach not only overcomes the limitations of conventional T-cell therapies but also facilitates the development of novel therapeutic paradigms, including "smart" cellular systems capable of sensing and responding to dynamic biological cues. Furthermore, synthetic circuits allow for the incorporation of logic-gated mechanisms to minimize off-target effects and enhance therapeutic precision. Despite these advancements, challenges remain in optimizing safety, scalability, and regulatory compliance. This research aims to explore the intersection of synthetic biology and T-cell engineering, highlighting cutting-edge methodologies, therapeutic applications, and emerging trends. By addressing current limitations and envisioning future possibilities, this work seeks to contribute to the growing body of knowledge driving synthetic biology toward clinical and industrial breakthroughs in cellular immunotherapy.

AI in Biomedical Imaging and Diagnostics

Advances in artificial intelligence (AI) and synthetic biology have profoundly influenced biomedical research, creating transformative opportunities in imaging, diagnostics, and therapeutic engineering. In biomedical imaging, AI-driven algorithms enhance precision and accuracy, enabling automated analysis of complex datasets, real-time imaging insights, and identification of disease biomarkers. Meanwhile, synthetic biology redefines cellular engineering, particularly in T-cell research, by enabling customized functionalities, such as precision-targeted antigen recognition and tunable immune responses. The integration of AI into T-cell engineering amplifies these capabilities, facilitating the design and optimization of synthetic circuits, predictive modeling of cellular behaviors, and dynamic monitoring of therapeutic outcomes. This interdisciplinary approach is revolutionizing diagnostics and immunotherapy by streamlining the identification of disease-specific markers, improving diagnostic accuracy, and enabling real-time modulation of T-cell functionality within the tumor microenvironment. By combining AI-powered insights with synthetic biology's ability to engineer living systems, this research aims to address critical challenges in disease treatment, including tumor heterogeneity and immune evasion. This work explores the synergistic application of AI and synthetic biology in biomedical imaging and T-cell engineering, highlighting state-of-the-art technologies, their therapeutic potential, and the future landscape of personalized medicine.

Deciphering substrate promiscuity and specificity of indolethylamine N-methyltransferase family enzymes from amphibian toads

N-methylation is a crucial post-modification process in natural product biosynthesis and also contributes to the metabolism of various physiological substances, such as neurotransmitter, hormone, and trace elements. In this study, we identified seven indolethylamine N-methyltransferase (INMT) family enzymes from the amphibian toad Bufo gargarizan with distinct catalytic properties. Among these enzymes, BNMT 1, BNMT 5, BNMT 6 and BNMT 7 exhibited notable promiscuity, demonstrating the ability to methylate multiple derivatives of indolethylamine, phenylethylamine, phenylethanolamine, and nicotinamide. Conversely, BNMT 3 and BNMT 4 exhibited more specific substrate preferences, targeting particular phenylethylamine, phenylethanolamine, and nicotinamide-type substrates. Additionally, one enzyme, BNMT 11, exhibiting high specificity towards phenylethanolamines. By employing molecular docking and mutating key amino acids, we provided a rational explanation for the promiscuity and specificity mechanisms exhibited by these enzymes. This research offers valuable insights into the catalytic mechanisms of INMT family enzymes in B. gargarizans, as well as other organisms. Moreover, the identification of these broadly substrate-specific enzymes holds promise for leveraging synthetic biology in the production of a wide variety of naturally occurring N-methylated compounds.

Abstraction-based segmental simulation of reaction networks using adaptive memoization

Background Stochastic models are commonly employed in the system and synthetic biology to study the effects of stochastic fluctuations emanating from reactions involving species with low copy-numbers. Many important models feature complex dynamics, involving a state-space explosion, stiffness, and multimodality, that complicate the quantitative analysis needed to understand their stochastic behavior. Direct numerical analysis of such models is typically not feasible and generating many simulation runs that adequately approximate the model’s dynamics may take a prohibitively long time.Results We propose a new memoization technique that leverages a population-based abstraction and combines previously generated parts of simulations, called segments, to generate new simulations more efficiently while preserving the original system’s dynamics and its diversity. Our algorithm adapts online to identify the most important abstract states and thus utilizes the available memory efficiently.Conclusion We demonstrate that in combination with a novel fully automatic and adaptive hybrid simulation scheme, we can speed up the generation of trajectories significantly and correctly predict the transient behavior of complex stochastic systems.

Modulating Microbial Materials - Engineering Bacterial Cellulose with Synthetic Biology.

The fusion of synthetic biology and materials science offers exciting opportunities to produce sustainable materials that can perform programmed biological functions such as sensing and responding or enhance material properties through biological means. Bacterial cellulose (BC) is a unique material for this challenge due to its high-performance material properties and ease of production from culturable microbes. Research in the past decade has focused on expanding the benefits and applications of BC through many approaches. Here, we explore how the current landscape of BC-based biomaterials is being shaped by progress in synthetic biology. As well as discussing how it can aid production of more BC and BC with tailored material properties, we place special emphasis on the potential of using BC for engineered living materials (ELMs); materials of a biological nature designed to carry out specific tasks. We also explore the role of 3D bioprinting being used for BC-based ELMs and highlight specific opportunities that this can bring. As synthetic biology continues to advance, it will drive further innovation in BC-based materials and ELMs, enabling many new applications that can help address problems in the modern world, in both biomedicine and many other application fields.

Development of a specific biosensor for sesquiterpene based on SELEX and directed evolution platforms

Biosensors are essential in synthetic biology, particularly for detecting compounds without distinct visual markers. In this study, a riboswitch biosensor specifically responsive to a sesquiterpene — amorpha-4,11-diene was developed for the first time. Through SELEX-SMB, a high-affinity aptamer library comprising 81,520 sequences was generated. Subsequent screening via the TBG-directed evolution platform identified riboswitch5, which downregulates downstream gene expression in response to amorpha-4,11-diene within a concentration range of 10–100 mg/L, achieving a log₂Foldchange of −0.51 at 100 mg/L. Structural predictions, combined with molecular docking and molecular dynamics simulations, revealed that ligand binding induced conformational changes that stabilize the riboswitch and enhance its repressive effect. This biosensor represents a powerful tool for the detection and regulation of small molecules, with broad applications in strain engineering, pathway regulation, and metabolic optimization in synthetic biology.

Reducing potential dual-use risks in synthetic biology laboratory research: a dynamic model of analysis

Synthetic biology (SynBio) is an interdisciplinary field that includes biology, genomics, engineering, and informatics. The introduction of machine learning technology reduces the requirement for human participation in SynBio research, which may lead to increased biosafety and biosecurity concerns within the complex human–technology-management system in SynBio laboratories compared with traditional biology. The rapid development of SynBio and machine learning technology has added to the systemic complexity of management and technology subsystems, the risks of which remain unclear. To fully understand and address these risks, there is an urgent need to develop a quantitative model. In this study, three-round Delphi interviews were conducted with SynBio experts who have published articles in top journals (e.g., Nature, Science, Cell, etc.) and applied the system dynamics method to build a quantitative analysis model. The model explores the potential risk factors and their complex relationships in the SynBio research. We analyzed three subsystems (biotechnology, information technology, and management), identifying seven key risk factors. Among these, the Average Experience of Newcomers and Tolerable Error Frequency play an essential role in laboratory management. Notably, the biosafety atmosphere has the greatest impact on reducing error rate and increasing safety awareness, as well as on the infrastructure safety of SynBio laboratory. In addition, we also consider the impact of the progressiveness of machine learning in the SynBio research and find that using more advanced machine learning technology leads to lower instrument safety. This study provides a framework for quantifying potential risk factors in SynBio research and lays a theoretical foundation for future laboratory safety management and the development of a global code of conduct for interdisciplinary scientists.

Advancements in Colloidosome Fabrication, Functionalization, and Applications: A Comprehensive Review

Abstract: Among the recent developments in the field of microcapsules, colloidosomes microcapsules made of colloidal particles have elicited much interest in promising perspectives in microencapsulation technologies. First reported by Dinsmore et al., these structures hold a lot of potential in different areas of application, which include gene delivery, targeting the brain, and tumor treatment. However, the challenges associated with their permeability and stability limit the use of these coatings. Nevertheless, these problems define colloidosomes as possessing some features of protocells that include metabolic activities and response to stimuli, which make them significant in synthetic biology and biomimetic systems. In drug delivery, colloidosomes have the potential of providing solubility, stability, and a controlled rate of drug release, which will have an impact on the therapeutic use of drugs. Fabrication of colloidosomes is done in many ways, and the common ones include the mulsion strategy, layer adsorption, and microfluidics strategy. Some of these fabrication techniques have been investigated in the recent past to determine how their application can lead to enhanced colloidosome characteristics. Some of these methods include spray drying, sol-gel processes, and electrostatic assembly. For example, the study by Payizila Zulipiker et al. showed that different levels of permeability are controllable through the use of spray drying technology. Another study by Jia Jia et al. presented the preparation of pH-responsive colloidosomes and the high efficiency of encapsulation. Consequently, improvements in the manufacturing process and new self-assembly technologies that include microfluidics systems have also improved the creation of colloidosomes. New advancements in the field of colloidosome synthesis, as well as their surface modification, will open up numerous application prospects in the fields of pharmaceutics, regenerative medicine, and bioinspired materials science. New trends range from 3D printing to stimuli-responsive materials and design, as well as hybrid systems that are application- specific. Despite the difficulties that remain to be faced, colloidosomes represent an extraordinary opportunity for the growth of biotechnology and materials science to open the path to new generations of drugs and bioinspired applications.