Pub Date : 2025-12-26DOI: 10.1016/j.biotechadv.2025.108789
Huan Liu , Lunjie Wu , Songyin Zhao , Yan Xu , Yao Nie
Artificial multi-enzyme cascades utilize enzymatic catalysis to achieve continuous complex biosynthesis, thus, standing out as a promising approach. The remarkable efficiency of cascade reactions arises from the precise coordination among multiple enzymes, which facilitates efficient intermediate transfer and maximizing pathway flux. This coordination closely parallels the precise and synchronized collaboration of performers in a symphony orchestra. This review summarizes the value-added biosynthetic capabilities of multi-enzyme cascades, focusing on their ability to convert inexpensive substrates into high-value complex products. Various working forms of multi-enzyme systems are presented and primarily classified into four approaches: free enzymes, assembled complexes, fusion enzymes, and bio-based immobilized enzymes, with emphasis on the unique value of confined microenvironments as crucial platforms for achieving highly efficient cascade reactions. Furthermore, we emphasize the spatial architecture and dynamic regulation of multi-enzyme complexes, while exploring strategies for the rational design of artificial multi-enzyme assemblies tailored to cascade reaction requirements. Finally, emerging trends in AI-assisted design of cascade reactions and multi-enzyme complexes are highlighted to guide future developments.
{"title":"Enzyme symphony in bio-inspired multi-enzyme cascades for enhanced biosynthesis","authors":"Huan Liu , Lunjie Wu , Songyin Zhao , Yan Xu , Yao Nie","doi":"10.1016/j.biotechadv.2025.108789","DOIUrl":"10.1016/j.biotechadv.2025.108789","url":null,"abstract":"<div><div>Artificial multi-enzyme cascades utilize enzymatic catalysis to achieve continuous complex biosynthesis, thus, standing out as a promising approach. The remarkable efficiency of cascade reactions arises from the precise coordination among multiple enzymes, which facilitates efficient intermediate transfer and maximizing pathway flux. This coordination closely parallels the precise and synchronized collaboration of performers in a symphony orchestra. This review summarizes the value-added biosynthetic capabilities of multi-enzyme cascades, focusing on their ability to convert inexpensive substrates into high-value complex products. Various working forms of multi-enzyme systems are presented and primarily classified into four approaches: free enzymes, assembled complexes, fusion enzymes, and bio-based immobilized enzymes, with emphasis on the unique value of confined microenvironments as crucial platforms for achieving highly efficient cascade reactions. Furthermore, we emphasize the spatial architecture and dynamic regulation of multi-enzyme complexes, while exploring strategies for the rational design of artificial multi-enzyme assemblies tailored to cascade reaction requirements. Finally, emerging trends in AI-assisted design of cascade reactions and multi-enzyme complexes are highlighted to guide future developments.</div></div>","PeriodicalId":8946,"journal":{"name":"Biotechnology advances","volume":"87 ","pages":"Article 108789"},"PeriodicalIF":12.5,"publicationDate":"2025-12-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145845091","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-12-26DOI: 10.1016/j.biotechadv.2025.108791
Wei Song , Duo Wang , Jinming Li , Rui Zhang
Spatial transcriptomics (ST) is a significant advancement in life science research, enabling transcriptome analysis to transition from traditional bulk and single-cell levels to spatial location levels, thereby expanding the boundaries of biological research and pathological diagnosis. This technological breakthrough has provided unprecedented insights into complex biological processes, disease mechanisms, and clinical diagnosis. Despite the impressive advances in the field in recent years, it still faces several challenges, including technical complexity, difficulties in data analysis, and the lack of standardization. This review provides a comprehensive comparison of the technical principles and data analysis processes of ST, while also summarizing its latest applications and the current state of standardization. It aims to provide researchers with a clear framework for understanding the progresses, challenges, and future directions, thereby promoting the further development and clinical transition of ST technologies.
{"title":"Spatial transcriptomics: integrating platforms and computational approaches for clinical insights","authors":"Wei Song , Duo Wang , Jinming Li , Rui Zhang","doi":"10.1016/j.biotechadv.2025.108791","DOIUrl":"10.1016/j.biotechadv.2025.108791","url":null,"abstract":"<div><div>Spatial transcriptomics (ST) is a significant advancement in life science research, enabling transcriptome analysis to transition from traditional bulk and single-cell levels to spatial location levels, thereby expanding the boundaries of biological research and pathological diagnosis. This technological breakthrough has provided unprecedented insights into complex biological processes, disease mechanisms, and clinical diagnosis. Despite the impressive advances in the field in recent years, it still faces several challenges, including technical complexity, difficulties in data analysis, and the lack of standardization. This review provides a comprehensive comparison of the technical principles and data analysis processes of ST, while also summarizing its latest applications and the current state of standardization. It aims to provide researchers with a clear framework for understanding the progresses, challenges, and future directions, thereby promoting the further development and clinical transition of ST technologies.</div></div>","PeriodicalId":8946,"journal":{"name":"Biotechnology advances","volume":"87 ","pages":"Article 108791"},"PeriodicalIF":12.5,"publicationDate":"2025-12-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145845087","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-12-23DOI: 10.1016/j.biotechadv.2025.108784
Dhruvkumar Hariharbhai Soni , V. Reghellin , G. Sbarufatti , P. Minghetti , A. Altomare
Viral safety remains a fundamental requirement in the manufacturing of monoclonal antibodies (mAbs), particularly due to the widespread use of mammalian cell lines susceptible to both endogenous and adventitious viral contamination. This review provides a comprehensive overview of current viral clearance strategies integrated into downstream processing (DSP), highlighting the mechanisms, performance, and practical implementation of key unit operations. Chromatographic methods, including Protein A affinity, ion exchange (CEX and AEX), hydrophobic interaction (HIC), and mixed-mode chromatography (MMC), contribute to virus removal to varying extents, depending on virus type, resin chemistry, and process conditions. Anion exchange membranes have demonstrated high log reduction values (LRVs), especially for small non-enveloped viruses, while mixed-mode resins enhance removal through dual-mode interactions. Dedicated viral inactivation steps, such as low-pH incubation and detergent treatment, remain effective against enveloped viruses, with the use of stabilizing agents like arginine and extremolytes increasingly adopted to preserve product quality. Virus filtration continues to represent the most robust barrier to small viruses, though its performance depends on parameters such as filter material, fouling tendency, and viral load. Emerging solutions, such as activated carbon filtration and membrane chromatography, offer scalable, orthogonal alternatives compatible with disposable and continuous processing formats. Notably, viral clearance strategies have been successfully incorporated into continuous downstream workflows, including multicolumn capture, inline inactivation, and extended-duration filtration. Collectively, these advances support the transition toward more flexible, efficient, and sustainable viral safety frameworks, paving the way for next-generation biomanufacturing platforms.
{"title":"Viral clearance in biopharmaceutical manufacturing: Current strategies, challenges, and future directions","authors":"Dhruvkumar Hariharbhai Soni , V. Reghellin , G. Sbarufatti , P. Minghetti , A. Altomare","doi":"10.1016/j.biotechadv.2025.108784","DOIUrl":"10.1016/j.biotechadv.2025.108784","url":null,"abstract":"<div><div>Viral safety remains a fundamental requirement in the manufacturing of monoclonal antibodies (mAbs), particularly due to the widespread use of mammalian cell lines susceptible to both endogenous and adventitious viral contamination. This review provides a comprehensive overview of current viral clearance strategies integrated into downstream processing (DSP), highlighting the mechanisms, performance, and practical implementation of key unit operations. Chromatographic methods, including Protein A affinity, ion exchange (CEX and AEX), hydrophobic interaction (HIC), and mixed-mode chromatography (MMC), contribute to virus removal to varying extents, depending on virus type, resin chemistry, and process conditions. Anion exchange membranes have demonstrated high log reduction values (LRVs), especially for small non-enveloped viruses, while mixed-mode resins enhance removal through dual-mode interactions. Dedicated viral inactivation steps, such as low-pH incubation and detergent treatment, remain effective against enveloped viruses, with the use of stabilizing agents like arginine and extremolytes increasingly adopted to preserve product quality. Virus filtration continues to represent the most robust barrier to small viruses, though its performance depends on parameters such as filter material, fouling tendency, and viral load. Emerging solutions, such as activated carbon filtration and membrane chromatography, offer scalable, orthogonal alternatives compatible with disposable and continuous processing formats. Notably, viral clearance strategies have been successfully incorporated into continuous downstream workflows, including multicolumn capture, inline inactivation, and extended-duration filtration. Collectively, these advances support the transition toward more flexible, efficient, and sustainable viral safety frameworks, paving the way for next-generation biomanufacturing platforms.</div></div>","PeriodicalId":8946,"journal":{"name":"Biotechnology advances","volume":"87 ","pages":"Article 108784"},"PeriodicalIF":12.5,"publicationDate":"2025-12-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145822889","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Uridine Diphosphate glycosyltransferases (UGTs) catalyze the transfer of glycosyl moieties from donors to acceptors, a modification critical for plant growth, development, and metabolic homeostasis. These enzymes are ubiquitous across all life domains, playing key roles in the biosynthesis of diverse glycosides. This review focuses on the plant UGT708 family, which is uniquely characterized by its ability to form stable C-glycosidic bonds on flavonoid backbones, enhancing metabolic stability and bioactivity. We conducted a comprehensive analysis of UGT distribution and functional evolution across life forms, highlighting their evolutionary significance and diversification. Emphasizing plant-specific adaptations, UGT708 enzymes specialize in C-glycosylation of flavonoid, polyphenolic compounds, diketones and aromatic hydrocarbons, particularly 2-hydroxyflavanones and polyhydroxy ketones, facilitating the production of defense metabolites such as schaftosides and isoschaftosides which enhance plant resilience to environmental stresses. Clade-specific variations in the conserved PSPG motif, notably in monocots, correlate with differences in sugar-donor specificity and substrate promiscuity, reflecting structural and functional diversity within the family. Structural analyses reveal key active site residues responsible for selective C-glycosylation, resulting in a broad spectrum of glycosides with ecological and therapeutic importance. While UGT708s hold great promise as biocatalysts for generating bioactive compounds, more detailed studies on their molecular interactions and catalytic mechanisms are essential to fully exploit their potential in agriculture, medicine, and industry.
{"title":"UGT708 glycosyltransferases: Nature's architects of C-glycosides","authors":"Bhawna Verma , Palak Arora , Shahnawaz Hussain , Ritu Devi , Suphla Gupta","doi":"10.1016/j.biotechadv.2025.108787","DOIUrl":"10.1016/j.biotechadv.2025.108787","url":null,"abstract":"<div><div>Uridine Diphosphate glycosyltransferases (UGTs) catalyze the transfer of glycosyl moieties from donors to acceptors, a modification critical for plant growth, development, and metabolic homeostasis. These enzymes are ubiquitous across all life domains, playing key roles in the biosynthesis of diverse glycosides. This review focuses on the plant UGT708 family, which is uniquely characterized by its ability to form stable <em>C</em>-glycosidic bonds on flavonoid backbones, enhancing metabolic stability and bioactivity. We conducted a comprehensive analysis of UGT distribution and functional evolution across life forms, highlighting their evolutionary significance and diversification. Emphasizing plant-specific adaptations, UGT708 enzymes specialize in <em>C</em>-glycosylation of flavonoid, polyphenolic compounds, diketones and aromatic hydrocarbons, particularly 2-hydroxyflavanones and polyhydroxy ketones, facilitating the production of defense metabolites such as schaftosides and isoschaftosides which enhance plant resilience to environmental stresses. Clade-specific variations in the conserved PSPG motif, notably in monocots, correlate with differences in sugar-donor specificity and substrate promiscuity, reflecting structural and functional diversity within the family. Structural analyses reveal key active site residues responsible for selective <em>C</em>-glycosylation, resulting in a broad spectrum of glycosides with ecological and therapeutic importance. While UGT708s hold great promise as biocatalysts for generating bioactive compounds, more detailed studies on their molecular interactions and catalytic mechanisms are essential to fully exploit their potential in agriculture, medicine, and industry.</div></div>","PeriodicalId":8946,"journal":{"name":"Biotechnology advances","volume":"87 ","pages":"Article 108787"},"PeriodicalIF":12.5,"publicationDate":"2025-12-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145822890","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Enzyme engineering involves enhancing enzyme function and application through multidimensional technological systems. Its development encompasses elucidating sequence-structure-function relationships, exploring fitness landscapes, and multiscale regulation. Conventional enzyme engineering strategies include directed evolution (DE), rational/semi-rational design, residue co-evolution, and de novo design. DE mimics natural selection but is limited by high-throughput screening efficiency. Rational/semi-rational design integrates computational simulation with experimental validation to regulate enzyme performance. Residue co-evolution combines sequence co-evolution analysis and kinetic simulations, and de novo design is dedicated to achieving precise protein folding through physical modeling. In recent years, the breakthrough progress in machine learning (ML), especially deep learning (DL), has significantly enhanced the efficiency of all the above-mentioned methods; by accurately predicting mutational effects and efficiently exploring discontinuous sequence space, it provides powerful tools to improve or supplement the above strategies, thereby aiding in escaping local fitness optima traps. This review summarizes common strategies in enzyme engineering, including directed evolution, rational/semi-rational design, residue co-evolution, and de novo design, and further introduces the latest applications of ML and DL models in each of these fields. Although challenges persist, such as force field accuracy limitations, mutation sampling constraints, experimental throughput limitations, and epistatic effects, more comprehensive multimodal foundation models in the future are expected to integrate cross-scale parameters for intelligent design, and the establishment of standardized enzymology databases will enhance prediction reliability. Overall, AI-empowered enzyme engineering will drive a profound transformation toward a predictable and highly efficient pathway for enzyme design, providing more precise and powerful solutions for biocatalysis.
{"title":"From traditional to AI-driven: The evolution of intelligent enzyme engineering for biocatalysis","authors":"Feifei Lv , Jiaxing Zhang , Shengping You , Wei Qi","doi":"10.1016/j.biotechadv.2025.108788","DOIUrl":"10.1016/j.biotechadv.2025.108788","url":null,"abstract":"<div><div>Enzyme engineering involves enhancing enzyme function and application through multidimensional technological systems. Its development encompasses elucidating sequence-structure-function relationships, exploring fitness landscapes, and multiscale regulation. Conventional enzyme engineering strategies include directed evolution (DE), rational/semi-rational design, residue co-evolution, and de novo design. DE mimics natural selection but is limited by high-throughput screening efficiency. Rational/semi-rational design integrates computational simulation with experimental validation to regulate enzyme performance. Residue co-evolution combines sequence co-evolution analysis and kinetic simulations, and de novo design is dedicated to achieving precise protein folding through physical modeling. In recent years, the breakthrough progress in machine learning (ML), especially deep learning (DL), has significantly enhanced the efficiency of all the above-mentioned methods; by accurately predicting mutational effects and efficiently exploring discontinuous sequence space, it provides powerful tools to improve or supplement the above strategies, thereby aiding in escaping local fitness optima traps. This review summarizes common strategies in enzyme engineering, including directed evolution, rational/semi-rational design, residue co-evolution, and de novo design, and further introduces the latest applications of ML and DL models in each of these fields. Although challenges persist, such as force field accuracy limitations, mutation sampling constraints, experimental throughput limitations, and epistatic effects, more comprehensive multimodal foundation models in the future are expected to integrate cross-scale parameters for intelligent design, and the establishment of standardized enzymology databases will enhance prediction reliability. Overall, AI-empowered enzyme engineering will drive a profound transformation toward a predictable and highly efficient pathway for enzyme design, providing more precise and powerful solutions for biocatalysis.</div></div>","PeriodicalId":8946,"journal":{"name":"Biotechnology advances","volume":"87 ","pages":"Article 108788"},"PeriodicalIF":12.5,"publicationDate":"2025-12-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145823734","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-12-21DOI: 10.1016/j.biotechadv.2025.108774
Thomas G. Neuman , Surya Karla , James (Chip) Kilduff , Joel Plawsky , Georges Belfort
The RNA revolution, advancing beyond traditional vaccines to new therapeutic modalities and constructs such as self-amplifying RNA (saRNA) and circular RNA (circRNA), continues to place increasing pressure on downstream purification. Diffusion-limited resins, the time-tested workhorse of protein purification, are fundamentally incompatible because mRNA is an enormous (>40 nm) molecule with low diffusivity (10−11–10−12 m2/s). Our perspective, rooted in core chemical engineering principles, applies transport analysis and re-examines published performance data to demonstrate why even optimized perfusion chromatographic resin systems, exhibiting 0.1 % of total flow through the resin particles, cannot overcome the inherent diffusional barriers preventing efficient RNA purification. Alternatively, convection-based devices, notably membranes and monoliths, are well situated as their transport characteristics are not limited by the molecular transport properties of RNA. Ultimately, pressure-driven flow enables the potential for increased device capacity at orders of magnitude (103x) lower process time and smaller device footprint contributing to markedly improved productivity. Taken together, these findings suggest that a paradigm shift is required toward convective membrane systems to create a platform capable of delivering scalable, economic, and ultimately industrially attractive mRNA purification.
{"title":"Membranes: The only chance for the mRNA tortoise to win, economically","authors":"Thomas G. Neuman , Surya Karla , James (Chip) Kilduff , Joel Plawsky , Georges Belfort","doi":"10.1016/j.biotechadv.2025.108774","DOIUrl":"10.1016/j.biotechadv.2025.108774","url":null,"abstract":"<div><div>The RNA revolution, advancing beyond traditional vaccines to new therapeutic modalities and constructs such as self-amplifying RNA (saRNA) and circular RNA (circRNA), continues to place increasing pressure on downstream purification. Diffusion-limited resins, the time-tested workhorse of protein purification, are fundamentally incompatible because mRNA is an enormous (>40 nm) molecule with low diffusivity (10<sup>−11</sup>–10<sup>−12</sup> m<sup>2</sup>/s). Our perspective, rooted in core chemical engineering principles, applies transport analysis and re-examines published performance data to demonstrate why even optimized perfusion chromatographic resin systems, exhibiting 0.1 % of total flow through the resin particles, cannot overcome the inherent diffusional barriers preventing efficient RNA purification. Alternatively, convection-based devices, notably membranes and monoliths, are well situated as their transport characteristics are not limited by the molecular transport properties of RNA. Ultimately, pressure-driven flow enables the potential for increased device capacity at orders of magnitude (10<sup>3</sup>x) lower process time and smaller device footprint contributing to markedly improved productivity. Taken together, these findings suggest that a paradigm shift is required toward convective membrane systems to create a platform capable of delivering scalable, economic, and ultimately industrially attractive mRNA purification.</div></div>","PeriodicalId":8946,"journal":{"name":"Biotechnology advances","volume":"87 ","pages":"Article 108774"},"PeriodicalIF":12.5,"publicationDate":"2025-12-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145813855","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-12-20DOI: 10.1016/j.biotechadv.2025.108783
Zheng-Hui Li , Shuang Li , Hai-Bo Chen , Yi-Zhi Ji , Chong Zhang
The growing demand for novel antibiotics, industrial enzymes, and environmentally sustainable biotechnological solutions is drawing increasing attention to more efficient strategies for microbial resource discovery across biomedical, industrial, and ecological domains. To meet this need, high-throughput microbial culturomics has emerged as a powerful strategy that integrates advanced cultivation platforms, diverse growth conditions, and rapid identification technologies. This review explores two principal paradigms within high-throughput microbial culturomics: the function-driven screening-first strategy and the enrichment-based cultivation-first strategy. Their technical foundations, application scenarios, and inherent limitations are examined in detail, providing a comparative analysis of their respective advantages and challenges. We further emphasize the importance of high-throughput identification methods, which play a crucial role in classifying isolates and revealing their functional potential. Ultimately, the review explores future directions for the development of automated, fully integrated cultivation platforms that integrate large-scale experimentation with data-driven optimization. By offering a structured comparison of culturomics strategies, integration pathways, and key obstacles, this review serves as a methodological reference for advancing microbial isolation, functional screening, and biotechnological innovation.
{"title":"The advances of strategies and technologies in high-throughput microbial culturomics","authors":"Zheng-Hui Li , Shuang Li , Hai-Bo Chen , Yi-Zhi Ji , Chong Zhang","doi":"10.1016/j.biotechadv.2025.108783","DOIUrl":"10.1016/j.biotechadv.2025.108783","url":null,"abstract":"<div><div>The growing demand for novel antibiotics, industrial enzymes, and environmentally sustainable biotechnological solutions is drawing increasing attention to more efficient strategies for microbial resource discovery across biomedical, industrial, and ecological domains. To meet this need, high-throughput microbial culturomics has emerged as a powerful strategy that integrates advanced cultivation platforms, diverse growth conditions, and rapid identification technologies. This review explores two principal paradigms within high-throughput microbial culturomics: the function-driven screening-first strategy and the enrichment-based cultivation-first strategy. Their technical foundations, application scenarios, and inherent limitations are examined in detail, providing a comparative analysis of their respective advantages and challenges. We further emphasize the importance of high-throughput identification methods, which play a crucial role in classifying isolates and revealing their functional potential. Ultimately, the review explores future directions for the development of automated, fully integrated cultivation platforms that integrate large-scale experimentation with data-driven optimization. By offering a structured comparison of culturomics strategies, integration pathways, and key obstacles, this review serves as a methodological reference for advancing microbial isolation, functional screening, and biotechnological innovation.</div></div>","PeriodicalId":8946,"journal":{"name":"Biotechnology advances","volume":"87 ","pages":"Article 108783"},"PeriodicalIF":12.5,"publicationDate":"2025-12-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145796178","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-12-18DOI: 10.1016/j.biotechadv.2025.108782
Jiwoo Nam , Yuna Lee , Sion Lee , Hyungjun Choi , Sang Yup Lee , Dongsoo Yang
Microorganisms inhabit diverse environments, including nearly every organ in the human body. The human microbiome—a complex community of microorganisms residing in the human body—has gained increasing attention as a key contributor to human health and disease, making it an important target for the development of diagnostic and therapeutic strategies. However, the inherent complexity of microbial communities and the challenges of engineering diverse non-model microorganisms present significant barriers. To address these challenges, synthetic biology has provided powerful tools and strategies to engineer microorganisms capable of sensing disease-specific environments and performing targeted therapeutic functions. In particular, the development of synthetic genetic circuits has significantly improved the precision and reliability of disease diagnosis and treatment, enabling real-time disease monitoring, therapeutic, and even preventive interventions. This review highlights state-of-the-art synthetic biology tools and strategies for engineering the probiotics and commensal bacteria aimed at the diagnosis and treatment of human diseases, with accompanying examples. Future challenges and prospects are also discussed.
{"title":"Synthetic biology strategies for engineering probiotics and commensal bacteria for diagnostics and therapeutics","authors":"Jiwoo Nam , Yuna Lee , Sion Lee , Hyungjun Choi , Sang Yup Lee , Dongsoo Yang","doi":"10.1016/j.biotechadv.2025.108782","DOIUrl":"10.1016/j.biotechadv.2025.108782","url":null,"abstract":"<div><div>Microorganisms inhabit diverse environments, including nearly every organ in the human body. The human microbiome—a complex community of microorganisms residing in the human body—has gained increasing attention as a key contributor to human health and disease, making it an important target for the development of diagnostic and therapeutic strategies. However, the inherent complexity of microbial communities and the challenges of engineering diverse non-model microorganisms present significant barriers. To address these challenges, synthetic biology has provided powerful tools and strategies to engineer microorganisms capable of sensing disease-specific environments and performing targeted therapeutic functions. In particular, the development of synthetic genetic circuits has significantly improved the precision and reliability of disease diagnosis and treatment, enabling real-time disease monitoring, therapeutic, and even preventive interventions. This review highlights state-of-the-art synthetic biology tools and strategies for engineering the probiotics and commensal bacteria aimed at the diagnosis and treatment of human diseases, with accompanying examples. Future challenges and prospects are also discussed.</div></div>","PeriodicalId":8946,"journal":{"name":"Biotechnology advances","volume":"87 ","pages":"Article 108782"},"PeriodicalIF":12.5,"publicationDate":"2025-12-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145784803","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-12-17DOI: 10.1016/j.biotechadv.2025.108781
Xuan Zhou, Wenyan Cao, Chao Huang, Xiaojuan Zhang, Shenghu Zhou, Yu Deng
Gene expression regulatory elements (GEREs) play a pivotal role in the control of gene transcription and translation. The design of GEREs with precise and tunable activity remains a major challenge in synthetic biology. Over the past decades, engineering strategies have evolved from empirical sequence mining and random mutagenesis to increasingly rational approaches guided by biophysical models and artificial intelligence. In this review, we systematically examine the design principles, representative studies, and implementation strategies for each GERE class, highlighting how mining, modular recombination, targeted mutagenesis, and deep generative modeling contribute to the development of functional regulatory elements. We further discuss the strengths and limitations of these strategies, offering practical guidance for optimizing microbial cell factory bioproduction through the fine-tuning of gene expression.
{"title":"Designing prokaryotic gene expression regulatory elements: From genomic mining to artificial intelligence-driven generation","authors":"Xuan Zhou, Wenyan Cao, Chao Huang, Xiaojuan Zhang, Shenghu Zhou, Yu Deng","doi":"10.1016/j.biotechadv.2025.108781","DOIUrl":"10.1016/j.biotechadv.2025.108781","url":null,"abstract":"<div><div>Gene expression regulatory elements (GEREs) play a pivotal role in the control of gene transcription and translation. The design of GEREs with precise and tunable activity remains a major challenge in synthetic biology. Over the past decades, engineering strategies have evolved from empirical sequence mining and random mutagenesis to increasingly rational approaches guided by biophysical models and artificial intelligence. In this review, we systematically examine the design principles, representative studies, and implementation strategies for each GERE class, highlighting how mining, modular recombination, targeted mutagenesis, and deep generative modeling contribute to the development of functional regulatory elements. We further discuss the strengths and limitations of these strategies, offering practical guidance for optimizing microbial cell factory bioproduction through the fine-tuning of gene expression.</div></div>","PeriodicalId":8946,"journal":{"name":"Biotechnology advances","volume":"87 ","pages":"Article 108781"},"PeriodicalIF":12.5,"publicationDate":"2025-12-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145784804","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Flavin reductases (FRs) are essential redox enzymes that supply reduced flavin cofactors (FMNH−/FADH−) to various monooxygenase partners in two-component flavin-dependent monooxygenase (TC-FDMO) systems. These enzymes play critical roles in numerous biological processes and industrial biocatalytic reactions, including hydroxylation, halogenation, and epoxidation. In this review, we provide a comprehensive analysis of the structural features, oligomeric states, kinetic mechanisms, and newly proposed classification strategies of FRs. We highlight the limitations of existing classification systems that rely solely on physiological function and propose a more informative framework based on amino acid sequences and domain architectures. Detailed mechanistic insights from transient kinetics, charge-transfer complex formation, and flavin transfer pathways are discussed, with emphasis on enzyme-specific features such as half-site reactivity and substrate-enhanced catalysis. Advances in protein engineering and fusion protein design aimed at improving FR stability, catalytic performance, and cofactor regeneration are also critically evaluated. In addition, we explore alternative strategies for supplying reduced flavin to monooxygenase partners, including non-enzymatic regeneration methods and the use of nicotinamide analogs. Finally, we outline key challenges and future directions for developing next-generation FRs with enhanced industrial applicability. This knowledge provides a foundation for engineering TC-FDMO systems for scalable, sustainable, and industrially relevant biocatalysis.
{"title":"Flavin reductases in two-component systems: Mechanistic insights, structural classification, and biotechnological advances","authors":"Panu Pimviriyakul , Piyanuch Anuwan , Pimchai Chaiyen , Thanyaporn Wongnate","doi":"10.1016/j.biotechadv.2025.108779","DOIUrl":"10.1016/j.biotechadv.2025.108779","url":null,"abstract":"<div><div>Flavin reductases (FRs) are essential redox enzymes that supply reduced flavin cofactors (FMNH<sup>−</sup>/FADH<sup>−</sup>) to various monooxygenase partners in two-component flavin-dependent monooxygenase (TC-FDMO) systems. These enzymes play critical roles in numerous biological processes and industrial biocatalytic reactions, including hydroxylation, halogenation, and epoxidation. In this review, we provide a comprehensive analysis of the structural features, oligomeric states, kinetic mechanisms, and newly proposed classification strategies of FRs. We highlight the limitations of existing classification systems that rely solely on physiological function and propose a more informative framework based on amino acid sequences and domain architectures. Detailed mechanistic insights from transient kinetics, charge-transfer complex formation, and flavin transfer pathways are discussed, with emphasis on enzyme-specific features such as half-site reactivity and substrate-enhanced catalysis. Advances in protein engineering and fusion protein design aimed at improving FR stability, catalytic performance, and cofactor regeneration are also critically evaluated. In addition, we explore alternative strategies for supplying reduced flavin to monooxygenase partners, including non-enzymatic regeneration methods and the use of nicotinamide analogs. Finally, we outline key challenges and future directions for developing next-generation FRs with enhanced industrial applicability. This knowledge provides a foundation for engineering TC-FDMO systems for scalable, sustainable, and industrially relevant biocatalysis.</div></div>","PeriodicalId":8946,"journal":{"name":"Biotechnology advances","volume":"87 ","pages":"Article 108779"},"PeriodicalIF":12.5,"publicationDate":"2025-12-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145780161","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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