ACS Synthetic Biology最新文献_第10页

Establishing a High-Yield Chloroplast Cell-Free System for Prototyping Genetic Parts. 建立用于基因部件原型设计的高产无叶绿体细胞系统。

IF 3.7 2区生物学

ACS Synthetic Biology Pub Date : 2024-08-16 Epub Date: 2024-07-18 DOI: 10.1021/acssynbio.4c00111

Lauren Clark, Christopher A Voigt, Michael C Jewett

{"title":"Establishing a High-Yield Chloroplast Cell-Free System for Prototyping Genetic Parts.","authors":"Lauren Clark, Christopher A Voigt, Michael C Jewett","doi":"10.1021/acssynbio.4c00111","DOIUrl":"10.1021/acssynbio.4c00111","url":null,"abstract":"Plastid engineering offers the potential to carry multigene traits in plants; however, it requires reliable genetic parts to balance expression. The difficulty of chloroplast transformation and slow plant growth makes it challenging to build plants just to characterize genetic parts. To address these limitations, we developed a high-yield cell-free system from Nicotiana tabacum chloroplast extracts for prototyping genetic parts. Our cell-free system uses combined transcription and translation driven by T7 RNA polymerase and works with plasmid or linear template DNA. To develop our system, we optimized lysis, extract preparation procedures (e.g., runoff reaction, centrifugation, and dialysis), and the physiochemical reaction conditions. Our cell-free system can synthesize 34 ± 1 μg/mL luciferase in batch reactions and 60 ± 4 μg/mL in semicontinuous reactions. We apply our batch reaction system to test a library of 103 ribosome binding site (RBS) variants and rank them based on cell-free gene expression. We observe a 1300-fold dynamic range of luciferase expression when normalized by maximum mRNA expression, as assessed by the malachite green aptamer. We also find that the observed normalized gene expression in chloroplast extracts and the predictions made by the RBS Calculator are correlated. We anticipate that chloroplast cell-free systems will increase the speed and reliability of building genetic systems in plant chloroplasts for diverse applications.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":null,"pages":null},"PeriodicalIF":3.7,"publicationDate":"2024-08-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141631934","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

Balancing the AspC and AspA Pathways of Escherichia coli by Systematic Metabolic Engineering Strategy for High-Efficient l-Homoserine Production. 通过系统代谢工程策略平衡大肠杆菌的 AspC 和 AspA 途径以高效生产 l-高丝氨酸。

IF 3.7 2区生物学

ACS Synthetic Biology Pub Date : 2024-08-16 Epub Date: 2024-07-23 DOI: 10.1021/acssynbio.4c00208

Yuanyuan Chen, Lianggang Huang, Tao Yu, Yuan Yao, Mingming Zhao, Aiping Pang, Junping Zhou, Bo Zhang, Zhiqiang Liu, Yuguo Zheng

{"title":"Balancing the AspC and AspA Pathways of Escherichia coli by Systematic Metabolic Engineering Strategy for High-Efficient l-Homoserine Production.","authors":"Yuanyuan Chen, Lianggang Huang, Tao Yu, Yuan Yao, Mingming Zhao, Aiping Pang, Junping Zhou, Bo Zhang, Zhiqiang Liu, Yuguo Zheng","doi":"10.1021/acssynbio.4c00208","DOIUrl":"10.1021/acssynbio.4c00208","url":null,"abstract":"l-Homoserine is a promising C4 platform compound used in the agricultural, cosmetic, and pharmaceutical industries. Numerous works have been conducted to engineer Escherichia coli to be an excellent l-homoserine producer, but it is still unable to meet the industrial-scale demand. Herein, we successfully engineered a plasmid-free and noninducible E. coli strain with highly efficient l-homoserine production through balancing AspC and AspA synthesis pathways. First, an initial strain was constructed by increasing the accumulation of the precursor oxaloacetate and attenuating the organic acid synthesis pathway. To remodel the carbon flux toward l-aspartate, a balanced route prone to high yield based on TCA intensity regulation was designed. Subsequently, the main synthetic pathway and the cofactor system were strengthened to reinforce the l-homoserine synthesis. Ultimately, under two-stage DO control, strain HSY43 showed 125.07 g/L l-homoserine production in a 5 L fermenter in 60 h, with a yield of 0.62 g/g glucose and a productivity of 2.08 g/L/h. The titer, yield, and productivity surpassed the highest reported levels for plasmid-free strains in the literature. The strategies adopted in this study can be applied to the production of other l-aspartate family amino acids.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":null,"pages":null},"PeriodicalIF":3.7,"publicationDate":"2024-08-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141746731","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 a Novel Probiotic Toolkit in Escherichia coli Nissle 1917 for Sensing and Mitigating Gut Inflammatory Diseases. 在大肠杆菌 Nissle 1917 中设计新型益生菌工具包，用于感知和缓解肠道炎症性疾病。

IF 3.7 2区生物学

ACS Synthetic Biology Pub Date : 2024-08-16 Epub Date: 2024-08-08 DOI: 10.1021/acssynbio.4c00036

Nathalie Weibel, Martina Curcio, Atilla Schreiber, Gabriel Arriaga, Marine Mausy, Jana Mehdy, Lea Brüllmann, Andreas Meyer, Len Roth, Tamara Flury, Valerie Pecina, Kim Starlinger, Jan Dernič, Kenny Jungfer, Fabian Ackle, Jennifer Earp, Martin Hausmann, Martin Jinek, Gerhard Rogler, Cauã Antunes Westmann

{"title":"Engineering a Novel Probiotic Toolkit in Escherichia coli Nissle 1917 for Sensing and Mitigating Gut Inflammatory Diseases.","authors":"Nathalie Weibel, Martina Curcio, Atilla Schreiber, Gabriel Arriaga, Marine Mausy, Jana Mehdy, Lea Brüllmann, Andreas Meyer, Len Roth, Tamara Flury, Valerie Pecina, Kim Starlinger, Jan Dernič, Kenny Jungfer, Fabian Ackle, Jennifer Earp, Martin Hausmann, Martin Jinek, Gerhard Rogler, Cauã Antunes Westmann","doi":"10.1021/acssynbio.4c00036","DOIUrl":"10.1021/acssynbio.4c00036","url":null,"abstract":"Inflammatory bowel disease (IBD) is characterized by chronic intestinal inflammation with no cure and limited treatment options that often have systemic side effects. In this study, we developed a target-specific system to potentially treat IBD by engineering the probiotic bacterium Escherichia coli Nissle 1917 (EcN). Our modular system comprises three components: a transcription factor-based sensor (NorR) capable of detecting the inflammation biomarker nitric oxide (NO), a type 1 hemolysin secretion system, and a therapeutic cargo consisting of a library of humanized anti-TNFα nanobodies. Despite a reduction in sensitivity, our system demonstrated a concentration-dependent response to NO, successfully secreting functional nanobodies with binding affinities comparable to the commonly used drug Adalimumab, as confirmed by enzyme-linked immunosorbent assay and in vitro assays. This newly validated nanobody library expands EcN therapeutic capabilities. The adopted secretion system, also characterized for the first time in EcN, can be further adapted as a platform for screening and purifying proteins of interest. Additionally, we provided a mathematical framework to assess critical parameters in engineering probiotic systems, including the production and diffusion of relevant molecules, bacterial colonization rates, and particle interactions. This integrated approach expands the synthetic biology toolbox for EcN-based therapies, providing novel parts, circuits, and a model for tunable responses at inflammatory hotspots.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":null,"pages":null},"PeriodicalIF":3.7,"publicationDate":"2024-08-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11334186/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141900039","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

ShuffleAnalyzer: A Comprehensive Tool to Visualize DNA Shuffling. ShuffleAnalyzer：可视化 DNA 洗牌的综合工具

IF 3.7 2区生物学

ACS Synthetic Biology Pub Date : 2024-08-16 Epub Date: 2024-07-11 DOI: 10.1021/acssynbio.4c00251

Franz Schweiggert, Gregor Habeck, Patrick Most, Martin Busch, Jörg Schweiggert

{"title":"ShuffleAnalyzer: A Comprehensive Tool to Visualize DNA Shuffling.","authors":"Franz Schweiggert, Gregor Habeck, Patrick Most, Martin Busch, Jörg Schweiggert","doi":"10.1021/acssynbio.4c00251","DOIUrl":"10.1021/acssynbio.4c00251","url":null,"abstract":"DNA shuffling is a powerful technique for generating synthetic DNA via recombination of homologous parental sequences. Resulting chimeras are often incorporated into complex libraries for functionality screenings that identify novel variants with improved characteristics. To survey shuffling efficiency, subsequences of chimeras can be computationally assigned to their corresponding parental counterpart, yielding insight into frequency of recombination events, diversity of shuffling libraries and actual composition of final variants. Whereas tools for parental assignment exist, they do not provide direct visualization of the results, making the analysis time-consuming and cumbersome. Here we present ShuffleAnalyzer, a comprehensive, user-friendly, Python-based analysis tool that directly generates graphical outputs of parental assignments and is freely available under a BSD-3 license (https://github.com/joerg-swg/ShuffleAnalyzer/releases). Besides DNA shuffling, peptide insertions can be simultaneously analyzed and visualized, which makes ShuffleAnalyzer a highly valuable tool for integrated approaches often used in synthetic biology, such as AAV capsid engineering in gene therapy applications.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":null,"pages":null},"PeriodicalIF":3.7,"publicationDate":"2024-08-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141588903","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

Dual-Regulation in Peroxisome and Cytoplasm toward Efficient Limonene Biosynthesis with Rhodotorula toruloides. 过氧物酶体和细胞质中的双重调控实现了 Rhodotorula toruloides 的高效柠檬烯生物合成。

IF 3.7 2区生物学

ACS Synthetic Biology Pub Date : 2024-08-16 Epub Date: 2024-06-11 DOI: 10.1021/acssynbio.4c00306

Qidou Gao, Yaqi Dong, Ying Huang, Sasa Liu, Xiaochun Zheng, Yiming Ma, Qingsheng Qi, Xue Wang, Zongbao Kent Zhao, Xiaobing Yang

{"title":"Dual-Regulation in Peroxisome and Cytoplasm toward Efficient Limonene Biosynthesis with Rhodotorula toruloides.","authors":"Qidou Gao, Yaqi Dong, Ying Huang, Sasa Liu, Xiaochun Zheng, Yiming Ma, Qingsheng Qi, Xue Wang, Zongbao Kent Zhao, Xiaobing Yang","doi":"10.1021/acssynbio.4c00306","DOIUrl":"10.1021/acssynbio.4c00306","url":null,"abstract":"Rhodotorula toruloides is a potential workhorse for production of various value-added chemicals including terpenoids, oleo-chemicals, and enzymes from low-cost feedstocks. However, the limited genetic toolbox is hindering its metabolic engineering. In the present study, four type I and one novel type II peroxisomal targeting signal (PTS1/PTS2) were characterized and employed for limonene production for the first time in R. toruloides. The implant of the biosynthesis pathway into the peroxisome led to 111.5 mg/L limonene in a shake flask culture. The limonene titer was further boosted to 1.05 g/L upon dual-metabolic regulation in the cytoplasm and peroxisome, which included employing the acetoacetyl-CoA synthase NphT7, adding an additional copy of native ATP-dependent citrate lyase, etc. The final yield was 0.053 g/g glucose, which was the highest ever reported. The newly characterized PTSs should contribute to the expansion of genetic toolboxes forR. toruloides. The results demonstrated that R. toruloides could be explored for efficient production of terpenoids.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":null,"pages":null},"PeriodicalIF":3.7,"publicationDate":"2024-08-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141299339","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

Towards Self-regeneration: Exploring the Limits of Protein Synthesis in the Protein Synthesis Using Recombinant Elements (PURE) Cell-free Transcription-Translation System. 走向自我再生：利用重组元件（PURE）无细胞转录-翻译系统探索蛋白质合成的极限。

IF 3.7 2区生物学

ACS Synthetic Biology Pub Date : 2024-08-16 Epub Date: 2024-07-27 DOI: 10.1021/acssynbio.4c00304

Ragunathan B Ganesh, Sebastian J Maerkl

引用次数: 0

From De Novo to Xeno: Advancing Macromolecule Design beyond Proteins. 从 De Novo 到 Xeno：推进超越蛋白质的大分子设计。

IF 3.7 2区生物学

ACS Synthetic Biology Pub Date : 2024-08-16 DOI: 10.1021/acssynbio.4c00179

Tyler Stukenbroeker

引用次数: 0

The BioRECIPE Knowledge Representation Format. BioRECIPE 知识表示格式。

IF 3.7 2区生物学

ACS Synthetic Biology Pub Date : 2024-08-16 Epub Date: 2024-07-25 DOI: 10.1021/acssynbio.4c00096

Emilee Holtzapple, Gaoxiang Zhou, Haomiao Luo, Difei Tang, Niloofar Arazkhani, Casey Hansen, Cheryl A Telmer, Natasa Miskov-Zivanov

引用次数: 0

Toward Practical Applications of Engineered Living Materials with Advanced Fabrication Techniques. 利用先进制造技术实现工程活体材料的实际应用。

IF 3.7 2区生物学

ACS Synthetic Biology Pub Date : 2024-08-16 Epub Date: 2024-07-13 DOI: 10.1021/acssynbio.4c00259

Chenjing Lu, Yaying Huang, Jian Cui, Junhua Wu, Chunping Jiang, Xiaosong Gu, Yi Cao, Sheng Yin

{"title":"Toward Practical Applications of Engineered Living Materials with Advanced Fabrication Techniques.","authors":"Chenjing Lu, Yaying Huang, Jian Cui, Junhua Wu, Chunping Jiang, Xiaosong Gu, Yi Cao, Sheng Yin","doi":"10.1021/acssynbio.4c00259","DOIUrl":"10.1021/acssynbio.4c00259","url":null,"abstract":"Engineered Living Materials (ELMs) are materials composed of or incorporating living cells as essential functional units. These materials can be created using bottom-up approaches, where engineered cells spontaneously form well-defined aggregates. Alternatively, top-down methods employ advanced materials science techniques to integrate cells with various kinds of materials, creating hybrids where cells and materials are intricately combined. ELMs blend synthetic biology with materials science, allowing for dynamic responses to environmental stimuli such as stress, pH, humidity, temperature, and light. These materials exhibit unique \"living\" properties, including self-healing, self-replication, and environmental adaptability, making them highly suitable for a wide range of applications in medicine, environmental conservation, and manufacturing. Their inherent biocompatibility and ability to undergo genetic modifications allow for customized functionalities and prolonged sustainability. This review highlights the transformative impact of ELMs over recent decades, particularly in healthcare and environmental protection. We discuss current preparation methods, including the use of endogenous and exogenous scaffolds, living assembly, 3D bioprinting, and electrospinning. Emphasis is placed on ongoing research and technological advancements necessary to enhance the safety, functionality, and practical applicability of ELMs in real-world contexts.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":null,"pages":null},"PeriodicalIF":3.7,"publicationDate":"2024-08-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141602889","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

Heterologous Biosynthesis of Methanobactin from Methylocystis sp. Strain SB2 in Methylosinus trichosporium OB3b. 三孢甲烷菌株 SB2 在三孢甲烷菌 OB3b 中异源合成甲烷菌素。

IF 3.7 2区生物学

ACS Synthetic Biology Pub Date : 2024-08-16 Epub Date: 2024-08-07 DOI: 10.1021/acssynbio.4c00026

Peng Peng, Alan A DiSpirito, Braden J Lewis, Joel D Nott, Jeremy D Semrau

{"title":"Heterologous Biosynthesis of Methanobactin from Methylocystis sp. Strain SB2 in Methylosinus trichosporium OB3b.","authors":"Peng Peng, Alan A DiSpirito, Braden J Lewis, Joel D Nott, Jeremy D Semrau","doi":"10.1021/acssynbio.4c00026","DOIUrl":"10.1021/acssynbio.4c00026","url":null,"abstract":"Aerobic methanotrophs, or methane-consuming microbes, are strongly dependent on copper for their activity. To satisfy this requirement, some methanotrophs produce a copper-binding compound, or chalkophore, called methanobactin (MB). In addition to playing a critical role in methanotrophy, MB has also been shown to have great promise in treating copper-related human diseases, perhaps most significantly Wilson's disease. In this congenital disorder, copper builds up in the liver, leading to irreversible damage and, in severe cases, complete organ failure. Remarkably, MB has been shown to reverse such damage in animal models, and there is a great deal of interest in upscaling MB production for expanded clinical trials. Such efforts, however, are currently hampered as (1) the natural rate of MB production rate by methanotrophs is low, (2) the use of methane as a substrate for MB production is problematic as it is explosive in air, (3) there is limited understanding of the entire pathway of MB biosynthesis, and (4) the most attractive form of MB is produced by Methylocystis sp. strain SB2, a methanotroph that is genetically intractable. Herein, we report heterologous biosynthesis of MB from Methylocystis sp. strain SB2 in an alternative methanotroph, Methylosinus trichosporium OB3b, not only on methane but also on methanol. As a result, the strategy described herein not only facilitates enhanced MB production but also provides opportunities to construct various mutants to delineate the entire pathway of MB biosynthesis, as well as the creation of modified forms of MB that may have enhanced therapeutic value.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":null,"pages":null},"PeriodicalIF":3.7,"publicationDate":"2024-08-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141896041","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