ACS Synthetic Biology最新文献

Engineering Cell Fate with Adaptive Feedback Control. 基于自适应反馈控制的工程细胞命运。

IF 3.7 2区生物学

ACS Synthetic Biology Pub Date : 2025-07-23 DOI: 10.1021/acssynbio.5c00299

Frank Britto Bisso, Giulia Giordano, Christian Cuba Samaniego

{"title":"Engineering Cell Fate with Adaptive Feedback Control.","authors":"Frank Britto Bisso, Giulia Giordano, Christian Cuba Samaniego","doi":"10.1021/acssynbio.5c00299","DOIUrl":"https://doi.org/10.1021/acssynbio.5c00299","url":null,"abstract":"Engineering cell fate is fundamental to optimizing therapies based on stem cells, which are aimed at replacing cells in patients suffering from trauma or disease. By timely administering molecular regulators (e.g., transcription factors, RNAs, or small molecules) in a process that mimics in vivo embryonic development, stem cell differentiation can be guided toward a specific cell fate. However, scaling up these therapies is extremely challenging because such differentiation strategies often result in mixed cellular populations. While synthetic biology approaches have been proposed to increase the yield of desired cell types, designing gene circuits that effectively redirect cell fate decisions requires mechanistic insight into the dynamics of the endogenous regulatory networks that govern this type of decision-making. In this work, we present a biomolecular adaptive controller designed to favor a specific cell fate. The controller, whose topology is akin to that of an Incoherent Feedforward Loop (IFFL), requires minimal knowledge of the endogenous network as it exhibits adaptive, non-reference-based behavior. The synthetic circuit operates through a sequestration mechanism and a delay introduced by an intermediate species, producing an output that asymptotically approximates a discrete temporal derivative of its input if the sequestration rate is sufficiently fast. Allowing the controller to actuate over a target species involved in the decision-making process creates a tunable synthetic bias that favors the production of the desired species with minimal alteration to the overall equilibrium landscape of the endogenous network. Through theoretical and computational analysis, we provide design guidelines for the controller's optimal operation, evaluate its performance under parametric perturbations, and extend its applicability to various examples of common multistable systems in biology.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":" ","pages":""},"PeriodicalIF":3.7,"publicationDate":"2025-07-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144688321","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

Peroxisomal Compartmentalization of the Methylerythritol-4-phosphate Pathway Alleviates Cellular Stress and Enhances Geraniol Production in Saccharomyces cerevisiae. 甲基赤藓糖醇-4-磷酸途径的过氧化物酶体区隔化缓解了细胞应激并促进了酿酒酵母香叶醇的产生。

IF 3.7 2区生物学

ACS Synthetic Biology Pub Date : 2025-07-23 DOI: 10.1021/acssynbio.4c00830

Jerome R Lon, Xuemei Zhao, Gulkiz Mamatrixat, Zhoukang Zhuang, Zhehao Jin, Tao Yu, Jufang Wang, Hongting Tang

{"title":"Peroxisomal Compartmentalization of the Methylerythritol-4-phosphate Pathway Alleviates Cellular Stress and Enhances Geraniol Production in Saccharomyces cerevisiae.","authors":"Jerome R Lon, Xuemei Zhao, Gulkiz Mamatrixat, Zhoukang Zhuang, Zhehao Jin, Tao Yu, Jufang Wang, Hongting Tang","doi":"10.1021/acssynbio.4c00830","DOIUrl":"https://doi.org/10.1021/acssynbio.4c00830","url":null,"abstract":"Terpenoids are a diverse class of compounds with significant application potential. While prokaryotic bacteria synthesize terpenoids via the methylerythritol-4-phosphate (MEP) pathway, fungi utilize the mevalonate (MVA) pathway. The MVA pathway has been widely employed for efficient terpenoid production in bacteria such as Escherichia coli, but the MEP pathway performs poorly for biosynthesis in yeast. In this study, we constructed a compartmentalized MEP pathway to enhance monoterpenoid production in Saccharomyces cerevisiae. By introducing a geraniol synthase, we initially achieved the production of geraniol from glucose. Further effective incorporation of a cytosolic MEP pathway with nine enzymes increased geraniol production by 174.5%. However, this also significantly inhibited cell growth. Overexpression analysis revealed that flavodoxin and flavodoxin reductase were major contributors to growth inhibition, which could also be a factor limiting the application of the MEP pathway. To address these issues, we employed peroxisomal compartmentalization to isolate the MEP pathway from cytosolic metabolism. This strategy alleviated growth inhibition and improved geraniol production by 93.18% compared to that of cytosolic expression. Through additional metabolic engineering, we optimized peroxisomal geraniol production, achieving a yield of 30.64 mg/L. Our findings demonstrate the potential of compartmentalized MEP pathway expression as a viable approach for enhancing terpenoid biosynthesis in yeast, offering valuable insights for future metabolic engineering efforts.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":" ","pages":""},"PeriodicalIF":3.7,"publicationDate":"2025-07-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144697086","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

Systematic Mapping of Bacterial CRISPRa Systems for Synergistic Gene Activation Reveals Antagonistic Effects. 细菌CRISPRa系统的系统定位协同基因激活揭示拮抗作用。

IF 3.7 2区生物学

ACS Synthetic Biology Pub Date : 2025-07-22 DOI: 10.1021/acssynbio.5c00358

Cholpisit Kiattisewee, Ava V Karanjia, Ryan A L Cardiff, Kira E Olander, Pansa Leejareon, Sarah S Alvi, James M Carothers, Jesse G Zalatan

{"title":"Systematic Mapping of Bacterial CRISPRa Systems for Synergistic Gene Activation Reveals Antagonistic Effects.","authors":"Cholpisit Kiattisewee, Ava V Karanjia, Ryan A L Cardiff, Kira E Olander, Pansa Leejareon, Sarah S Alvi, James M Carothers, Jesse G Zalatan","doi":"10.1021/acssynbio.5c00358","DOIUrl":"https://doi.org/10.1021/acssynbio.5c00358","url":null,"abstract":"CRISPR gene activation (CRISPRa) tools have shown great promise for bacterial strain engineering but often require customization for each intended application. Our goal is to create generalizable CRISPRa tools that can overcome previous limitations of gene activation in bacteria. In eukaryotic cells, multiple activators can be combined for synergistic gene activation. To identify potential effectors for synergistic activation in bacteria, we systematically characterized bacterial activator proteins with a set of engineered synthetic promoters. We found that optimal target sites for different activators could vary by up to 200 bases in the region upstream of the transcription start site (TSS). These optimal target sites qualitatively matched previous reports for each activator, but the precise targeting rules varied between different promoters. By characterizing targeting rules in the same promoter context, we were able to test activator combinations with each effector positioned at its optimal target site. We did not find any activator combinations that produced synergistic activation, and we found that many combinations were antagonistic. This systematic investigation highlights fundamental mechanistic differences between bacterial and eukaryotic transcriptional activation systems and suggests that alternative strategies will be necessary for strong bacterial gene activation at arbitrary endogenous targets.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":" ","pages":""},"PeriodicalIF":3.7,"publicationDate":"2025-07-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144681665","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

Hydrogel-Immobilized Multienzyme Systems for Cell-Free Chemical Bioproduction. 用于无细胞化学生物生产的水凝胶固定化多酶系统。

IF 3.7 2区生物学

ACS Synthetic Biology Pub Date : 2025-07-21 DOI: 10.1021/acssynbio.5c00234

Widianti Sugianto, Ryan A L Cardiff, Claire Benstead, Gokce Altin-Yavuzarslan, Lilo Pozzo, Alshakim Nelson, James M Carothers

{"title":"Hydrogel-Immobilized Multienzyme Systems for Cell-Free Chemical Bioproduction.","authors":"Widianti Sugianto, Ryan A L Cardiff, Claire Benstead, Gokce Altin-Yavuzarslan, Lilo Pozzo, Alshakim Nelson, James M Carothers","doi":"10.1021/acssynbio.5c00234","DOIUrl":"https://doi.org/10.1021/acssynbio.5c00234","url":null,"abstract":"Cell-free gene expression systems derived from bacterial lysates enable the expression of biosynthetic pathways from inexpensive and easily prepared DNA templates. These systems hold great promise for modular and on-demand bioproduction of valuable small molecules in resource-limited settings but are constrained in their long-term stability, reusability, and deployability. In this work, we demonstrate that multiple cell-free expressed enzymes can be co-immobilized in biocompatible hydrogels made from poly(ethylene glycol) diacrylate (PEGDA) with added glycerol for enhanced gel integrity. Using small-angle X-ray scattering (SAXS), we show that the mesh size of PEGDA-glycerol hydrogels is comparable to the globular sizes of many proteins and enzymes, which could be used for protein entrapment. We found that the combination between entrapment and chemical ligation of the enzymes was effective to retain proteins. By employing a method for direct fluorescence measurement from hydrogels, we found that proteins can be retained in PEGDA-glycerol for at least a week. By separating the cell-free enzyme expression from the immobilization step, we successfully fabricated enzyme-laden hydrogels with three heterologous cell-free enzymes for the bioconversion of pyruvic acid to malic acid, an industrially valuable and versatile precursor chemical. Both heterologous and endogenous enzymes from the lysate remain functional in photo-cross-linked hydrogels and can be reused for multiple biocatalytic cycles. Moreover, we also found that the immobilized enzymes exhibit up to 1.6-fold higher activity and 2-fold longer lifetimes than free enzymes in liquid reactions. These results could advance the deployment of cell-free synthetic biology because they show that reusable, stable, and durable multienzyme systems can be created using readily available materials and fabrication techniques.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":" ","pages":""},"PeriodicalIF":3.7,"publicationDate":"2025-07-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144673316","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

Fedbatchdesigner: A User-Friendly Dashboard for Modeling and Optimizing Growth-Arrested Fed-Batch Processes. Fedbatchdesigner：一个用户友好的仪表板，用于建模和优化增长受阻的feed批处理过程。

IF 3.7 2区生物学

ACS Synthetic Biology Pub Date : 2025-07-21 DOI: 10.1021/acssynbio.5c00357

Andrea C Graf, Julian Libiseller-Egger, Mathias Gotsmy, Jürgen Zanghellini

{"title":"Fedbatchdesigner: A User-Friendly Dashboard for Modeling and Optimizing Growth-Arrested Fed-Batch Processes.","authors":"Andrea C Graf, Julian Libiseller-Egger, Mathias Gotsmy, Jürgen Zanghellini","doi":"10.1021/acssynbio.5c00357","DOIUrl":"https://doi.org/10.1021/acssynbio.5c00357","url":null,"abstract":"Optimizing fed-batch fermentation strategies is key to maximizing bioprocess efficiency. While mathematical modeling can aid process design, its complexity often limits accessibility for experimental scientists. We present FedBatchDesigner, a user-friendly web tool for optimizing fed-batch processes with a growth-arrested production stage. With minimal input requirements, FedBatchDesigner enables rapid exploration of a process's titer, rate, and yield (TRY) landscape for constant, linear, and exponential feeding strategies. Interactive visualizations allow users to assess trade-offs between productivity and titer, supporting rational decision-making without the need for extensive modeling expertise. We demonstrate FedBatchDesigner's utility via two case studies: synthesis of (i) l-valine with a microaerobic production stage in Escherichia coli and (ii) ethanol under nitrogen starvation in Saccharomyces cerevisiae. FedBatchDesigner is freely available at https://chemnettools.anc.univie.ac.at/FedBatchDesigner, with the source code provided at https://github.com/julibeg/FedBatchDesigner under the MIT license.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":" ","pages":""},"PeriodicalIF":3.7,"publicationDate":"2025-07-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144681664","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

IF 3.7 2区生物学

ACS Synthetic Biology Pub Date : 2025-07-18

Matthias Recktenwald, Ritankar Bhattacharya, Mohammed Mehdi Benmassaoud, James MacAulay, Varun M. Chauhan, Leah Davis, Evan Hutt, Peter A. Galie, Mary M. Staehle, Nichole M. Daringer, Robert J. Pantazes and Sebastián L. Vega*,