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Table of Content

    31 August 2025, Volume 6 Issue 4
    Contents in Chinese and English
    2025, 6(4):  0. 
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    Perspective
    PE and PX values in biomanufacturing: definitions and applications
    ZHANG Yi-Heng P. Job, CHEN Xuemei, HAN Pingping
    2025, 6(4):  715-727.  doi:10.12211/2096-8280.2025-020
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    Biomanufacturing is one of the representatives of strategic emerging industries and represents a new quality productive force for the upcoming bioeconomy. The authors used to propose the philosophical guiding significance of “Tao-Fa-Shu-Qi” for industrial biomanufacturing. To further elaborate on the principle of “Shu is techniques” in biomanufacturing, for the first time this opinion paper introduces the new concepts of “Product-to-Enzyme Ratio” (abbreviated as PE value) that is the weight ratio of the product to the non-cell biocatalysts (i.e., enzyme(s)), such as enzyme molecules and multiple-enzyme molecular machines, and of “Product-to-X(Cell) Ratio” (abbreviated as PX value) that is the weight ratio of the product to cell mass. Both values feature simple, clear and quantitative properties. The PE value can be applied to enzyme-based biocatalysis or in vitro biotransformation; PX value can be applied to cell-based fermentation. Theoretical PE values can be calculated from total-turn-over number (TTN) of enzyme. First, the author defines and calculates the PE value. Second, authors summarize and classify industrial biomanufacturing cases and literature data catalyzed by enzymes, multiple-enzyme molecular machines, and cells. It was found that enzymatic starch-hydrolyzing enzymes has a PE value 50-100 times of that of enzymatic cellulose hydrolysis, resulting in an extremely high cellulase dosage. This ultra-high enzyme cost is the biggest obstacle to the industrialization of the second-generation biorefineries that are based on the biological conversion and utilization of lignocellulosic biomass. Lastly, the PE/PX values can be used to quickly estimate and compare the costs of biocatalysts, such as enzyme molecules, multiple-enzyme molecular machines, and microbial cells in the biomanufacturing process, guiding the development path for further decreasing the cost of biocatalysts. Increasing PE values by enzyme (co-)immobilization could greatly decrease biocatalyst costs, surpassing cell-based fermentation. Cellulase could be the largest industrial enzyme with a potential market size of 500 billion RMB by considering the huge supplies of non-food lignocellulosic biomass and potential markets of renewable biocommodities and artificial food/feed. The calculation and analysis of the PE/PX values would provide a new perspective for the future development of the strategic emerging industry of biomanufacturing, deepen the understanding of the cost of biocatalysts in biomanufacturing, promote the high-quality development of the bioeconomy, predict the future cost trend of biological products, and evaluate the industrialization potential of emerging biotechnologies. {L-End}

    Invited Review
    Halogenases in biocatalysis: advances in mechanism elucidation, directed evolution, and green manufacturing
    WANG Mingpeng, CHEN Lei, ZHAO Yiran, ZHANG Yimin, ZHENG Qifan, LIU Xinyang, WANG Yixue, WANG Qinhong
    2025, 6(4):  728-763.  doi:10.12211/2096-8280.2024-091
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    Organic halides, which serve as critical structural motifs in pharmaceuticals, agrochemicals, and advanced materials, are typically synthesized using energy-intensive processes that involve toxic reagents and generate hazardous waste. In contrast, halogenases-nature’s biocatalytic tools-catalyze regio- and stereoselective halogenation under environmentally benign conditions, offering a paradigm shift towards sustainable chemistry. This review systematically consolidates recent breakthroughs in halogenase research, emphasizing mechanistic insights, engineering innovations, and scalable industrial applications. Halogenases are mechanistically classified into three major families: flavin-dependent enzymes that mediate electrophilic halogenation throughtransient hypohalous acid intermediates; non-heme iron/α-ketoglutarate-dependent oxygenases that drive radical-based halogenation pathways; and S-adenosylmethionine (SAM)-dependent enzymes that facilitate rare nucleophilic halogenation. Cutting-edge structural biology techniques, enhanced by computational simulations, have elucidated dynamic substrate-enzyme interactions and transient catalytic states, facilitating the rational design of halogenases with tailored reactivity. , The integration of bioinformatics tools with high-throughput screening platforms has concurrently accelerated the discovery of novel halogenases from underexplored microbial niches, revealing unprecedented catalytic diversity. To bridge natural enzymatic capabilities with industrial demands, interdisciplinary strategies are being deployed: Directed evolution optimizes activity and stability under non-native conditions; computational protein design rebuilds substrate-binding pockets for non-canonical substrates; and synthetic biology frameworks reconstruct halogenation pathways in engineered microbial hosts. These efforts collectively expand the biocatalytic toolbox, enabling precise halogenation of complex scaffolds, including aromatic systems, aliphatic chains, and heterocycles. In industrial contexts, enzymatic halogenation is gaining traction for synthesizing high-value compounds, ranging from antibiotic derivatives and antitumor agents to crop protection molecules, while circumventing the traditional reliance on heavy metal catalysts, extreme temperatures, and halogenated solvents. Emerging applications further extend to the functionalization of biomaterials and fine chemicals, underscoring the versatility of halogenases. Future advancements will likely harness machine learning algorithms to decode sequence-activity landscapes and predict multi-enzyme cascades for tandem halogenation-functional group interconversions. Such developments are in line with global sustainability agendas, positioning halogenases as key biocatalysts in the transition towards circular chemical economies. This review highlights the convergence of enzymology, systems biology, and green chemistry in unlocking the full potential of halogenases, paving the way for next-generation biomanufacturing. {L-End}

    Recent advancements in non-biological component-augmented synthetic bio-hybrid systems
    HUANG Yuqing, WU Han, LI Xiaobin, LIU Junyu, MA Shaohua, GE Jun, XING Xinhui, ZHANG Canyang
    2025, 6(4):  764-788.  doi:10.12211/2096-8280.2025-048
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    The integration of non-biological components into living systems represents a pivotal advance in synthetic biology. This approach facilitates the creation of synthetic bio-hybrid systems effectively overcoming inherent limitations of natural biological systems. By leveraging the synergistic enhancement and superior functionalities derived from both biological and non-biological constituents, these hybrid systems exhibit immense potential across diverse applications, including bio-manufacturing, precise diagnostics, and biomedicine, establishing them as a cutting-edge frontier. However, despite the vast functional diversity of non-biological components, current bio-hybrid systems often present functional singularity, and their underlying synergistic mechanisms remain insufficiently elucidated. These limitations hinder their broader adoption and sophisticated applications. To address these challenges, this review systematically summarizes recent advancements in non-biological component-enhanced synthetic bio-hybrid systems. We categorize these systems based on the nature of the non-biological components (e.g., nanomaterials, polymers, semiconductors) and their integration strategies with diverse biological entities (e.g., enzymes, nucleic acids, cells). Through in-depth analysis of representative studies, we elucidate construction methodologies, functional realization pathways, and performance characteristics across various hybrid configurations. A central focus is to critically identify existing limitations, particularly concerning functional modularity, fine-tuned control, and the comprehensive elucidation of complex underlying mechanisms. We also explore strategies to overcome these challenges, emphasizing rational design and advanced characterization. Looking ahead, we present a forward-looking perspective on the future trajectory of this burgeoning field. Key areas for advancement include multi-platform integration, combining various non-biological components with multiple biological parts for highly sophisticated systems. Furthermore, we highlight the importance of advanced engineering design and high-throughput screening to accelerate discovery and optimization. The refinement of precise spatiotemporal regulation is crucial for controlling complex assemblies. Moreover, the integration of artificial intelligence and machine learning for rational design promises to revolutionize development. This review aims to serve as a valuable resource, providing critical insights and inspiring further research into the design, construction, and application of non-biological component-enhanced synthetic bio-hybrid systems, therefore paving the way for groundbreaking innovations in healthcare and biotechnology. {L-End}

    Application of plasma microbial breeding technology in biofabrication
    ZHONG Naicai, CHEN Yuan, PAN Wenfeng, SU Xiaofeng, LIAO Jingwen, ZHAI Yinglei, ZHONG Jinyi
    2025, 6(4):  789-805.  doi:10.12211/2096-8280.2025-005
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    Sustainable and green biomanufacturing has emerged as a strategic priority for nations globally; however, microbial performance remains one of the key constraints hindering the industrialization of biomanufacturing. Traditional breeding methods often face challenges such as long breeding cycles, low efficiency, and high costs, making it difficult to meet the demands for highly efficient and stable microbial strains in industrial-scale production. Low-temperature plasma technology, as an efficient and environmentally friendly method for microbial breeding, can stimulate mutation hotspots, enhance mutation efficiency, and expand mutation ranges, effectively improving the performance of target strains and product yields. By combining plasma mutagenesis’s advanced capabilities with other techniques, significant improvements in the performance and productivity of microbial strains can be achieved, thus driving the commercialization of sustainable bioprocesses. This review outlines the theoretical basis of plasma mutagenesis technology, the technical characteristics of three plasma sources (ARTP, DBD, and CD), the mechanisms of plasma mutagenesis, and the progress of combining this technology with high-throughput screening, classic mutagenesis, and rational breeding methods. Furthermore, it summarizes typical plasma breeding cases in biomanufacturing fields such as bio-enzyme, organic acids, bioenergy, and biomaterials. These insights offer important references for research and industrialization in related fields. By combining plasma mutagenesis’s advanced capabilities with other techniques, significant improvements in the performance and productivity of microbial strains can be achieved, thus driving the commercialization of sustainable bioprocesses. The cases discussed in this review provide a practical understanding of how plasma mutagenesis can be applied to optimize microbial strains for industrial-scale production of valuable bioproducts, providing a reference for research and industrialization in related fields. In the future, it is essential to develop novel plasma generators based on air source, which, while being miniaturized, achieve low cost, low energy consumption, and minimal temperature rise. Integrating them with high-throughput screening and AI technologies will enable precise microbial mutagenesis and efficient strain breeding, thereby overcoming technical bottlenecks and ultimately advancing the biomanufacturing industry. {L-End}

    Synthetic biology and applications of high-adhesion protein materials
    LI Quanfei, CHEN Qian, LIU Hao, HE Kundong, PAN Liang, LEI Peng, GU Yi’an, SUN Liang, LI Sha, QIU Yibin, WANG Rui, XU Hong
    2025, 6(4):  806-828.  doi:10.12211/2096-8280.2025-043
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    Due to their exceptional bioadhesive properties and potential biocompatibility, high-viscosity protein materials exhibit significant application prospects in the fields of biomedical materials and adhesives. However, traditionally sourced high-viscosity protein materials encounter numerous challenges, including low yields, structural complexity, and difficulties in scaling up production. Synthetic biology, as an emerging interdisciplinary field, offers innovative strategies to address these bottlenecks. This review systematically summarizes recent advances in the biosynthesis, modification, and applications of high-viscosity protein materials, focusing on the advantages of synthetic biology in addressing issues related to the yield, controllability, and functional diversity of these materials. The precise design and efficient expression of adhesive proteins, such as mussel adhesive proteins, barnacle cement proteins, and scallop foot proteins, achieved through genetic engineering, are comprehensively reviewed, demonstrating the overcoming of limitations in the production and controllability of high-viscosity protein materials. Furthermore, the unique advantages of these protein materials in bioadhesives and functional medical coatings, such as the wet adhesion of mussel proteins, the strong adhesion of barnacle cement proteins, and the tunable properties of elastin-like proteins, are summarized. By employing synthetic biology approaches, limitations in the yield, performance, and functionality of high-viscosity protein materials can be overcome, thereby accelerating their application in areas such as tissue engineering and surface modification. Finally, the latest advancements and innovations in the field of synthetic biology for high-viscosity protein materials are summarized, and future development directions are envisioned, offering new ideas and strategies for the development of high-performance, multifunctional high-viscosity protein materials.

    Construction and advances in the applications of transcription factor-based biosensors
    WANG Hong, LU Kongyong, ZHENG Yangyang, CHEN Tao, WANG Zhiwen
    2025, 6(4):  829-845.  doi:10.12211/2096-8280.2025-030
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    Microbial cell factories are pivotal in green biomanufacturing, with applications spanning various sectors, including food production, chemical engineering, pharmaceuticals, and energy. However, traditional metabolic engineering strategies, which reply on static regulation and are hindered by the inherent latency in real-time metabolic flux monitoring, face significant limitations in constructing microbial systems that efficiently synthesize target products. These constraints severely hinder the high-yield biosynthesis of bio-based compounds. Transcription factor-based biosensors (TFBs), which are cornerstone tools in synthetic biology and metabolic engineering, offer innovative solutions by dynamically linking real-time perception of metabolite concentration signals or environmental cues with autonomous regulation of target gene expression. This integration allows for intelligent optimization and efficient construction of microbial production systems. This review systematically examines the molecular architecture, functional classification, and signal transduction mechanisms of TFBs, focusing on the rational design of ligand-recognition modules and the reconfiguration of signal-output components. Key strategies for constructing TFBs are summarized, including directed evolution and rational redesign of transcription factor ligand-binding domains (LBD), modular engineering of responsive promoters, and optimization of ribosome binding sites (RBS) for reporter genes. The review also highlights cutting-edge applications of TFBs in microbial cell factories, such as high-throughput screening platforms, identification of metabolic engineering targets, and dynamic regulation of metabolic pathways. Despite their transformative potential, several challenges remain, including the scarcity of metabolite-responsive elements, narrow ligand detection ranges, insufficient substrate recognition specificity, time-consuming transcription-dependent processes, and poor robustness of sensor components under industrial conditions. To address these bottlenecks, future research must prioritize the integration of synthetic biology with artificial intelligence (AI)-driven big data modeling. Such interdisciplinary efforts will accelerate the development of customizable, standardized plug-and-play modular components to overcome limitations like the shortage of responsive elements. Concurrently, the establishment of scalable validation platforms across “lab-scale, pilot-scale, and industrial production” stages is essential to validate system scalability, laying the foundation for next-generation TFBs capable of supporting large-scale industrial biomanufacturing. These advancements are set to enhance the efficiency and intelligence of microbial cell factories while expanding their applications in critical areas such as food safety testing, environmental monitoring, and medical diagnostics and therapeutics. By offering critical insights into the design and application of TFBs, this review aims to drive the evolution of microbial cell factories into multifunctional, smart bioproduction systems that integrate precision, adaptability, and industrial robustness, ultimately fostering sustainable innovation in the bioeconomy. {L-End}

    Research progress in plant-derived vaccines
    SONG Xinyu, PAN Weisong, WU Tairu, PAN Jiahao, WU Chuan, LI Waichin
    2025, 6(4):  846-872.  doi:10.12211/2096-8280.2025-029
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    Plant-derived vaccines represent an innovative vaccine production technology that employs plants as bioreactors to express specific antigenic proteins within the plant system. This technology has demonstrated tremendous potential and application prospects in the field of vaccines in recent years. Compared to traditional vaccine production methods, plant-derived vaccines offer distinct advantages in cost control, scalability, and safety. Firstly, the production cost of plant-derived vaccines is relatively low. This is due to the short growth cycle of plants, their strong reproductive capacity, and the lack of need for complex bioreactors or expensive culture media. This makes the large-scale production of vaccines more economical and efficient. Secondly, plant-derived vaccines are easy to scale up. Due to the renewable nature and rapid growth characteristics of plants, they can quickly respond to large-scale vaccine demands, which is particularly important in dealing with public health emergencies. In addition, there is no risk of contamination in the production process of plant-derived vaccines that is typically associated with traditional vaccine production. Plant cells possess inherent biosafety, which can effectively avoid contamination from animal-derived pathogens and endotoxins, thus ensuring the safety of the vaccine. Plants can perform post-translational modifications on foreign proteins, a characteristic that is conducive to the formation of virus-like particles (VLPs). VLPs are non-infectious particles that structurally resemble viruses; they can mimic the immunogenicity of viruses, stimulating the body to produce an immune response. However, they lack the ability to replicate, which makes them safer. This article first introduces the basic concept of plant-derived vaccines by using plants as vectors to express antigenic proteins. Then, the article emphasizes the important role of plant-derived vaccines in the field of global public health and epidemic prevention, especially in providing rapid, economical, and safe vaccines. The article then details the development history of plant-derived vaccines, from early exploration to modern commercial applications. At the same time, the article provides a comprehensive description of the different classifications, expression platforms, and expression systems of plant-derived vaccines, covering various technological pathways from genetically engineered plants to plant viral expression vectors. The analysis focused on how vaccine optimization and application enhance the expression and immunogenicity of antigenic proteins through gene editing and protein engineering, as well as how to improve the efficacy and stability of vaccines by optimizing their formulation and adjuvants. Furthermore, current cases of developed plant-derived vaccines were analyzed, especially their application advantages in addressing human and animal diseases. These cases demonstrate the potential of plant-derived vaccines in rapidly responding to epidemics, reducing costs, and improving accessibility. Finally, the article discusses and summarizes the development progress of plant-derived vaccines domestically and internationally, providing references and insights for the research and application of plant-derived vaccines in our country. Through these analyses, the article aims to promote the development of plant-derived vaccine technology and contribute to global public health security. {L-End}

    Trends and challenges in microbial synthesis of higher alcohols
    FANG Xinyi, SUN Lichao, HUO Yixin, WANG Ying, YUE Haitao
    2025, 6(4):  873-898.  doi:10.12211/2096-8280.2025-006
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    Higher alcohols refer to alcohols containing three or more carbon atoms and represent an important class of chemicals widely used in various industries, such as fuels, solvents, coatings, and specialty chemicals. Traditionally, the production of these higher alcohols has depended heavily on petrochemical processes, which not only rely on non-renewable resources but also contribute significantly to environmental pollution. The finite nature of fossil fuels and the associated environmental concerns have prompted researchers to explore alternative, sustainable production methods. Consequently, the development of sustainable bio-based higher alcohol production technologies has emerged as a critical research focus. Recent advances in metabolic engineering and synthetic biology have paved the way for developing engineered microbial strains capable of producing higher alcohols through biological fermentation. By optimizing metabolic pathways, these engineered strains can channel more carbon flux toward the desired alcohol products. In addition, enhancing the tolerance of these strains to high concentrations of the produced alcohols and improving their overall biosynthetic capabilities are key strategies that have been successfully implemented. Such innovations have enabled the production of higher alcohols from renewable feedstocks in a more environmentally friendly and cost-effective manner. Renewable raw materials, such as lignocellulose, waste proteins, waste lipids, and carbon dioxide, provide diverse possibilities for the environmentally friendly and sustainable synthesis of higher alcohols. Lignocellulosic biomass, for instance, is abundant and renewable, making it an attractive alternative to conventional sugars. Waste proteins and lipids, often derived from industrial by-products, provide additional inexpensive substrates that not only help in waste valorization but also reduce the overall production cost. Carbon dioxide, as an abundant greenhouse gas, can be captured and converted into valuable higher alcohols, contributing to carbon sequestration and climate change mitigation. Despite these promising prospects, several challenges remain to be addressed. Issues such as low substrate conversion efficiency, the formation of inhibitory byproducts during fermentation, and high costs associated with downstream separation and purification continue to hinder commercial viability. This review provides a comprehensive overview of the current market size, major applications, and economic value of higher alcohols, with particular emphasis on the market performance of isobutanol, 1,3-butanediol, and 2,3-butanediol. In addition, the review explores the biosynthetic pathways utilized for higher alcohol production, including the acetyl-CoA-dependent pathway, the branched-chain amino acid synthesis pathway, and the fatty acid chain elongation pathway. It also summarizes key metabolic engineering strategies, such as cofactor balancing, competitive pathway elimination, enzyme optimization, and high-yield strain selection. Moreover, the utilization of extremophiles as chassis cells, in combination with next-generation industrial biotechnology (NGIB), represents a promising new direction for sustainable production. Looking ahead, the integration of biosensors, advanced gene editing technologies, and computer-aided metabolic engineering is expected to further optimize microbial cell factory design, thereby enhancing the industrial production efficiency of higher alcohols and promoting the development of renewable energy and green chemical industries. {L-End}

    Advancements in the study of probiotics for adjunctive prevention and treatment of malignancies
    ZHU Xinyue, CHEN Tiantian, SHAO Hengxuan, TANG Manyu, HUA Wei, CHENG Yanling
    2025, 6(4):  899-919.  doi:10.12211/2096-8280.2025-004
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    Cancer continues to pose a significant global public health challenge, as its incidence and mortality rates persistently rise. Conventional cancer treatments, which include chemotherapy, radiotherapy, and surgery, often involves severe side effects and potential drug resistance. This comprehensive review examines the pivotal role of probiotics in cancer prevention, treatment, and management, elucidating their underlying mechanisms and clinical applications. Probiotics, defined as beneficial microorganisms that colonize the human gastrointestinal tract and other mucosal surfaces, have emerged as potential adjuncts in the prevention and treatment of cancer. The mechanisms of action include modulating the tumor microenvironment (TME), enhancing immune responses, and inhibiting carcinogenesis. In cancer prevention, probiotics can modulate the gut microbiota to inhibit carcinogen generation. For example, specific strains of Lactobacillus and Bifidobacterium have been shown to decrease the activity of enzymes involved in carcinogen production, such as β-glucuronidase and nitroreductase. Moreover, Probiotics and their metabolites, such as short-chain fatty acids (SCFAs) and indole compounds, play an antitumor role by regulating the tumor microenvironment such as regulating cancer-related gene expression, the PI3K-AKT signaling pathway, and the tryptophan-indole metabolic pathway. In the context of adjuvant therapy for malignant tumors, probiotics have shown inhibitory effects on various cancers in the digestive and reproductive systems. They can modulate the intestinal microenvironment, influence tumor cell proliferation and apoptosis, and ultimately suppress tumor growth. Additionally, probiotics can alleviate the adverse effects of cancer therapies. For example, they can mitigate chemotherapy-induced diarrhea and radiation-induced mucositis, and promote postoperative recovery by enhancing gut barrier function and reducing inflammation. This review offers a comprehensive and systematic synthesis of research on the role of probiotics in the prevention and adjuvant treatment of malignant tumors. It delves into their potential mechanisms of action and explores their clinical applications, aiming to establish a solid theoretical foundation and practical guidance for the integrated management of cancer. Looking ahead, the integration of synthetic biology with probiotics holds significant potential for cancer therapy. Advances in synthetic biology have enabled the enhancement of the anti-tumor efficacy of probiotics through genetic engineering. Engineered strains, such as Escherichia coli Nissle 1917 and attenuated Salmonella typhimurium VNP20009, have shown potential in tumor-targeted therapy. When combined with emerging technologies such as nanotechnology and photodynamic therapy, the application of probiotics in cancer treatment is expected to become more precise and effective. However, the safety and efficacy of engineered probiotics require further validation, particularly regarding the potential risks associated with long-term use. Future research should concentrate on personalized probiotic applications, the development of engineered strains, and their synergistic effects with other therapeutic modalities to advance this field. In conclusion, probiotics hold significant promise as adjuncts in cancer prevention and treatment, with the potential to modulate the TME, enhance immune responses, and alleviate treatment-related side effects. Further research is necessary to fully elucidate their mechanisms of action and optimize their clinical application, thereby facilitating their integration into comprehensive cancer care strategies. {L-End}

    Extracellular multi-enzyme assembly and biocatalytic cascade: advances and prospects
    MA Muqing, WU Yan, QU Maohua, LU Xiafeng, CAO Min, DU Feng, JI Rongtao, DONG Leichi, LUO Zhibo
    2025, 6(4):  920-939.  doi:10.12211/2096-8280.2025-056
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    Amidst the accelerating global efforts toward carbon neutrality, sustainable biomanufacturing has emerged as a crucial alternative for energy-intensive and environmentally harmful chemical synthesis. Within this transformative landscape, Multi-Enzyme Cascade Reactions (MECRs) represent a paradigm-shifting biocatalytic platform, harnessing the orchestrated activity of spatially organized enzyme modules to enable efficient, one-pot transformations with inherent cofactor recycling capabilities. This comprehensive review synthesizes cutting-edge advances in the design, optimization, and deployment of MECRs, offering a unified analysis across molecular, technological, and application dimensions. We critically dissect the mechanistic foundations of MECRs, including enzyme-enzyme synergy, substrate channeling phenomena, and allosteric regulation governing reaction flux. A systematic classification framework delineates cascade topologies—linear, convergent, parallel, and cyclic systems—elucidating their distinct kinetic advantages and thermodynamic constraints. Transformative technological innovations are highlighted, encompassing AI-driven de novo enzyme design, advanced co-immobilization strategies (such as protein scaffolds and biomimetic mineralization), nano-confined spatial organization for enhanced mass transfer, and novel cofactor regeneration systems utilizing light or electrical energy. The integration of computational fluid dynamics (CFD) modeling with microenvironmental optimization (such as pH and ionic strength gradients) has been shown to significantly enhance biocatalytic efficiency, stability, and operational lifetime. The relevance of industrial application is substantiated by compelling case studies that demonstrate the synthesis of high-value compounds driven by MECR, including optically pure pharmaceutical intermediates, bio-based polymers, and platform chemicals. These processes achieve superior atom economy (>90%), significant reductions in the E-factor, and near-quantitative yields under mild aqueous conditions. The inherent competitive advantages of MECRs—unmatched stereoselectivity, ambient operational parameters, and intrinsic sustainability—are rigorously contrasted against conventional chemocatalytic processes. Persistent challenges in enzyme inactivation, cofactor economics, and reactor scalability are objectively evaluated, alongside emerging mitigation strategies such as continuous-flow membrane bioreactors, artificial metabolon engineering, and machine learning-guided network optimization. Forward-looking perspectives outline a roadmap for next-generation MECRs, prioritizing dynamic spatiotemporal control of microenvironments, AI-accelerated evolution of hyperstable enzymes, and modular continuous manufacturing platforms. The strategic convergence of synthetic biology, computational enzyme engineering, and intelligent process control is poised to unlock programmable biocatalytic systems for complex, multi-step syntheses. This review establishes a foundational framework for advancing MECRs from bench-scale curiosities to industrially robust, environmentally transformative technologies, ultimately positioning enzyme cascades as a cornerstone of carbon-negative manufacturing and a pivotal enabler of global net-zero ambitions. {L-End}

    A comparative analysis of global research and development competition in synthetic biology
    WU Xiaoyan, SONG Qi, XU Rui, DING Chenjun, CHEN Fang, GUO Qing, ZHANG Bo
    2025, 6(4):  940-955.  doi:10.12211/2096-8280.2024-087
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    Synthetic biology is an advanced interdisciplinary field that merges biology, engineering, and technology, emerging as a strategic focus for major economies globally. This study offers a detailed comparative assessment of synthetic biology research and development in the United States, Europe, and China. Through systematic analysis of policy infrastructure, research achievements, technological advancements, and industrial applications, we have identified distinct competitive patterns across these regions. Our findings show that the United States maintains a clear leadership position, particularly in fundamental research, with significant technological capabilities and intellectual property assets. Europe has built an effective regional innovation ecosystem, achieving notable success in research commercialization through integrated collaboration among academic, research, and industrial sectors. While China has made substantial recent investments in synthetic biology, there remain opportunities to enhance high-impact research output, valuable patent development, and technology transfer efficiency compared to Western nations. Notably, China has achieved a leading position in specific areas (cell-free synthetic biology) and major applications (such as CO2-to-starch synthesis and cellulose-to-starch conversion). Based on the above analysis, this paper recommends promoting the high-quality development of China’s synthetic biology industry through five key approaches: improving top-level policy design, strengthening enterprises’ role as primary innovators, achieving breakthroughs in core technologies, optimizing intellectual property strategy, and enhancing the industrial ecosystem. These recommendations aim to promote the sustainable development of China’s synthetic biology sector. This study provides valuable strategic guidance for decision-makers, researchers, and industry stakeholders, aiding them in navigating the global competitive landscape and making informed decisions regarding the development of synthetic biology. {L-End}

    Research Article
    Development and application of a high-throughput microbial clone picking workstation based on machine vision
    ZHANG Jiankang, WANG Wenjun, GUO Hongju, BAI Beichen, ZHANG Yafei, YUAN Zheng, LI Yanhui, LI Hang
    2025, 6(4):  956-971.  doi:10.12211/2096-8280.2025-038
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    Microbial clone picking, a crucial step in genetic engineering and biological experiments, involves the accurate and rapid isolation of single colonies with desired characteristics from petri dishes teeming with numerous clones, followed by their inoculation into culture media for further propagation or analysis. In high-throughput settings, this task becomes burdensome due to its vast volume, complex record-keeping requirements, and the risk of cross-contamination, rendering manual operations impractical for achieving timely and precise results. To address this challenge, we present the design and manufacture of an automated clone picking workstation that performs efficient clone picking using 96-channel pneumatic pick-up pins, eliminating the need for consumables. The pins can be reused after ultrasonic cleaning and sterilization at high temperature following the previous picking cycle, making it more economical and environmentally friendly, compared with other methods that use disposable pipettes or picking needles. The pins can be replaced to adapt to different types of bacterial strains to meet various experimental requirements. The grab integrated on the picking head can rotate 360° and transfer the plates to different work positions.In the aspect of colony detection, the photos are automatically taken by an optical system, and the positioning and screening of colonies are achieved through the deep learning of numerous colony images by the software, which was designed independently. The precision image recognition technology is coupled with robotic and automated control technologies to enable seamless processes for picking, inoculating, cleaning, and drying. The High-Efficiency Particulate Air Filter and ultraviolet sterilization prevent cross-contamination, ensuring the experimental environment meets the required standards. This workstation is equipped with an independent operation computer and has developed a set of user-friendly software that enables personalized editing of multiple experimental protocols tailored to diverse microbial clone types. It can also communicate with external devices via TCP/IP protocols, facilitating the integration for conducting experiments such as fully automated synthetic biology. The validation experiment of bacterial colony picking was conducted by a prototype machine to test the selection efficiency. The success of the experiment suggests that the proposed system and method are feasible and effective, offering a valuable tool and a practical approach for the automation development of high-throughput laboratories. {L-End}

    Biosynthesis of xylo-oligosaccharides from wheat straw xylan through the synergistic hydrolysis by xylanase Xyn11A and arabinofuranosidase Abf62A
    HU Die, XU Daozhu, LU Zhiyi, TANG Wei, FAN Bo, HE Yucai
    2025, 6(4):  972-986.  doi:10.12211/2096-8280.2025-037
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    Xylo-oligosaccharides (XOSs) are a category of functional oligosaccharides primarily composed of 2-7 xylose units linked by β-1,4 glycosidic bonds. They are recognized as soluble dietary fibers with prebiotic properties. Recently, there has been significant interest in manufacturing XOSs from xylan extracted from lignocellulosic biomass using enzyme catalysis under mild conditions. In this work, the arabinofuranosidase Abf62A gene was cloned from Aspergillus usamii genomic DNA through sequential molecular processes and expressed in Pichia pastoris X33. The xylan (100 g/L) extracted xylan in wheat straw (WS) was biologically hydrolyzed into 50.32 g/L of XOSs by xylanase Xyn11A (300 U/g substrate) and arabinofuranase Abf62A (20 U/g substrate), which indicated a notable synergistic effect compared to the 34.42 g/L XOSs produced via Xyn11A. The 50.32 g/L of XOSs products comprised xylobiose (31.71 g/L), xylotriose (15.92 g/L), xylotetraose (1.65 g/L) and xylopentaose (1.04 g/L). Notably, the combined content of xylobiose and xylotriose accounted for up to 94.7%. The XOSs purified from the enzyme hydrolysate could effectually scavenge free radicals, and the antioxidant activity was more than 90%. In summary, XOSs were biologically manufactured from wheat straw xylan through the synergistic biocatalysis via xylanase and arabinofuranosidase Abf62A in a green and sustainable way, rending one kind of prebiotic oligosaccharides with substantial positive effects on human and animal health. {L-End}