 Your new post is loading...
|
Scooped by
?
Today, 1:26 PM
|
Although the α-amylase (AmyS) from Geobacillus stearothermophilus exhibits high thermostability, its low enzymatic activity (44.36 ± 2.02 U/mL) severely hinders industrial applications. Given its strong protein secretion capability and GRAS (generally recognized as safe) status, this study employed Bacillus subtilis as the host to enhance AmyS production. Through error-prone PCR and high-throughput screening, a triple mutant T151A/K178E/T458A (AmySM) was generated, showing a 17.67% increase in activity (51.34 ± 1.11 U/mL). AmySM activity was further increased by 29.32% to 65.23 ± 2.33 U/mL using the signal peptide SPykwD, selected from a comprehensive library for superior secretion efficiency. The promoter PgsiB enhanced activity by 37.61% to 89.54 ± 2.95 U/mL. Optimizing the ribosome binding site (RBS) resulted in an additional 48.83% increase in activity, yielding a final activity of 134.02 ± 3.54 U/mL, which corresponds to a 3.02-fold improvement over the initial strain WBSW (44.36 ± 2.02 U/mL). Ultimately, scale-up fermentation in a 5-L bioreactor yielded a maximum extracellular activity of 1244.17 ± 48.66 U/mL at 72 hours, a 2.84-fold increase over the control. This multi-level strategy provides a rational framework for high-efficiency AmySM production, paving the way for extracellular production of high-value proteins in the GRAS host B. subtilis.
|
|
Scooped by
?
Today, 12:45 PM
|
Recent advances in AI have substantially accelerated in silico drug discovery, yet most computational approaches remain task-specific and require expert knowledge. We present BioChemAIgent, an agentic framework that orchestrates state-of-the-art AI models and established computational chemistry tools to support end-to-end small-molecule analysis, protein modeling, molecular docking, and interaction analysis through a unified interface. BioChemAIgent emphasizes transparent reasoning, reproducible workflows, and community-oriented extensibility for structural biology and drug discovery applications.
|
|
Scooped by
?
Today, 11:23 AM
|
Predicting the species composition of microbial communities is a problem that has proven surprisingly difficult, including in fully controlled microbial systems in a lab setting. Few organisms are able to establish themselves in an community with no others, even if nutrients are provided. At the same time some species are observed to be unable to stably coexist with certain other species. We here present a model attempting to reduce the dynamics of community assembly to its most basic components, while preserving its salient features. The model deals not with abundances or densities of bacterial populations or resources, but only with their presence or absence. Similarly, only three types of discrete relationships between species are considered: species excluding each other, and mutualistic dependence on and production of nutrients in the form of exometabolites. Despite these simplifications, the model system still exhibits rich dynamics, including extinction cascades and emerging stability against invasion. We derive conditions for these to occur and compare our results with existing knowledge of microbial communities. Our results provide a novel approach for the theoretical study of microbial communities and their stability.
|
|
Scooped by
?
December 19, 5:04 PM
|
Remarkable progress has been achieved by machine learning, particularly in accurate prediction of protein tertiary structures. Despite these advances, accurately annotating protein functions through machine learning approaches remains challenging, primarily due to the limited availability of large-scale benchmarking data. In this study, we addressed this gap by systematically screening proteins from the UniProt database for functional annotations, resulting in the creation of a benchmarking dataset that includes protein sequences and their corresponding annotations. The Protein Annotation Dataset (PAD) is a resource available to train a wide range of machine learning models for assignment of function annotations to previously unlabeled proteins. We curated a comprehensive dataset comprising four categories of functional annotations using enzyme commission (EC) numbers and gene ontology (GO) terms. The dataset was subsequently partitioned into training, validation, and test subsets. Furthermore, we incorporated an independent set from 12 diverse species, enabling the development and evaluation of innovative machine learning models.
|
|
Scooped by
?
December 19, 4:50 PM
|
Bradyrhizobium and Sinorhizobium are dominant soybean microsymbionts in acidic/neutral and alkaline soils, respectively. However, the molecular mechanisms underlying this pH-dependent adaptation remain elusive. In this study, phylogenomic analysis of 286 Bradyrhizobium and 322 Sinorhizobium genomes revealed that Bradyrhizobium possesses abundant xeno-siderophore receptors but has limited siderophore biosynthesis functions. In contrast, gene clusters directing siderophore biosynthesis are enriched in Sinorhizobium. As siderophores can chelate the prevalent insoluble Fe3+ under neutral and alkaline conditions, whereas being less important in acidic environments where soluble Fe2+ is readily accessible, we hypothesized that the genus-dependent phyletic distribution of siderophore biosynthesis and exploitation functions may contribute to the pH adaptation of these two genera. Indeed, Bradyrhizobium species barely grow under iron-limiting conditions, and this growth defect can be rescued by xeno-siderophores produced by Sinorhizobium. Using a xeno-siderophore-exploiting Bradyrhizobium diazoefficiens strain, an engineered xeno-siderophore exploiter, and an altruistic siderophore-producing strain derived from Sinorhizobium fredii, we revealed the competitive advantage of xeno-siderophore exploitation during soybean nodulation. Heterologous expression of certain Bradyrhizobium xeno-siderophore receptors, along with various adaptive mutations in the genome of the S. fredii receptor-lacking mutant, allowed this mutant to rapidly restore growth under iron-limiting conditions. These adaptive events in experimental evolution depend on the siderophore biosynthetic function of S. fredii. Taken together, these findings suggest that the siderophore utilization ability of soybean rhizobia can be positively selected under iron-limiting conditions: by maintaining abundant xeno-siderophore receptors in acid-tolerant Bradyrhizobium and by the rapid adaptive evolution of utilization machinery for self-produced siderophores in alkaline-tolerant Sinorhizobium.
|
|
Scooped by
?
December 19, 4:35 PM
|
Fungal plant biomass conversion (FPBC) is of great importance to the global carbon cycle and has been increasingly applied for the production of biofuel and biochemicals from lignocellulose. However, the comprehensive understanding of relevant molecular mechanisms in different fungi remains challenging. Here, we comparatively analyzed the transcriptome, proteome and metabolome profile of four ascomycetes and one basidiomycete fungi during their growth on two common agricultural feedstocks (soybean hulls and corn stover). We revealed strong time-, substrate- and species-specific responses at multi-omics levels for the tested fungi, highlighting species-specific carbon utilization approaches and evolutionary adaptation to environmental niches. Notably, a remarkable expressional diversity of lignocellulose degrading enzymes, sugar transporter and metabolic genes, as well as industrially relevant metabolites were identified across different fungi and cultivation conditions. The findings improves our understanding of complex molecular networks underlying FPBC and fungal ecological roles, offering novel insights that can guide future genetic engineering of fungi for valorization of agriculture waste into value-added bioproducts.
|
|
Scooped by
?
December 19, 4:32 PM
|
Violacein is a natural purple secondary metabolite with a wide range of biological activities including antibacterial, anticancer, antioxidant, and antiparasitic properties, rendering it a highly promising candidate for applications in medicine, agriculture, and food industries. Despite its availability from natural sources, a profound understanding of its production mechanisms has long been lacking. High-level production of violacein has been achieved through integrated strategies, including heterologous expression of its biosynthetic pathway in recombinant strains, enhancement of tryptophan precursor supply, and optimization of fermentation conditions. These approaches offer a flexible and scalable platform for violacein biosynthesis. Furthermore, recent efforts have focused on utilizing agro-industrial waste as a cost-effective and sustainable feedstock to further improve production efficiency and environmental compatibility. This review provides a comprehensive overview of the latest advancements in violacein production, examines the challenges associated with its application, and proposes strategies for optimizing gene expression, refining fermentation protocols, and utilizing low-cost raw materials to facilitate the efficient and sustainable violacein production.
|
|
Scooped by
?
December 19, 4:27 PM
|
Lignocellulose-derived fuels and chemicals are vital to breaking the world's dependence on fossil fuels. Though plant biomass is notoriously resistant to deconstruction, lignocellulolytic thermophiles are especially adept at degrading its constituent polysaccharides into mono- and oligo-saccharides for catabolism. And many thermophiles, whether lignocellulolytic or not, can be engineered to ferment lignocellulose-derived sugars into valuable fuels and chemicals as part of consolidated bioprocesses. Although the past 20 years have seen major advances in the genetic and metabolic engineering of individual thermophiles, the strategy of co-culturing thermophilic strains as part of synthetic communities has not been well established. Synthetic communities unlock synergistic interactions that outperform monocultures, thereby enhancing product titers, rates, and yields. While limited genetic tools once hindered the development of synthetic thermophilic communities, recent advances now offer robust systems for engineering these industrially relevant organisms. Here, we propose that this expanded genetic malleability enables engineering of 1) transport specialization to reduce substrate competition between strains and 2) division of labor strategies whereby one strain focuses on lignocellulose deconstruction while another strain dedicates metabolic burden for product synthesis. We draw on examples of engineered thermophiles like Clostridium thermocellum, Thermoanaerobacter saccharolyticum, and Anaerocellum bescii to illustrate how these mechanisms have been applied in thermophilic co-cultures. In brief, this perspective outlines design principles to construct effective thermophilic communities for lignocellulose bioprocessing.
|
|
Scooped by
?
December 19, 4:20 PM
|
Throughout thousands of years, the yeast Saccharomyces cerevisiae has been acting as a cell factory in food production. Lately, it has further functioned as a platform cell factory for the production of a multitude of different compounds, spanning from high-volume fuels to high-value pharmaceuticals. The past decade has witnessed the fact that the innovative tools in synthetic biology have driven the rapid progress of the yeast cell factory, enabling us to edit the genetic systems of organisms efficiently and "reprogram" elements or systems such as genes, circuits, pathways, and networks of organisms. Here, we will offer a brief review that highlights the most recent significant advances and perspectives regarding the innovative tools in yeast synthetic biology. These tools encompass genome editing tools, computational tools, adaptive laboratory evolution, and the standardization of biological DNA parts, with the intention of providing a practical guide for the implementation of novel, effective, and efficient development of the customized yeast cell factory.
|
|
Scooped by
?
December 19, 4:13 PM
|
Class 1 CRISPR–Cas systems utilize multi-subunit effector ribonucleoprotein complexes to identify and target DNA. Upon recognition, type I systems recruit the helicase/nuclease Cas3 for DNA degradation, while type IV-A systems use the helicase CasDinG for transcriptional repression. Here, we developed two recombinant class 1 CRISPR–Cas genome editing tools for inducing large genomic deletions: the compact type I-Fv (also termed I-F2) system from Shewanella putrefaciens and the type IV-A1 system from Pseudomonas oleovorans. In the latter, CasDinG was engineered to include a C-terminal HNH nuclease domain, conferring DNA cleavage activity and enabling analysis of CasDinG processivity. Whole-genome sequencing of Escherichia coli BL21-AI was used to monitor genome reduction and DNA repair mechanisms in response to CRISPR–Cas-induced damage. Small deletions were flanked by microhomologies, consistent with repair via alternative end joining, whereas deletions larger than 10 kb consistently terminated at nearby IS1 elements, implicating these sequences in the repair process. This study introduces compact type I and engineered type IV-A genome editing tools with distinct protospacer-adjacent motif requirements and provides new insights into CasDinG evolution and the DNA repair pathways engaged during CRISPR–Cas-mediated genome editing.
|
|
Scooped by
?
December 19, 2:00 AM
|
Rhizosphere microorganisms are known to be able to modify the plant's ability to resist abiotic stresses. It is, however, difficult to modify microbial communities to improve plant phenotypes. Here we tested if a rhizosphere microbial community from a water-stress naive soil could be modified by adding DNA extracted from soils with a water stress history. Six-week-old wheat plants growing under low or high-water availabilities were inoculated with DNA extracted from soils with contrasting long-term histories of water availability - one continuously and the other intermittently exposed to water deficit. The fate of the inoculated DNA in the rhizosphere microbial communities was assessed by shotgun metagenomics. Putatively transferred inoculum genes were disproportionately found in the Acidobacteria and Bacteroidetes and belonged to functional category such as antibiotics, biofilm, and carboxylates metabolism, among others. These functional categories were shared by pre-inoculation laterally transferred genes in the recipient soil, highlighting their usefulness for life in soil. The "continuous" inoculum reduced the stress levels of wheat under reduced soil water content, suggesting that the natural genetic transformation of the rhizosphere community can feedback to the plant. Altogether, we are providing evidence for an ecological mechanism that could be harnessed to modify plant-associated microbial communities and help plants sustain water stress.
|
|
Scooped by
?
December 19, 1:56 AM
|
Magnetotactic bacteria (MTB) are capable of accumulating and storing intercellular pools containing phosphorus, sulfur, nitrogen and carbon. Yet, the metabolic pathways connected with their intracellular storage of polyphosphate, elemental sulfur, nitrate, and calcium carbonate, and the environmental influence on MTB inclusions and their quantitative contribution to sedimentary chemical budgets remain underexplored. Using a combination of chemical characterization, ultrastructural and compositional analyses, and phylogenetic and genomic insights into microbial assemblages, we investigated the influence of geochemical parameters on the diversity, ecophysiology, and distribution of MTB in freshwater and brackish sediments. Special focus is given to intracellular inclusions and their metabolic pathways to uncover functional traits and ecological roles in elemental cycling. Of the 118 examined MTB cells, polyphosphate granules (75%) and nitrate-containing vacuoles (67%) were the most common inclusions followed by sulfur globules (25%) and calcium carbonate granules (8%). The intracellular phosphorus, nitrogen, and sulfur stored in MTB cells were conservatively estimated to account for 0.60%, 0.91%, and 0.23% of the total sedimentary P, N, and S in the investigated freshwater sediments, respectively. The ubiquitous nature and important ecological role of MTB can be explained by their ability to sequester chemical elements into intracellular inclusions, giving them a metabolic advantage in dynamic chemically stratified environments. The extraordinary phylogenetic diversity of MTB, coupled with their capability to hyperaccumulate and store a wide range of elements and compounds, represents a significant resource for biotechnology innovation.
|
|
Scooped by
?
December 19, 1:45 AM
|
The Gene Ontology (GO) knowledgebase https://geneontology.org is a comprehensive resource describing the functions of genes. The GO knowledgebase is regularly updated and improved. We describe here the major updates that have been made in the past 3 years. The ontology and annotations have been expanded and revised, particularly in several areas of biology: cellular metabolism, multi-organism interactions (e.g. host-pathogen), extracellular matrix proteins, chromatin remodeling (e.g. the “histone code”), and noncoding RNA functions. We have released version 2 of a comprehensive set of integrated, reviewed annotations for human genes, which we call the “functionome.” We have also dramatically increased the number of GO-CAM models, with over 1500 models of metabolic and signaling pathways, primarily in human, mouse, budding and fission yeast, and fruit fly. Finally, we discuss our current recommendations and future prospects of AI in the use and development of GO.
|
|
|
Scooped by
?
Today, 12:54 PM
|
Driven by the persistence of microplastics and an overdependence on non-renewable sources, mycelium materials have emerged as alternatives to traditional materials, owing to their sustainable production and versatility. Engineered living materials, composite material systems incorporating biological components to enable function, possess desirable regenerative properties but have yet to be fully applied due to their lack of strength versus synthetic or natural materials. We leverage two strategies used by nature to generate mechanical strength: structural hierarchy and biomineralization. We extrude a bioink composed of alginate and a modified strain of the fungus Aspergillus niger capable of silica mineralization, and modulate the morphology by the growth and processing conditions. Mineralization results in significantly stronger and stiffer dried fibers. We demonstrate their potential as textile materials through twisting and braiding to significantly increase the fracture strain. Our results show that mycelium morphology and mechanics can be tuned through mineralization, growth conditions, and processing.
|
|
Scooped by
?
Today, 11:29 AM
|
Bacterial extracellular vesicles (BEVs) are known to enhance infection susceptibility in vivo, yet the mechanistic basis for this remote preconditioning of host cells is unknown. Here, we discover an evolutionarily conserved, lipid-driven physical mechanism by which pathogenic bacterial EVs systemically arrest phagosome maturation in bystander host cells. Using live-cell fluorescence lifetime imaging, in vitro reconstitution and micromanipulation we show that EVs from diverse pathogens - Mycobacterium tuberculosis, Klebsiella pneumoniae, and Staphylococcus aureus - fuse with host plasma, phagosomal, and lysosomal membranes. This fusion increases membrane tension and perturbs early phagosomal maturation. Transcriptomic profiling confirms a broad downregulation of phagosome maturation genes while upregulation of lysosomal stress responsive genes. Crucially, in vitro reconstitution shows that EVs, and their purified lipids alone, are sufficient to induce phase separation and increase membrane tension, directly inhibiting phago-lysosomal fusion. Our findings establish a paradigm in which pathogens exploit EVs not merely as delivery vehicles, but as tools to remotely rewire host cell membrane mechanics to hijack phagosome maturation and host defense - a strategy that moves beyond canonical effector-based models of pathogenesis.
|
|
Scooped by
?
Today, 11:13 AM
|
DNA-based information storage offers a promising alternative to conventional media due to its high density, long-term stability, and low energy requirements. However, its application remains hindered by synthesis costs, limited sequence length and poor scalability. DNA polymerase is a critical enzymatic tool in the DNA storage systems by enabling high-fidelity data writing and targeted sequence amplification. In this study, we engineered chimeric DNA polymerases by fusing the high-fidelity 9°N DNA polymerase with double-stranded DNA binding proteins derived from thermophilic archaea. These fusions significantly enhanced processivity, thermal stability, and salt tolerance by stabilizing enzyme-template interactions, mimicking sliding clamps while preserving catalytic efficiency. Leveraging these properties, we demonstrated precise file retrieval from a mixed oligonucleotide pool using orthogonal barcode primers. Compared with wild-type 9°N, the chimeric polymerases, particularly PLS, exhibited reduced substitution error rates and improved read accuracy. We then applied these enzymes to a DNA movable type storage system, where prefabricated DNA modules were assembled into encoding blocks. Using engineered polymerases, these blocks were recombined to enable flexible data rewriting without de novo DNA synthesis. Moreover, a multi-enzyme assembly strategy enabled the construction of kilobase-scale DNA sequences encoding a classical Chinese poem, achieving complete data recovery. All assembled fragments remained stable in E. coli over 100 generations, exhibiting the potential for in vivo storage. Collectively, our findings demonstrated the role of engineered DNA polymerases for DNA-based information storage. Moreover, this system reduced synthesis demands, supported scalable rewriting, and ensured long-term preservation, offering a practical route to sustainable, high-fidelity DNA data storage.
|
|
Scooped by
?
December 19, 4:57 PM
|
Cyclic disulfide-rich peptides have become increasingly popular in drug development because their structures enhance molecular stability and allow for mutagenesis to introduce non-native functions. This review focuses on yeast-based platform technologies and their utility in advancing cyclic disulfide-rich peptides as drug modalities and for large-scale biomanufacturing. These technologies include yeast surface display which facilitates the screening of large libraries to develop peptide binders with strong affinity and selectivity for protein targets, while maintaining the innate high stability of the peptide scaffold via protease-based selection pressure. We also describe a recently developed platform that leverages yeast’s ability to secrete correctly folded disulfide-rich peptides while simultaneously displaying peptide or protein tags on their surfaces. In combination with microfluidics technology, the platform creates single-cell yeast-in-droplets reactors, enabling the screening of large libraries based on functional output rather than solely on binding affinity. After identifying cyclic peptide candidates through library-based discovery, these candidates can be produced using a versatile yeast-based bioproduction platform. Traditionally, cyclic disulfide-rich peptides are produced through solid-phase synthesis, a method that generates significant amounts of toxic waste. In contrast, yeast-based bioproduction offers an environmentally sustainable alternative. It has the capability to produce structurally distinct peptides with minimal adjustments and is easily scalable using microbial fermenters, making it an ideal choice for large-scale production.
|
|
Scooped by
?
December 19, 4:41 PM
|
Tissue- or cell-type-specific expression of transgenes is often essential for interrogation of the biological phenomenon or predictable engineering of multicellular organisms but can be stymied by cryptic enhancers that make identification of promoters that generate desired expression profiles challenging. In plants, the months-to-years long timeline associated with prototyping putative tissue-specific promoters in transgenic lines deepens this challenge. We have developed a novel strategy called Ribozyme-Enabled Tissue Specificity (RETS) that leverages the knowledge of where and when genes are expressed derived from transcriptomic studies to enable tissue-specific expression without needing characterized promoters. It uses a split self-splicing ribozyme based on a group I intron from Tetrahymena thermophila to enable the conditional reconstitution of a transgene mRNA in the presence of a secondary tissue-specific mRNA of choice. We elucidate the design features that enable flexible swapping of transgenes and targets, enhancing transgene expression, and circumventing host RNA interference responses. We then show that these innovations enable tissue-specific and dose-dependent expression of transgenes in Arabidopsis thaliana. Finally, we demonstrate the utility of RETS both for creating genetically encoded biosensors to study the spatiotemporal patterns of gene expression in planta and for engineering tissue-specific changes in organ size. RETS provides a novel avenue to study expression patterns of native loci with nondestructive imaging, complementing the weakness of existing approaches. Additionally, the spatiotemporal control of transgene expression afforded by RETS enables precision engineering of plant phenotypes, which will facilitate enhancing crops without the trade-offs associated with constitutive expression.
|
|
Scooped by
?
December 19, 4:33 PM
|
A great number of multifactorial diseases, including neoplastic, metabolic, and autoimmune diseases, have been associated with microbiota dysbiosis. Recently, there has been an increasing understanding of the importance of microbiome and their impact on human health. Advances in synthetic biology have led to the development of probiotics as diagnostic tools and disease treatment approaches. In this review, we briefly summarize recent examples of engineered probiotic-based therapeutics in human diseases, including cancers, gastrointestinal disorders, infectious diseases, and metabolic disorders. Finally, we discuss the challenges and opportunities in developing engineered probiotics for disease treatments.
|
|
Scooped by
?
December 19, 4:30 PM
|
Plant-microbe interactions are critical to ecosystem resilience and substantially influence crop production. From the perspective of plant science, two important focus areas concerning plant-microbe interactions include: 1) understanding plant molecular mechanisms involved in plant-microbe interfaces and 2) engineering plants for increasing plant disease resistance or enhancing beneficial interactions with microbes to increase their resilience to biotic and abiotic stress conditions. Molecular biology and genetics approaches have been used to investigate the molecular mechanisms underlying plant responses to various beneficial and pathogenic microbes. While these approaches are valuable for elucidating the functions of individual genes and pathways, they fall short of unraveling the complex cross-talk across pathways or systems that plants employ to respond and adapt to environmental stresses. Also, genetic engineering of plants to increase disease resistance or enhance symbiosis with microbes has mainly been attempted or conducted through targeted manipulation of single genes/pathways of plants. Recent advancements in synthetic biology tool development are paving the way for multi-gene characterization and engineering in plants in relation to plant-microbe interactions. Here, we briefly summarize the current understanding of plant molecular pathways involved in plant interactions with beneficial and pathogenic microorganisms. Then, we highlight the progress in applying plant synthetic biology to elucidate the molecular basis of plant responses to microbes, enhance plant disease resistance, engineer synthetic symbiosis, and conduct in situ microbiome engineering. Lastly, we discuss the challenges, opportunities, and future directions for advancing plant-microbe interactions research using the capabilities of plant synthetic biology.
|
|
Scooped by
?
December 19, 4:23 PM
|
Approaches are needed to reduce the impact of climate change and provide freshwater. Synthetic biology, and plant synthetic biology specifically, is in an exceptional position to address these needs. Plants are powerful synthetic biology platforms as they derive their energy from the sun (photosynthesis) while capturing the potent greenhouse gas, carbon dioxide. Historically, plants were used in specialized circumstances to provide and store freshwater. Even today many mangrove species can filter seawater while some plants can store massive amounts of water. I propose that starting from nature's own design principles we could engineer plants to filter seawater and/or store freshwater. Moreover, synthetic biologists could derive components found in diverse life forms, modified by water's evolutionarily adaptive properties, to provide source material for engineering plants to filter and transport water. By combining components and inspiration from nature, with advanced synthetic biology tools such as computational protein design and directed evolution, we have the knowledge and technology to make an impact on climate change. Synthetic biologists could engineer plant platforms with the ability to filter saltwater while producing freshwater thus providing a powerful and sustainable means to address freshwater needs while reducing the impact of climate change. With creativity and time, plant synthetic biology could thus help provide sustainable solutions.
|
|
Scooped by
?
December 19, 4:17 PM
|
White biotechnology stands as a major sustainable alternative to address pressing environmental issues arising from our heavy dependence on petrochemical synthesis. However, reaching this goal, both technologically and economically, will take time, resources and money. A major reason is within the biological system itself, as it has evolved into a bow-tie structure in which carbon and energy are converted, via highly regulated, complex and interconnected metabolic networks, into cellular components for growth and homeostasis. This objective is fundamentally at odds with that of biotechnology, which aims to convert carbon and energy into bioproducts. Engineering of microorganism using systems and synthetic biological systems tools has been developed to provide a compromise between these two objectives. However, these genetic and metabolic interventions have revealed often unexpected physiological behaviors, in part due to the fact that a large proportion of metabolic enzymes are catalyzing other reactions than those for which they were evolved. While this promiscuity is the source of an underground metabolism that can prove very advantageous in building high-performance production routes, it is also responsible for loss of yield and production due to metabolic disturbances, negative cross-talks between natural and heterologous pathways as well as it is at the onset of metabolic damages. Identifying these promiscuous enzymes and thus anticipating their opportunities or weaknesses in engineering microbial cell factories for bioproduction is a major challenge in order to improve their performance. It is foreseen that machine learning tools operating on databases continuously fed by genetic, metabolic, enzymatic and fermentation processes data can help to overcome these challenges and provide a better understanding of the physiological functioning of the microbial system.
|
|
Scooped by
?
December 19, 4:07 PM
|
Owing to the complexity of living cells, multicellular systems exhibit heterogeneity at both the macro (different cell types) and the micro (molecular states within a cell type) levels. Traditionally, such heterogeneity has challenged the yield and quality of stem cell-derived cell therapy manufacturing. Here, we argue that heterogeneity can instead be harnessed as a design feature in the quality-by-design toolkit to optimize cell therapy yield and quality, thereby improving bioprocess robustness. We propose a framework for mapping input cell state to output cell fate using systems and synthetic biology tools. This framework can be used to define material and critical quality attributes at the molecular level that better predict drug safety and efficacy. By understanding the sources and consequences of heterogeneity, we can harness it to conquer complex cell therapy manufacturing and bring it to the level of robustness currently only achieved for biologics and small molecules.
|
|
Scooped by
?
December 19, 1:58 AM
|
Type IV secretion systems (T4SS) enable the spread of antibiotic resistance and other virulence factors. In Gram-positive bacteria, T4SSs have long been thought to lack VirB2-like proteins that form conjugative pili and instead rely on adhesins for cell-cell contacts. Yet, it has remained unclear how subsequent DNA transfer (conjugation) is physically mediated. Here we identify a VirB2-like protein, PrgFB2, from the clinically isolated conjugative plasmid pCF10 in Enteroccocus faecalis and show that it is essential for conjugation. Structural modeling confidently predicts a pilus-like assembly for PrgFB2. We validate this prediction through mutagenesis, conjugation assays, and targeted chemical labeling. By combining various machine learning bioinformatic techniques, we analysed >1000 Gram-positive conjugative plasmids from diverse species, including major pathogens, and identified pili forming VirB2-like proteins in almost all of them. Our findings overturn the prevailing view that Gram-positive T4SSs lack pili. This discovery provides a new framework for understanding horizontal gene transfer and highlights critical targets for combating antimicrobial resistance and virulence in Gram-positive bacteria.
|
|
Scooped by
?
December 19, 1:51 AM
|
Prime editing (PE) enables the precise installation of intended base substitutions, small deletions or small insertions into the genome of living cells. While the use of Cas9 nickase can avoid DNA double-strand breaks (DSB), undesired insertions and deletions (indels) often accompany the correct edits, particularly when PE activity increased. Here we show that the anti-CRISPR (Acr) protein AcrIIA5 can significantly enhance PE activity by up to 8.2-fold while markedly reducing byproduct indels. Further investigation reveals that AcrIIA5 can promote PE across various approaches (PE2, PE3, PE4, PE5, and PE6), edit types (substitutions, insertions and deletions), and endogenous loci. Mechanistically, AcrIIA5 appears to inhibit the re-nicking activity of PE complex rather than enhancing the core editing machinery itself, suggesting a distinct mode of interaction with Cas9. Overall, we demonstrate that a known “inhibitor” Acr protein can unexpectedly acting as an “enhancer” of CRISPR/Cas-based genome editing, providing an effective strategy to optimize PE specificity and activity. Prime editing enables precise genetic modification but often suffers from unwanted byproducts. Here, authors show that the anti-CRISPR protein AcrIIA5 unexpectedly enhances prime editing efficiency while reducing unintended indels, offering an effective strategy to improve activity and specificity.
|
