ACS Synthetic Biology最新文献_第10页

Engineering Modular Cargo Loading Strategies for Carboxysome-Derived Protein Particles 羧酶体衍生蛋白颗粒的工程模块化装载策略。

IF 4.5 2区生物学

ACS Synthetic Biology Pub Date : 2026-08-21 Epub Date: 2026-07-01 DOI: 10.1021/acssynbio.6c00354

Claudia A. Mak, Rachelle M. Baumann, Anthony G. Vecchiarelli

{"title":"Engineering Modular Cargo Loading Strategies for Carboxysome-Derived Protein Particles","authors":"Claudia A. Mak, Rachelle M. Baumann, Anthony G. Vecchiarelli","doi":"10.1021/acssynbio.6c00354","DOIUrl":"10.1021/acssynbio.6c00354","url":null,"abstract":"Bacterial microcompartments(BMCs) are a diverse and widespread classof self-assembling protein-based organelles consisting of a semipermeableprotein shell encapsulating an enzymatic core. Isolated BMC shellproteins have been shown to assemble into alternative superstructuressuch as flat sheets and nanotubes. The self-assembly and modularityof BMC shell proteins make them of great interest as modular platformsfor applications involving scaffolding, immobilization, and compartmentalization.While the assembly of BMC shell proteins into higher-order structureshas been well-studied, the design of controllable and modular cargoloading is underdeveloped in comparison. Recently, we reported thepH-controlled assembly of CcmK2, the major hexameric shell proteinof the β-carboxysome BMC, into monodisperse mesh-like microscaleparticles. Here, we develop a suite of encapsulation strategies forstochastic or targeted loading of various cargos, as well as the directconjugation of cargo to CcmK2 particles. Our systematic analysis demonstratesthat cargo loading and particle assembly can be modulated by the choiceof recruitment strategy and the order of cargo introduction. Our findingsalso reveal a cooperative cargo loading mechanism during assemblythat influences particle sizing and apparent morphology. Our studyserves as a blueprint for the rational design of tunable cargo loadinginto engineered BMC-derived microcompartment systems for diverse biotechnologicalapplications.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":"15 8","pages":"3462–3473"},"PeriodicalIF":4.5,"publicationDate":"2026-08-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148358481","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Engineering 2,4-Diacetylphloroglucinol-Hyper-Tolerant Escherichia coli via Adaptive Evolution for Efficient Biosynthesis and an Organic Solvent-Free Downstream Process 2,4-二乙酰间苯三酚-超耐药大肠杆菌的适应性进化及其高效生物合成和无溶剂下游工艺。

IF 4.5 2区生物学

ACS Synthetic Biology Pub Date : 2026-08-21 Epub Date: 2026-07-10 DOI: 10.1021/acssynbio.6c00194

Jingnan Wang, Mingyue Liu, Cheng Li, Bo Jiang, Yunhuan Lu, Jiajing Ma, Min Liu, Qingsheng Qi, Jichao Wang, Guang Zhao

{"title":"Engineering 2,4-Diacetylphloroglucinol-Hyper-Tolerant Escherichia coli via Adaptive Evolution for Efficient Biosynthesis and an Organic Solvent-Free Downstream Process","authors":"Jingnan Wang, Mingyue Liu, Cheng Li, Bo Jiang, Yunhuan Lu, Jiajing Ma, Min Liu, Qingsheng Qi, Jichao Wang, Guang Zhao","doi":"10.1021/acssynbio.6c00194","DOIUrl":"10.1021/acssynbio.6c00194","url":null,"abstract":"2,4-Diacetylphloroglucinol(DAPG) is a valuable antimicrobial compoundwith significant agricultural potential, suffers bioproduction limitationsfrom host toxicity and inefficient downstream processing. This studyengineered DAPG-hyper-tolerant Escherichia coli via adaptive laboratory evolution (ALE) starting from a phloroglucinol-tolerantstrain. Optimized shake-flask fermentation of evolved Bdt03 yielded330.52 mg/L DAPG (17.84-fold of the wild type), and the yield fromthe whole fermentation broth could be further increased to 391.36mg/L. A novel organic solvent-free extraction method recovered DAPGfrom fermentation broth with over 98% yield via acidification, coldincubation and centrifugation, simplifying downstream processing.Genomic resequencing identified several key mutations underlying DAPGtolerance, which were validated and stacked to precisely constructstrain Bb03 with enhanced production and tolerance. This work addressesthe critical bottlenecks in DAPG biosynthesis by enhancing host toleranceand developing a sustainable downstream processing strategy, and alsooffers valuable genetic insights for constructing high-yield DAPG-producingstrains and advancing the application of DAPG-responsive genetic circuitsin synthetic biology.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":"15 8","pages":"3322–3335"},"PeriodicalIF":4.5,"publicationDate":"2026-08-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148416925","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Machine-Learning-Assisted Pathway Optimization in Large Combinatorial Design Spaces: A p-Coumaric Acid Case Study 大型组合设计空间中的机器学习辅助路径优化：对香豆酸案例研究。

IF 4.5 2区生物学

ACS Synthetic Biology Pub Date : 2026-08-21 Epub Date: 2026-07-22 DOI: 10.1021/acssynbio.5c00864

Paul van Lent, Rianne van der Hoek, Sara Moreno Paz, Irsan Kooi, Moniek Jonkers, Priscilla Zwartjens, Joep Schmitz, Thomas Abeel

{"title":"Machine-Learning-Assisted Pathway Optimization in Large Combinatorial Design Spaces: A p-Coumaric Acid Case Study","authors":"Paul van Lent, Rianne van der Hoek, Sara Moreno Paz, Irsan Kooi, Moniek Jonkers, Priscilla Zwartjens, Joep Schmitz, Thomas Abeel","doi":"10.1021/acssynbio.5c00864","DOIUrl":"10.1021/acssynbio.5c00864","url":null,"abstract":"Combinatorial pathwayoptimization is a powerful approach in metabolicengineering to improve strain performance. While machine learning(ML) has shown promise in guiding the Design–Build–Test–Learn(DBTL) cycle, most applications have been limited to small designspaces, thereby restricting the potential of predictive and exploration-exploitationstrategies. In this work, we applied two DBTL cycles to optimize p-coumaric acid production in Saccharomyces cerevisiae. The first cycle involved constructinga large combinatorial library of 18 genes and 20 promoters (170 millionpossible designs). In the second cycle, we employed a gradient bandit-basedmachine learning recommendation strategy, tuned to balance explorationand exploitation. Our results show that this balanced strategy outperformsgreedy, feature importance-based approaches, leading to greater diversityin strain performance and improved top-producer identification. Notably,applying the same strategy to an alternative parent strain yieldedthe highest p-coumaric acid titer (1.23 g/L), a 2.37-foldimprovement over the original. These findings highlight the valueof ML-guided exploration in large design spaces and demonstrate thatbalancing exploration and exploitation is critical for successfulstrain optimization.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":"15 8","pages":"3146–3157"},"PeriodicalIF":4.5,"publicationDate":"2026-08-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148560136","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Harnessing the Power of SMART Single-Molecule Display for Enzyme Evolution: A Focus on Oxidase 利用SMART单分子显示的力量促进酶的进化：以氧化酶为重点。

IF 4.5 2区生物学

ACS Synthetic Biology Pub Date : 2026-08-21 Epub Date: 2026-02-23 DOI: 10.1021/acssynbio.5c00968

Kalhari Munaweera, Nana Odake, Hannah Patricia Halim, Kakeru Ikeda, Bo Zhu, Maurizio Camagna, Tomokazu Ito, Tetsuya Kitaguchi, Naoto Nemoto, Hideo Nakano, Jasmina Damnjanović

{"title":"Harnessing the Power of SMART Single-Molecule Display for Enzyme Evolution: A Focus on Oxidase","authors":"Kalhari Munaweera, Nana Odake, Hannah Patricia Halim, Kakeru Ikeda, Bo Zhu, Maurizio Camagna, Tomokazu Ito, Tetsuya Kitaguchi, Naoto Nemoto, Hideo Nakano, Jasmina Damnjanović","doi":"10.1021/acssynbio.5c00968","DOIUrl":"10.1021/acssynbio.5c00968","url":null,"abstract":"For a rapid and cost-effective evolution of tailor-madeenzymes,we established a high-throughput in vitro selectionplatform named SMART (Single-Molecule Assay on Ribonucleic acid byTranslated product), integrating mRNA display, next-generation sequencing,and bioinformatics. SMART represents a versatile system where a moduletermed an auxiliary unit allows enzyme-specific selection under variousexperimental conditions. Here, we report on the establishment of SMARTfor oxidases using a model enzyme, Schizosaccharomyces pombed-amino acid oxidase (SpDAAO), andascorbate peroxidase 2 as the auxiliary enzyme to detect hydrogenperoxide produced by the oxidase, and mediate biotinylation of activesingle-molecule display complexes. As a proof-of-concept, a libraryincluding site-saturation mutagenesis at the catalytic residue Y232of SpDAAO was subjected to a single SMART selection round, yieldingenrichment of the active enzyme variant. The results demonstrate theutility of SMART as a fast, robust, and efficient platform with thepotential of customization for other enzyme chemistries through appropriatemodifications of the auxiliary unit. Using SMART, desired enzyme variantscan be selected in just a few hours by a single person without theneed for costly equipment or any bias or limitations.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":"15 8","pages":"3168–3178"},"PeriodicalIF":4.5,"publicationDate":"2026-08-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147275191","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Engineering of Riboflavin Transporter and Enzymes in Bacillus subtilis Facilitates High-Efficiency FAD Synthesis from RF 枯草芽孢杆菌核黄素转运体及酶工程促进RF高效合成FAD。

IF 4.5 2区生物学

ACS Synthetic Biology Pub Date : 2026-08-21 Epub Date: 2026-05-11 DOI: 10.1021/acssynbio.6c00153

Yunlong Wu, Yuhao Sun, Huan Miao, Xiaole Xia, Ling Gao

{"title":"Engineering of Riboflavin Transporter and Enzymes in Bacillus subtilis Facilitates High-Efficiency FAD Synthesis from RF","authors":"Yunlong Wu, Yuhao Sun, Huan Miao, Xiaole Xia, Ling Gao","doi":"10.1021/acssynbio.6c00153","DOIUrl":"10.1021/acssynbio.6c00153","url":null,"abstract":"Thebiosynthesis of flavin adenine dinucleotide (FAD),an essentialredox coenzyme, is limited by inefficient substrate utilization andlow catalytic activity. To address this, an “Uptake-Catalysis”system was constructed in Bacillus subtilis for FAD synthesis from riboflavin (RF). First, the Streptomyces davawensis transporter (RibM) was coexpressedwith native FAD synthase (BsFADs) at a 2:1 ratio,which increased RF consumption to 51%. Subsequently, the isothermalcompressibility perturbation engineering (ICPE) strategy was adopted,generating the optimum mutant BsFADsP61W+R202Y, which increased catalytic efficiency (kcat/Km) toward RF and FMN by 83% and 73%,respectively. Mechanistic analysis revealed that this improvementwas due to a 39% shortening of RF substrate channel. Finally, FADtiter peaked at 563 mg/L with a conversion rate (cr-FAD) of 92% in shaking-flask fermentation optimization. This workachieved a 225-fold increase in FAD production, validating the effectivenessof the uptake-catalysis synergistic strategy, thereby establishingan efficient and scalable synthesis paradigm.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":"15 8","pages":"3272–3284"},"PeriodicalIF":4.5,"publicationDate":"2026-08-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147865861","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Engineering an Artificial Taxol Biosynthetic Pathway from Baccatin III in Yeast 酵母Baccatin III生物合成紫杉醇的人工途径

IF 4.5 2区生物学

ACS Synthetic Biology Pub Date : 2026-08-21 Epub Date: 2026-05-26 DOI: 10.1021/acssynbio.6c00148

Yuanwei Gou, Bin Jiang, Xiaojing Jiang, Yiwei Zhang, Binbin Chen, Jiaojiao Zhang, Dongfang Li, Yajie Wang, Jianbin Yan, Jiazhang Lian

{"title":"Engineering an Artificial Taxol Biosynthetic Pathway from Baccatin III in Yeast","authors":"Yuanwei Gou, Bin Jiang, Xiaojing Jiang, Yiwei Zhang, Binbin Chen, Jiaojiao Zhang, Dongfang Li, Yajie Wang, Jianbin Yan, Jiazhang Lian","doi":"10.1021/acssynbio.6c00148","DOIUrl":"10.1021/acssynbio.6c00148","url":null,"abstract":"Paclitaxel (Taxol) is among the most successful naturalanticanceragents, yet its manufacturing still relies on expensive semisyntheticmethods carried out under harsh alkaline conditions. Deciphering theentire Taxol biosynthetic pathway has been a longstanding challengefor over 50 years because of its complex regulatory network, but ithas only recently been understood. Differences between plant and microbialsystems render current knowledge insufficient for scalable biotechnologicalproduction, necessitating systematic metabolic engineering to overcomemultiple bottlenecks. Artificial pathways can compensate for an incompleteunderstanding of native biosynthesis and offer an alternative strategyto alleviate supply constraints. Here, we report the constructionof an artificial pathway in yeast that uses (2R,3S)-3-phenylisoserine as the substrate and requires only three enzymaticsteps to convert baccatin III to Taxol, demonstrating the broad substratescope of β-phenylalanoyl-CoA ligases (PCLs). Structure–activityanalysis of Baccatin III:3-amino-3-phenylpropanoyltransferase (BAPT)is further employed to enhance its catalytic efficiency toward non-naturalsubstrates. This work offers a practical strategy for the first stepof sustainable production of Taxol using synthetic biology.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":"15 8","pages":"3261–3271"},"PeriodicalIF":4.5,"publicationDate":"2026-08-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148024939","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

A Multitask Prediction Framework for CircRNAs, Drugs, and Diseases Based on Multi-View Information Integration and Graph Contrastive Learning 基于多视图信息集成和图对比学习的circrna、药物和疾病多任务预测框架

IF 4.5 2区生物学

ACS Synthetic Biology Pub Date : 2026-08-21 Epub Date: 2026-07-01 DOI: 10.1021/acssynbio.6c00184

Shudong Wang, Bo Yue, Tiyao Liu, Wenhao Wu, Wenjing Yin, Shanchen Pang

{"title":"A Multitask Prediction Framework for CircRNAs, Drugs, and Diseases Based on Multi-View Information Integration and Graph Contrastive Learning","authors":"Shudong Wang, Bo Yue, Tiyao Liu, Wenhao Wu, Wenjing Yin, Shanchen Pang","doi":"10.1021/acssynbio.6c00184","DOIUrl":"10.1021/acssynbio.6c00184","url":null,"abstract":"Asa part of drug discovery, the exploration of circRNA-drug sensitivityassociations (CRAs), drug-disease associations (RDAs) has been animportant means of understanding disease mechanisms and proposingnew treatment strategies. Due to the time-consuming and labor-intensivenature of biological experiments, computational methods have becomeeffective solutions to this problem. However, existing computationalmethods for inferring CRAs, RDAs typically operate in isolation, failingto exploit the rich interplay across circRNAs, drugs, and diseases.In this paper, we propose MVII-GCL, a novel multitask prediction frameworkbuilt upon multiview information integration and graph contrastivelearning. MVII-GCL first constructs a multiview network encompassingattribute, topology, and association perspectives, explicitly tailoredto capture the unique characteristics of each domain. To map multiviewinformation into a unified latent space, our multiview attention encodermodule employs domain-specific encoders to learn specialized representations,followed by a Top-k sparse attention mechanism. To further bolsterrobustness against data sparsity and noise, a symmetric graph contrastivelearning objective is seamlessly integrated as a regularization termwithin the multitask learning paradigm. 5-fold cross-validation demonstratesour model’s strong performance, achieving AUC, AUPR, and F1 scores of 0.9764, 0.9750, and 0.9234 in drug-diseaseassociation prediction, and 0.9301, 0.9339, and 0.8609 in circRNA-drugsensitivity prediction.Furthermore, case studies further support themodel’s ability to discover potential associations.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":"15 8","pages":"3310–3321"},"PeriodicalIF":4.5,"publicationDate":"2026-08-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148358453","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Joined Signatures of Erythropoietin and Interleukin-6 Signaling by Synthetic Heterodimeric gp130:Epo Receptor Complexes 合成异二聚体gp130:Epo受体复合物的促红细胞生成素和白细胞介素-6信号的联合信号。

IF 4.5 2区生物学

ACS Synthetic Biology Pub Date : 2026-08-21 Epub Date: 2026-08-02 DOI: 10.1021/acssynbio.6c00029

Jonas Schlei, Karlo F. Franke, Dmitriy Holzmann, Christoph Wittich, Doreen M. Floss, Puyan Rafii, Kevin Brors, Juergen Scheller

{"title":"Joined Signatures of Erythropoietin and Interleukin-6 Signaling by Synthetic Heterodimeric gp130:Epo Receptor Complexes","authors":"Jonas Schlei, Karlo F. Franke, Dmitriy Holzmann, Christoph Wittich, Doreen M. Floss, Puyan Rafii, Kevin Brors, Juergen Scheller","doi":"10.1021/acssynbio.6c00029","DOIUrl":"10.1021/acssynbio.6c00029","url":null,"abstract":"Synthetic engineering has shown that cytokine receptorsfrom differentfamilies can be functionally combined to a large extent. We and othershave developed fully synthetic cytokine ligand–receptor systems.Here, our synthetic cytokine receptor (SyCyR) system has been usedto generate highly effective non-natural cytokine receptor complexesfor the interleukin (IL)-6-type signaling receptors gp130, LIFR, andthe erythropoietin receptor (EpoR). Epo promotes the formation ofred blood cells in the bone marrow, while IL-6 plays a role in theimmune response and regulation of inflammation. Their synergisticeffects in combination could contribute to effectiveness of anemiatreatment in future in vivo studies. Activation of these SyCyRs isbased on homo- and heterodimeric GFP-mCherry fusion proteins as syntheticcytokine ligands and VHH nanobodies (variable domain of single-domainantibodies) against GFP and mCherry fused to the transmembrane andintracellular domains of engineered cytokine receptors. Although syntheticcytokine signaling was achieved for most designs, gp130:LIFR and gp130:EpoRdesigns were more efficient compared to LIFR:EpoR designs. The gp130:EpoRsignaling combines typical signaling signature of both pathways. Takentogether, our data show that synthetic cytokine receptors for membersof the IL-6 family and EpoR are generally feasible.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":"15 8","pages":"3485–3497"},"PeriodicalIF":4.5,"publicationDate":"2026-08-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148785414","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

A Highly Stringent Split Intein-Mediated DHFR Selectable Marker Enables Efficient Development of High-Producing CHO Cells for Therapeutic Proteins 一种高度严格的分裂蛋白介导的DHFR选择标记使高效开发用于治疗蛋白的高产CHO细胞成为可能。

IF 4.5 2区生物学

ACS Synthetic Biology Pub Date : 2026-08-21 Epub Date: 2026-07-02 DOI: 10.1021/acssynbio.6c00133

Eun-Ji Lee, Chansik Yoon, Na-Yeong Heo, So-Hui Ryu, Gyun Min Lee, Yeon-Gu Kim

{"title":"A Highly Stringent Split Intein-Mediated DHFR Selectable Marker Enables Efficient Development of High-Producing CHO Cells for Therapeutic Proteins","authors":"Eun-Ji Lee, Chansik Yoon, Na-Yeong Heo, So-Hui Ryu, Gyun Min Lee, Yeon-Gu Kim","doi":"10.1021/acssynbio.6c00133","DOIUrl":"10.1021/acssynbio.6c00133","url":null,"abstract":"Thedihydrofolate reductase (DHFR)-based selection systemwidelyused to develop Chinese hamster ovary (CHO) cell lines for the productionof therapeutic proteins is limited by low selection stringency, relianceon stepwise gene amplification, and restricted capacity for efficientmultigene selection. In this study, we developed a highly stringentsplit DHFR selectable marker by integrating a computationally guidedsplit-site design with split intein-mediated protein ligation (SiMPl)in CHO cells. Using the synthetic SiMPl-DHFR selection system, theproportion of Fc-fusion glycoprotein-producing cells in stable cellpools reached 98.5%, and specific protein productivity increased 22.5-foldcompared with the wild-type DHFR (WT-DHFR) selection system. Whencombined with dhfr/methotrexate-mediated gene amplification,the SiMPl-DHFR system further increased mRNA expression levels andFc-fusion glycoprotein production. Notably, dhfr-nonamplifiedcell pools generated using the SiMPl-DHFR selection system exhibitedhigher expression levels than all dhfr-amplifiedcell pools established using the WT-DHFR selection system. Furthermore,the SiMPl-DHFR selection system permitted the efficient generationof stable monoclonal antibody- and bispecific antibody-producing CHOcell pools that require coordinated expression of two and four genes,respectively. Overall, this study establishes a synthetic selectablemarker architecture based on intein-mediated protein splicing thatrewires DHFR selection logic and provides a highly stringent and extensibleplatform for mammalian cell line development.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":"15 8","pages":"3249–3260"},"PeriodicalIF":4.5,"publicationDate":"2026-08-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148373711","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Discovery of Novel Glycosidase-Derived Cell-Penetrating Peptides Encoded by Human Gut Commensals 人类肠道共生体编码的新型糖苷酶衍生细胞穿透肽的发现。

IF 4.5 2区生物学

ACS Synthetic Biology Pub Date : 2026-08-21 Epub Date: 2026-07-09 DOI: 10.1021/acssynbio.6c00031

Sarah E. Post, Hannah S. Ceisler, Rohit G. Lal, Adarsh Singh, Maya A. Deen, Locke E. Bonomo, Lille M. Cunic, Ilana L. Brito

{"title":"Discovery of Novel Glycosidase-Derived Cell-Penetrating Peptides Encoded by Human Gut Commensals","authors":"Sarah E. Post, Hannah S. Ceisler, Rohit G. Lal, Adarsh Singh, Maya A. Deen, Locke E. Bonomo, Lille M. Cunic, Ilana L. Brito","doi":"10.1021/acssynbio.6c00031","DOIUrl":"10.1021/acssynbio.6c00031","url":null,"abstract":"Intracellular delivery of therapeutics remains a majorchallengefor modern medicine. To enhance intracellular uptake, therapeuticscan be delivered with carrier proteins possessing an inherent cell-penetratingactivity. There is an increasing need for new cell-penetrating carrierswith diverse biophysical properties and mechanisms of action to transporta wide range of therapeutic cargo. As many cell-penetrating proteinsand peptides derive from natural proteins, we sought to mine a previouslyunexplored community, the human gut microbiome, for cell-penetratingsequences. Here, we performed a high-throughput functional metagenomicscreen to identify cell-penetrating protein fragments from the humangut microbiome. We identified protein fragments encoded within glycosidaseenzymes from members of the Bacteroidetes phylum that mediate internalizationinto human cell lines when displayed on the surface of nonpathogenic,noninvasive Escherichia coli. We investigateone fragment, dubbed Gh_112, that adheres to human fibronectin, activatesmultiple endocytic pathways, and specifically promotes uptake of E. coli into multiple cancerous epithelial cell linesrather than healthy epithelial tissue in vitro. Overall,this work demonstrates that the human gut microbiome is a source ofcell-penetrating sequences and expands the known repertoire of cell-penetratingcarrier systems.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":"15 8","pages":"3179–3192"},"PeriodicalIF":4.5,"publicationDate":"2026-08-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148416923","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0