Synthetic and Systems Biotechnology最新文献

英文中文

Engineering transcriptional regulatory networks for improving second-generation fuel ethanol production in Saccharomyces cerevisiae 改造转录调控网络，提高酿酒酵母的第二代燃料乙醇产量

IF 4.4 2区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Synthetic and Systems Biotechnology

Pub Date : 2024-10-28 DOI: 10.1016/j.synbio.2024.10.006

Presently, Saccharomyces cerevisiae demonstrates proficient co-fermentation of glucose and xylose, marking a significant advancement in second-generation fuel ethanol production. However, the presence of high concentrations of inhibitors in industrial lignocellulose hydrolysates and post-glucose effect caused by glucose consumption hinders severely impedes yeast robustness and xylose utilization for ethanol fermentation. Even worse, the antagonism between xylose utilization ability and strain robustness was observed, which proposes a difficult challenge in the production of second-generation fuel ethanol by S. cerevisiae. This review introduces the effect of engineering transcriptional regulatory networks on enhancing xylose utilization, improving strain robustness, alleviating antagonism between xylose utilization and strain robustness, and reducing post-glucose effect. Additionally, we provide an outlook on the developmental trends in this field, offering insights into future directions for increasing the production of second-generation fuel ethanol in S. cerevisiae.

目前，酿酒酵母能熟练地同时发酵葡萄糖和木糖，这标志着第二代燃料乙醇生产取得了重大进展。然而，工业木质纤维素水解物中存在的高浓度抑制剂以及葡萄糖消耗引起的后葡萄糖效应严重阻碍了酵母的生长和利用木糖发酵乙醇。更为严重的是，木糖利用能力与菌株的稳健性之间存在拮抗作用，这为利用酿酒酵母生产第二代燃料乙醇提出了艰巨的挑战。本综述介绍了转录调控网络工程对提高木糖利用率、改善菌株稳健性、缓解木糖利用率与菌株稳健性之间的拮抗作用以及降低葡萄糖后效应的影响。此外，我们还对这一领域的发展趋势进行了展望，为提高 S. cerevisiae 第二代燃料乙醇产量的未来方向提供了见解。

引用次数: 0

Biosensors for the detection of flaviviruses: A review 用于检测黄病毒的生物传感器：综述

IF 4.4 2区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Synthetic and Systems Biotechnology

Pub Date : 2024-10-26 DOI: 10.1016/j.synbio.2024.10.005

Flaviviruses affect the lives of millions of people in endemic regions and also have the potential to impact non-endemic areas. Factors such as climate change, global warming, deforestation, and increased travel and trade are linked to the spread of flaviviruses into new habitats and host species. Given the absence of specific treatments and the limited availability of vaccines, it is imperative to understand the biology of flaviviruses and develop rapid and sensitive diagnostic tests. These measures are essential for preventing the transmission of these potentially life-threatening pathogens. Flavivirus infections are mainly diagnosed using conventional methods. However, these techniques present several drawbacks, including high expenses, time-consuming procedures, and the need for skilled professionals. The search for fast, easy-to-use, and affordable alternative techniques as a feasible solution for developing countries is leading to the search for new methods in the diagnosis of flaviviruses, such as biosensors.

This review provides a comprehensive overview of different biosensor detection strategies for flaviviruses and describes recent advances in diagnostic technologies. Finally, we explore their future prospects and potential applications in pathogen detection. This review serves as a valuable resource to understand advances in ongoing research into new biosensor-based diagnostic methods for flaviviruses.

黄病毒影响着流行地区数百万人的生活，也有可能影响非流行地区。气候变化、全球变暖、森林砍伐以及旅行和贸易的增加等因素都与黄病毒向新的栖息地和宿主物种传播有关。由于缺乏特效治疗方法，疫苗供应有限，因此必须了解黄病毒的生物学特性，并开发快速、灵敏的诊断检测方法。这些措施对于防止这些可能危及生命的病原体的传播至关重要。黄病毒感染主要采用传统方法进行诊断。然而，这些技术存在一些缺点，包括费用高、程序耗时以及需要熟练的专业人员。本综述全面概述了针对黄病毒的不同生物传感器检测策略，并介绍了诊断技术的最新进展。最后，我们探讨了它们在病原体检测中的未来前景和潜在应用。这篇综述是了解正在进行的基于生物传感器的黄病毒新诊断方法研究进展的宝贵资料。

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引用次数: 0

Systematic metabolic engineering of Yarrowia lipolytica for efficient production of phytohormone abscisic acid 利用脂肪分解酵母的系统代谢工程高效生产植物激素脱落酸

IF 4.4 2区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Synthetic and Systems Biotechnology

Pub Date : 2024-10-18 DOI: 10.1016/j.synbio.2024.10.004

Abscisic acid (ABA) is an important phytohormone with diverse applications. It currently relies on the fermentation of Botrytis cinerea, which suffers from limited availability of genetic engineering tools. Here, Yarrowia lipolytica was engineered to enable de novo biosynthesis of ABA. To overcome the rate-limiting P450 enzymes, systematic engineering strategies were implemented. Firstly, the dissolved oxygen was increased to boost the activity of P450 enzymes. Secondly, the expansion of endoplasmic reticulum was implemented to improve the functional expression of P450 enzymes. Lastly, rate-limiting enzymes were assembled to facilitate substrate trafficking. Moreover, ABA production was further improved by strengthening the mevalonate pathway. Finally, the engineered strain produced 1221.45 mg/L of ABA in a 5-L bioreactor. The study provides effective approaches for alleviating rate-limiting P450 enzymes to enhance ABA production and achieve competitive industrial-level ABA production in Y. lipolytica.

脱落酸（ABA）是一种重要的植物激素，具有多种用途。目前，它依赖于西尼瑞氏菌的发酵，而西尼瑞氏菌的发酵受制于有限的基因工程工具。在这里，我们改造了脂肪分解蓍草菌（Yarrowia lipolytica），使其能够从头开始生物合成 ABA。为了克服 P450 酶的速率限制，我们实施了系统工程策略。首先，增加溶解氧以提高 P450 酶的活性。其次，扩大内质网以提高 P450 酶的功能表达。最后，组装限速酶以促进底物的运输。此外，还通过加强甲羟戊酸途径进一步提高了 ABA 的产量。最后，工程菌株在 5 升生物反应器中产生了 1221.45 毫克/升的 ABA。这项研究提供了有效的方法来减轻限速 P450 酶的作用，从而提高脂溶性酵母菌的 ABA 产量，并实现具有竞争力的工业级 ABA 生产。

引用次数: 0

Enhancing substrate specificity of microbial transglutaminase for precise nanobody labeling 增强微生物转谷氨酰胺酶的底物特异性，实现精确的纳米抗体标记

IF 4.4 2区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Synthetic and Systems Biotechnology

Pub Date : 2024-10-16 DOI: 10.1016/j.synbio.2024.10.003

Streptomyces mobaraenesis transglutaminase (smTG) can be used for site-specific labeling of proteins with chemical groups. Here, we explored the use of modified smTG for the biosynthesis of nanobody-fluorophore conjugates (NFC). smTG catalyzes the conjugation of acyl donors containing glutamine with lysine-containing acceptors, which can lead to non-specific cross-linking. To achieve precise site-specific labeling, we employed molecular docking and virtual mutagenesis to redesign the enzyme's substrate specificity towards the peptide GGGGQR, a non-preferred acyl donor for smTG. Starting with a thermostable and highly active smTG variant (TGm2), we identified that single mutations G250H and Y278E significantly enhanced activity against GGGGQR, increasing it by 41 % and 1.13-fold, respectively. Notably, the Y278E mutation dramatically shifted the enzyme's substrate preference, with the activity ratio against GGGGQR versus the standard substrate CBZ-Gln-Gly rising from 0.05 to 0.93. In case studies, we used nanobodies 1C12 and 7D12 as labeling targets, catalyzing their conjugation with a synthetic fluorophore via smTG variants. Nanobodies fused with GGGGQR were successfully site-specifically labeled by TGm2-Y278E, in contrast to non-specific labeling observed with other variants. These results suggest that engineering smTG for site-specific labeling is a promising approach for the biosynthesis of antibody-drug conjugates.

链霉菌转谷氨酰胺酶（smTG）可用于用化学基团对蛋白质进行位点特异性标记。smTG 催化含谷氨酰胺的酰基供体与含赖氨酸的受体的连接，这可能导致非特异性交联。为了实现精确的位点特异性标记，我们采用了分子对接和虚拟诱变的方法，重新设计了酶对肽 GGGGQR（smTG 的非首选酰基供体）的底物特异性。从一个恒温高活性的 smTG 变体（TGm2）开始，我们发现单个突变 G250H 和 Y278E 显著增强了对 GGGQR 的活性，分别提高了 41% 和 1.13 倍。值得注意的是，Y278E 突变极大地改变了酶的底物偏好，对 GGGGQR 与标准底物 CBZ-Gln-Gly 的活性比从 0.05 上升到 0.93。在案例研究中，我们使用纳米抗体 1C12 和 7D12 作为标记目标，通过 smTG 变体催化它们与合成荧光团的连接。与其他变体的非特异性标记不同，与 GGGGQR 融合的纳米抗体成功地被 TGm2-Y278E 进行了位点特异性标记。这些结果表明，进行位点特异性标记的 smTG 工程是生物合成抗体-药物共轭物的一种可行方法。

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引用次数: 0

Mechanistic insights into the orthogonal functionality of an AHL-mediated quorum-sensing circuit in Yersinia pseudotuberculosis 对耶尔森氏菌假结核中 AHL 介导的法定量感应回路的正交功能的机理认识

IF 4.4 2区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Synthetic and Systems Biotechnology

Pub Date : 2024-10-14 DOI: 10.1016/j.synbio.2024.10.002

YpsR, a pivotal regulatory protein in the quorum-sensing (QS) of Yersinia pseudotuberculosis(Y. pstb), is essential for molecular signaling, yet its molecular mechanisms remain poorly understood. Herein, this study systematically investigates the interactions between YpsR and acyl-homoserine lactones (AHLs), shedding light on the selective mechanism of YpsR to various AHL molecules. Using molecular docking and surface plasmon resonance (SPR) analysis, we confirmed YpsR's binding affinities, with the strongest observed for 3OC6-HSL, which notably inhibited Y. pstb growth. Additionally, we engineered a whole-cell biosensor based on YpsR-AHL interaction, which exhibited sensitivity to the signal molecule 3OC6-HSL produced by Y. pstb. Furthermore, key YpsR residues (S32, Y50, W54, D67) involved in AHL binding were identified and validated. Overall, this research elucidates the mechanisms of QS signal recognition in Y. pstb, providing valuable insights that support the development of diagnostic tools for detecting Y. pstb infections.

YpsR是假结核耶尔森菌（Y. pseudotuberculosis，Y. pstb）法定量感应（QS）过程中的关键调控蛋白，对分子信号转导至关重要，但其分子机制仍鲜为人知。本研究系统研究了 YpsR 与酰基高丝氨酸内酯（AHLs）之间的相互作用，揭示了 YpsR 对各种 AHL 分子的选择性机制。通过分子对接和表面等离子体共振（SPR）分析，我们证实了 YpsR 的结合亲和力，其中 3OC6-HSL 的结合亲和力最强，能显著抑制 Y. pstb 的生长。此外，我们还设计了一种基于 YpsR-AHL 相互作用的全细胞生物传感器，它对 Y. pstb 产生的信号分子 3OC6-HSL 具有敏感性。此外，我们还鉴定并验证了参与 AHL 结合的关键 YpsR 残基（S32、Y50、W54、D67）。总之，这项研究阐明了 Y. pstb 的 QS 信号识别机制，为开发检测 Y. pstb 感染的诊断工具提供了有价值的见解。

引用次数: 0

Revolutionizing biotechnology advances natural products discovery and industrial processing 生物技术的革命性进步推动了天然产品的发现和工业加工

IF 4.4 2区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Synthetic and Systems Biotechnology

Pub Date : 2024-10-04 DOI: 10.1016/j.synbio.2024.10.001

引用次数: 0

CRISPR/Cas12a-based genome editing for cyanophage of Anabeana sp. 基于 CRISPR/Cas12a 的蓝藻噬菌体基因组编辑技术

IF 4.4 2区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Synthetic and Systems Biotechnology

Pub Date : 2024-10-03 DOI: 10.1016/j.synbio.2024.09.011

Efforts have been conducted on cyanobacterial genome editing, yet achieving genome editing in cyanophages remains challenging. Editing cyanophage genomes is crucial for understanding and manipulating their interactions with cyanobacterial hosts, offering potential solutions for controlling cyanobacterial blooms. In this study, we developed a streamlined CRISPR-Cas12a-based method for efficient cyanophage genome editing and then applied this method to the cyanophages A-1(L) and A-4(L) of Anabeana sp. PCC.7120. Multiple hypothetical genes were edited and knocked out from these two cyanophage genomes, generating viable mutants with varying capabilities to inhibit cyanobacterial growth. All these mutants displayed significant inhibitory effects on the host, indicating that these genes were non-essential for phage life cycle and the deletion led to little impairment of the cyanophages in infectious efficiency to their host. By iterative and simultaneous gene knockouts in cyanophage A-4(L), we achieved the minimal genome mutant with a 2400 bp reduction in genome size, representing a 5.75 % decrease compared to the wild type (WT). In conclusion, these cyanophage mutants can facilitate the identification of nonessential genes for cyanophages biology and the insertion of foreign genes for synthetic biology research. This advancement holds promise in addressing the widespread issue of water blooms and the associated environmental hazards.

人们已经在蓝藻基因组编辑方面做出了努力，但在噬菌体中实现基因组编辑仍然具有挑战性。编辑蓝藻噬菌体基因组对于理解和操纵它们与蓝藻宿主的相互作用至关重要，从而为控制蓝藻藻华提供潜在的解决方案。在本研究中，我们开发了一种基于 CRISPR-Cas12a 的简化方法，用于高效编辑蓝藻噬菌体基因组，并将该方法应用于 Anabeana sp. PCC.7120 的蓝藻噬菌体 A-1(L) 和 A-4(L)。这两种蓝藻噬菌体基因组中的多个假定基因被编辑和敲除，产生了具有不同抑制蓝藻生长能力的可行突变体。所有这些突变体都对宿主有明显的抑制作用，表明这些基因对噬菌体的生命周期并不重要，而且基因缺失对蓝藻噬菌体感染宿主的效率几乎没有影响。通过在蓝藻噬菌体 A-4(L)中反复同时进行基因敲除，我们获得了基因组最小的突变体，其基因组大小减少了 2400 bp，与野生型（WT）相比减少了 5.75%。总之，这些蓝藻噬菌体突变体有助于鉴定蓝藻噬菌体生物学中的非必要基因，以及为合成生物学研究插入外来基因。这一进展有望解决普遍存在的水华问题及相关的环境危害。

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引用次数: 0

Developing polycistronic expression tool in Yarrowia lipolytica 在脂溶性亚罗酵母中开发多聚组蛋白表达工具

IF 4.4 2区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Synthetic and Systems Biotechnology

Pub Date : 2024-09-28 DOI: 10.1016/j.synbio.2024.09.010

Unconventional oleaginous yeast Yarrowia lipolytica has gained widespread applications as a microbial cell factory for synthesizing various chemicals and natural products. The construction of efficient cell factories requires intricate metabolic engineering. However, multi-gene expression in Y. lipolytica is labor-intensive. To facilitate multi-gene expression, we developed the polycistronic expression tool using 2A peptides. We first compared different 2A peptides in Y. lipolytica and identified two 2A peptides with high cleavage efficiency: P2A and ERBV-1. The effect of 2A peptides on the expression level of upstream and downstream genes was then determined. Ultimately, we applied the identified 2A peptides to express four genes in canthaxanthin biosynthetic pathway within one expression cassette for canthaxanthin production. This study enriches the multi-gene expression tools of Y. lipolytica, which will facilitate the cell factory construction of Y. lipolytica.

非常规油脂酵母亚罗酵母（Yarrowia lipolytica）作为合成各种化学品和天然产品的微生物细胞工厂，已获得广泛应用。构建高效细胞工厂需要复杂的代谢工程。然而，在脂溶性酵母中进行多基因表达是一项劳动密集型工作。为了促进多基因表达，我们利用 2A 肽开发了多聚组表达工具。我们首先比较了脂溶性酵母中不同的 2A 肽，确定了两种裂解效率较高的 2A 肽：P2A和ERBV-1。然后确定了 2A 肽对上游和下游基因表达水平的影响。最终，我们利用所鉴定的 2A 肽在一个表达盒中表达了角黄素生物合成途径中的四个基因，以生产角黄素。这项研究丰富了溶脂酵母的多基因表达工具，有助于溶脂酵母细胞工厂的构建。

引用次数: 0

Synthetic biotechnology for C1 bio-refinery 用于 C1 生物精炼厂的合成生物技术

IF 4.4 2区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Synthetic and Systems Biotechnology

Pub Date : 2024-09-23 DOI: 10.1016/j.synbio.2024.09.009

引用次数: 0

Transcriptome analysis reveals methanol metabolism variations for the growth damage caused by overexpression of chimeric transactivators in Pichia pastoris 转录组分析揭示了甲醇代谢变化对过量表达嵌合转录因子的 Pichia pastoris 生长造成的损害

IF 4.4 2区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Synthetic and Systems Biotechnology

Pub Date : 2024-09-23 DOI: 10.1016/j.synbio.2024.09.008

Methanol is a promising substrate for sustainable biomanufacturing, and Pichia pastoris has become a commonly used yeast for methanol utilization due to its powerful methanol metabolic pathways and methanol inducible promoter. Previous reconstruction of gene circuits highly improved transcriptional activity, but excessive expression of chimeric transactivator damaged cell growth on methanol. Here we employed transcriptome analysis to investigate the effects of chimeric transactivator overexpression on cellular metabolism and regulatory networks. The results showed that strong expression of chimeric transactivator unexpectedly downregulated methanol metabolism, especially the alcohol oxidase 1 (AOX1), but without remarkable changes in expression of transcriptional factors. Meanwhile, the synthesis of peroxisomes also varied with chimeric transactivator expression. In addition, the enrichment analysis of differentially expressed genes revealed their impact on cellular metabolism. The gene expression patterns caused by different expression levels of chimeric transactivators have also been clarified. This work provides useful information to understand the transcriptional regulation of the AOX1 promoter and methanol signaling. It revealed the importance of balancing transcription factor expression for the host improvement.

甲醇是一种很有前景的可持续生物制造底物，而 Pichia pastoris 因其强大的甲醇代谢途径和甲醇诱导启动子而成为甲醇利用的常用酵母。以前的基因回路重建大大提高了转录活性，但嵌合转录因子的过度表达会破坏细胞在甲醇中的生长。在此，我们利用转录组分析研究了嵌合型转录激活因子过度表达对细胞代谢和调控网络的影响。结果表明，嵌合型转录因子的强表达意外地下调了甲醇代谢，尤其是醇氧化酶1（AOX1），但转录因子的表达没有明显变化。同时，过氧化物酶体的合成也随嵌合转座子的表达而变化。此外，差异表达基因的富集分析显示了它们对细胞代谢的影响。此外，还阐明了嵌合转座因子不同表达水平所导致的基因表达模式。这项工作为了解 AOX1 启动子的转录调控和甲醇信号转导提供了有用的信息。它揭示了平衡转录因子表达对宿主改良的重要性。

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