BioTech最新文献

{"title":"Green Biocatalysis of Xylitol Monoferulate: <i>Candida antarctica</i> Lipase B-Mediated Synthesis and Characterization of Novel Bifunctional Prodrug.","authors":"Federico Zappaterra, Francesco Presini, Domenico Meola, Chaimae Chaibi, Simona Aprile, Lindomar Alberto Lerin, Pier Paolo Giovannini","doi":"10.3390/biotech14020025","DOIUrl":"https://doi.org/10.3390/biotech14020025","url":null,"abstract":"<p><p>Natural compounds with significant bioactive properties can be found in abundance within biomasses. Especially prominent for their anti-inflammatory, neuroprotective, antibacterial, and antioxidant activities are cinnamic acid derivatives (CAs). Ferulic acid (FA), a widely studied phenylpropanoid, exhibits a broad range of therapeutic and nutraceutical applications, demonstrating antidiabetic, anticancer, antimicrobial, and hepato- and neuroprotective activities. This research investigates the green enzymatic synthesis of innovative and potentially bifunctional prodrug derivatives of FA, designed to enhance solubility and stability profiles. Selective esterification was employed to conjugate FA with xylitol, a biobased polyol recognized for its bioactive antioxidant properties and safety profile. Furthermore, by exploiting <i>t</i>-amyl alcohol as a green solvent, the enzymatic synthesis of the derivative was optimized for reaction parameters including temperature, reaction time, enzyme concentration, and molar ratio. The synthesized derivative, xylitol monoferulate (XMF), represents a novel contribution to the literature. The comprehensive characterization of this compound was achieved using advanced spectroscopic methods, including ¹H-NMR, ¹³C-NMR, COSY, HSQC, and HMBC. This study represents a significant advancement in the enzymatic synthesis of high-value biobased derivatives, demonstrating increased biological activities and setting the stage for future applications in green chemistry and the sustainable production of bioactive compounds.</p>","PeriodicalId":34490,"journal":{"name":"BioTech","volume":"14 2","pages":""},"PeriodicalIF":2.7,"publicationDate":"2025-04-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12015828/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144056849","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Screening of Non-Conventional Yeasts on Low-Cost Carbon Sources and Valorization of Mizithra Secondary Cheese Whey for Metabolite Production.","authors":"Gabriel Vasilakis, Rezart Tefa, Antonios Georgoulakis, Dimitris Karayannis, Ioannis Politis, Seraphim Papanikolaou","doi":"10.3390/biotech14020024","DOIUrl":"https://doi.org/10.3390/biotech14020024","url":null,"abstract":"<p><p>The production of microbial metabolites such as (exo)polysaccharides, lipids, or mannitol through the cultivation of microorganisms on sustainable, low-cost carbon sources is of high interest within the framework of a circular economy. In the current study, two non-extensively studied, non-conventional yeast strains, namely, <i>Cutaneotrichosporon curvatus</i> NRRL YB-775 and <i>Papiliotrema laurentii</i> NRRL Y-3594, were evaluated for their capability to grow on semi-defined lactose-, glycerol-, or glucose-based substrates and produce value-added metabolites. Three different nitrogen-to-carbon ratios (i.e., 20, 80, 160 mol/mol) were tested in shake-flask batch experiments. Pretreated secondary cheese whey (SCW) was used for fed-batch bioreactor cultivation of <i>P. laurentii</i> NRRL Y-3594, under nitrogen limitation. Based on the screening results, both strains can grow on low-cost substrates, yielding high concentrations of microbial biomass (>20 g/L) under nitrogen-excess conditions, with polysaccharides comprising the predominant component (>40%, <i>w</i>/<i>w</i>, of dry biomass). Glucose- and glycerol-based cultures of <i>C. curvatus</i> promote the secretion of mannitol (13.0 g/L in the case of glucose, under nitrogen-limited conditions). The lipids (maximum 2.2 g/L) produced by both strains were rich in oleic acid (≥40%, <i>w</i>/<i>w</i>) and could potentially be utilized to produce second-generation biodiesel. SCW was nutritionally sufficient to grow <i>P. laurentii</i> strain, resulting in exopolysaccharides secretion (25.6 g/L), along with dry biomass (37.9 g/L) and lipid (4.6 g/L) production.</p>","PeriodicalId":34490,"journal":{"name":"BioTech","volume":"14 2","pages":""},"PeriodicalIF":2.7,"publicationDate":"2025-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12015851/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144022436","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Developments in Extracellular Matrix-Based Angiogenesis Therapy for Ischemic Heart Disease: A Review of Current Strategies, Methodologies and Future Directions.","authors":"Jad Hamze, Mark Broadwin, Christopher Stone, Kelsey C Muir, Frank W Sellke, M Ruhul Abid","doi":"10.3390/biotech14010023","DOIUrl":"https://doi.org/10.3390/biotech14010023","url":null,"abstract":"<p><p>Ischemic heart disease (IHD) is the leading cause of mortality worldwide, underscoring the urgent need for innovative therapeutic strategies. The cardiac extracellular matrix (ECM) undergoes extreme transformations during IHD, adversely influencing the heart's structure, mechanics, and cellular signaling. Researchers investigating the regenerative capacity of the diseased heart have turned their attention to exploring the modulation of ECM to improve therapeutic outcomes. In this review, we thoroughly examine the current state of knowledge regarding the cardiac ECM and its therapeutic potential in the ischemic myocardium. We begin by providing an overview of the fundamentals of cardiac ECM, focusing on the structural, functional, and regulatory mechanisms that drive its modulation. Subsequently, we examine the ECM's interactions within both chronically ischemic and acutely infarcted myocardium, emphasizing key ECM components and their roles in modulating angiogenesis. Finally, we discuss recent ECM-based approaches in biomedical engineering, focusing on different types of scaffolds as delivery tools and their compositions, and conclude with future directions for therapeutic research. By harnessing the potential of these emerging ECM-based therapies, we aim to contribute to the development of novel therapeutic modalities for IHD.</p>","PeriodicalId":34490,"journal":{"name":"BioTech","volume":"14 1","pages":""},"PeriodicalIF":2.7,"publicationDate":"2025-03-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11940646/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144015225","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Identifying Antimicrobial Agents from <i>Chlorella sorokiniana</i>: A Biotechnological Approach Utilizing Eco-Friendly Extraction and Characterization Methods.","authors":"Elia Lio, Martina Dramis, Gianluca Ottolina, Francesco Secundo","doi":"10.3390/biotech14010022","DOIUrl":"https://doi.org/10.3390/biotech14010022","url":null,"abstract":"<p><p>Natural compounds are increasingly favored over synthetic ones for their lower environmental impact. However, extraction and characterization processes typically rely on harsh conditions and conventional solvents, which are unsustainable and cause pollution. This study aimed to develop an eco-friendly extraction method to isolate and evaluate the antimicrobial properties of bioactive compounds from <i>Chlorella sorokiniana</i>. Using dimethyl carbonate (DMC), methoxycyclopentane (CPME), and butan-2-one (MEK) as green solvents alongside chloroform as a non-green reference solvent, on both untreated and sodium hydroxide pre-treated microalgae biomass, extract yields of up to 182 ± 27 mg/g DW were obtained using MEK. Extracts from untreated microalgae biomass exhibited lower MIC values compared to those obtained with the same solvent from pre-treated biomass, when tested as antimicrobial agents against <i>Escherichia coli</i>, <i>Bacillus megaterium</i>, and <i>Bacillus subtilis</i>. The lowest MIC value (4.89 ± 0.05 µg/mL) was observed against <i>E. coli</i> using the extract from the untreated microalgae biomass with CPME, which was comparable to the vancomycin control (1.55 ± 0.03 µg/mL). Principal component analysis highlighted correlations between GC-MS-identified compounds and antimicrobial activity. ANOVA and post hoc tests (<i>p</i> < 0.05) confirmed solvent choice, and pre-treatment influenced yield and bioactivity. The results underscore green solvents as sustainable alternatives for extracting bioactive compounds from autotrophic microalgae.</p>","PeriodicalId":34490,"journal":{"name":"BioTech","volume":"14 1","pages":""},"PeriodicalIF":2.7,"publicationDate":"2025-03-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11939975/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143988278","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Genomic Characterization of Extremely Antibiotic-Resistant Strains of <i>Pseudomonas aeruginosa</i> Isolated from Patients of a Clinic in Sincelejo, Colombia.","authors":"Nerlis Pajaro-Castro, Erick Diaz-Morales, Kenia Hoyos","doi":"10.3390/biotech14010021","DOIUrl":"https://doi.org/10.3390/biotech14010021","url":null,"abstract":"<p><p><i>Pseudomonas aeruginosa</i> is an opportunistic pathogen classified as a priority and a great public health concern; therefore, this research focuses on the genomic characterization of extremely resistant strains of <i>P. aeruginosa</i> isolated from patients in a clinic in Sincelejo, Colombia. Seven strains were analyzed by whole genome sequencing using the Illumina NovaSeq platform, with a focus on the identification of resistance genes and virulence factors through the CARD and VFDB databases. An ANI (Average Nucleotide Identity) analysis was carried out to determine the genetic relationship between the strains, complemented by a phylogenomic analysis to place the strains in different evolutionary clades. The results revealed that six of the strains are of Colombian origin, while one strain (547256) belongs to the high-risk clone ST773, previously unidentified in Colombia. Genome size ranged from 6 to 7.4 Mbp, indicating differences in genetic content among strains. Phylogenomic analysis confirmed that five strains belong to a multidrug-resistant (MDR) group, while one strain (572897) showed high alignment with a laboratory strain, and strain 547256 was not associated with any specific clade. Clinically, 100% of strains showed carbapenem resistance, resistance genes, and virulence factors that make them difficult to treat. This study provides key insights into the genetic diversity and resistance patterns of <i>P. aeruginosa</i> in this region, underscoring the need to monitor high-risk clones and optimize therapeutic strategies.</p>","PeriodicalId":34490,"journal":{"name":"BioTech","volume":"14 1","pages":""},"PeriodicalIF":2.7,"publicationDate":"2025-03-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11939883/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144052565","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Using Nano-Luciferase Binary (NanoBiT) Technology to Assess the Interaction Between Viral Spike Protein and Angiotensin-Converting Enzyme II by Aptamers.","authors":"Meng-Wei Lin, Cheng-Han Lin, Hua-Hsin Chiang, Irwin A Quintela, Vivian C H Wu, Chih-Sheng Lin","doi":"10.3390/biotech14010020","DOIUrl":"https://doi.org/10.3390/biotech14010020","url":null,"abstract":"<p><p>Nano-luciferase binary technology (NanoBiT)-based pseudoviral sensors are innovative tools for monitoring viral infection dynamics. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infects host cells via its trimeric surface spike protein, which binds to the human angiotensin-converting enzyme II (hACE2) receptor. This interaction is crucial for viral entry and serves as a key target for therapeutic interventions against coronavirus disease 2019 (COVID-19). Aptamers, short single-stranded DNA (ssDNA) or RNA molecules, are highly specific, high-affinity biorecognition elements for detecting infective pathogens. Despite their potential, optimizing viral infection assays using traditional protein-protein interaction (PPI) methods often face challenges in optimizing viral infection assays. In this study, we selected and evaluated aptamers for their ability to interact with viral proteins, enabling the dynamic visualization of infection progression. The NanoBiT-based pseudoviral sensor demonstrated a rapid increase in luminescence within 3 h, offering a real-time measure of viral infection. A comparison of detection technologies, including green fluorescent protein (GFP), luciferase, and NanoBiT technologies for detecting PPI between the pseudoviral spike protein and hACE2, highlighted NanoBiT's superior sensitivity and performance, particularly in aptamer selection. This bioluminescent system provides a robust, sensitive, and early-stage quantitative approach to studying viral infection dynamics.</p>","PeriodicalId":34490,"journal":{"name":"BioTech","volume":"14 1","pages":""},"PeriodicalIF":2.7,"publicationDate":"2025-03-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11940275/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144039064","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Characterization of a GH43 Bifunctional Glycosidase from Endophytic <i>Chaetomium globosum</i> and Its Potential Application in the Biotransformation of Ginsenosides.","authors":"Yao Lu, Qiang Jiang, Yamin Dong, Runzhen Ji, Yiwen Xiao, Du Zhu, Boliang Gao","doi":"10.3390/biotech14010018","DOIUrl":"https://doi.org/10.3390/biotech14010018","url":null,"abstract":"<p><p>The GH43 family of glycosidases represents an important class of industrial enzymes that are widely utilized across the food, pharmaceutical, and various other sectors. In this study, we identified a GH43 family glycoside hydrolytic enzyme, <i>Xyaf313</i>, derived from the plant endophytic fungus <i>Chaetomium globosum</i> DX-THS3, which is capable of transforming several common ginsenosides. The enzyme function analysis reveals that Xyaf313 exhibits dual functionality, displaying both α-L-arabinofuranosidase and β-D-xylosidase activity. When acting as an α-L-arabinofuranosidase, Xyaf313 achieves optimal enzyme activity of 23.96 U/mg at a temperature of 50 °C and a pH of 7. In contrast, its β-D-xylosidase activity results in a slight reduction in enzyme activity to 23.24 U/mg, with similar optimal temperature and pH conditions to those observed for the α-L-arabinofuranosidase activity. Furthermore, Xyaf313 demonstrates considerable resistance to most metal ions and common chemical reagents. Notably, while the maximum enzyme activity of Xyaf313 occurs at 50 °C, it maintains high activity at room temperature (30 °C), with relative enzyme activity exceeding 90%. Measurements of ginsenoside transformation show that Xyaf313 can convert common ginsenosides Rc, Rb₁, Rb₂, and Rb₃ into Rd, underscoring its potential for pharmaceutical applications. Overall, our findings contribute to the identification of a new class of bifunctional GH43 glycoside hydrolases, highlight the significance of plant endophytic fungi as a promising resource for the screening of carbohydrate-decomposing enzymes, and present new candidate enzymes for the biotransformation of ginsenosides.</p>","PeriodicalId":34490,"journal":{"name":"BioTech","volume":"14 1","pages":""},"PeriodicalIF":2.7,"publicationDate":"2025-03-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11940195/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144027439","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A Coupled Model of the Cardiovascular and Immune Systems to Analyze the Effects of COVID-19 Infection.","authors":"Camila Ribeiro Passos, Alexandre Altamir Moreira, Ruy Freitas Reis, Rodrigo Weber Dos Santos, Marcelo Lobosco, Bernardo Martins Rocha","doi":"10.3390/biotech14010019","DOIUrl":"https://doi.org/10.3390/biotech14010019","url":null,"abstract":"<p><p>The COVID-19 pandemic has underscored the importance of understanding the interplay between the cardiovascular and immune systems during viral infections. SARS-CoV-2 enters human cells via the ACE-2 enzyme, initiating a cascade of immune responses. This study presents a coupled mathematical model that integrates the cardiovascular system (CVS) and immune system (IS), capturing their complex interactions during infection. The CVS model, based on ordinary differential equations, describes heart dynamics and pulmonary and systemic circulation, while the IS model simulates immune responses to SARS-CoV-2, including immune cell interactions and cytokine production. A coupling strategy transfers information from the IS to the CVS at specific intervals, enabling the exploration of immune-driven cardiovascular effects. Numerical simulations examined how these interactions influence infection severity and recovery. The coupled model accurately replicated the evolution of cardiac function in survivors and non-survivors of COVID-19. Survivors exhibited a left ventricular ejection fraction (LVEF) reduction of up to 25% while remaining within normal limits, whereas non-survivors showed a severe 4-fold decline, indicative of myocardial dysfunction. Similarly, the right ventricular ejection fraction (RV EF) decreased by approximately 50% in survivors but underwent a drastic 5-fold reduction in non-survivors. These findings highlight the model's capacity to distinguish differential cardiac dysfunction across clinical outcomes and its potential to enhance our understanding of COVID-19 pathophysiology.</p>","PeriodicalId":34490,"journal":{"name":"BioTech","volume":"14 1","pages":""},"PeriodicalIF":2.7,"publicationDate":"2025-03-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11940274/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144017217","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Chemical Profiling and Bioactivity of Microalgae Extracts for Enhancing Growth and Anthracnose Resistance in the Common Bean (<i>Phaseolus vulgaris</i> L.).","authors":"Alessandro A Dos Santos, Camila Nader, Mateus B de Freitas, César F Ribeiro, Geovanna de Oliveira Costa, Louis P Sandjo, Alex S Poltronieri, Roberto B Derner, Marciel J Stadnik","doi":"10.3390/biotech14010017","DOIUrl":"https://doi.org/10.3390/biotech14010017","url":null,"abstract":"<p><p>The present study aimed to chemically profile the hydroalcoholic extracts from the microalgae (MEs) <i>Nannochloropsis oculata</i>, <i>Phaeodactylum tricornutum</i>, <i>Tetradesmus obliquus</i>, and <i>Tetraselmis tetrathele</i> and evaluate their effects on the development of <i>Colletotrichum lindemuthianum</i> and anthracnose symptoms, as well as on the initial growth of bean plants. For this, MEs were analyzed using UPLC coupled with a mass spectrometer, allowing the identification of peaks and annotation of potential metabolites. Fungal mycelial growth was assessed seven days after inoculation, and conidial germination was measured 72 h after incubation, using ME concentrations of 0, 0.1, 0.5, and 1.0 mg·mL^-1. Bean seeds of the IPR Uirapuru cultivar were sown and treated with 3 mL of extracts at four time points: at sowing and 72 h after each previous treatment. After 11 days of cultivation in a growth chamber, the plants were divided into two groups: one for anthracnose control assessment and the other for evaluating growth promotion by MEs. Plant length as well as fresh and dry weights of shoots and roots were determined, leaf pigments were quantified, and anthracnose severity was assessed using a diagrammatic scale. The UPLC analysis identified 32 compounds in the extracts of the four microalgae, belonging to different chemical and functional groups, with lipids being the most significant fraction. The extracts exhibited variability and diversity in chemical composition depending on the microalgal species. MEs did not affect mycelial growth yet increased the germination of <i>C. lindemuthianum</i> conidia, regardless of the dose or species used. Anthracnose severity was not affected by the microalgae extracts. Regarding growth promotion, the extracts showed varying effects but generally increased shoot and root length, fresh biomass, and leaf pigment content.</p>","PeriodicalId":34490,"journal":{"name":"BioTech","volume":"14 1","pages":""},"PeriodicalIF":2.7,"publicationDate":"2025-03-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11940543/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144022314","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Towards a Survival-Based Cellular Assay for the Selection of Protease Inhibitors in <i>Escherichia coli</i>.","authors":"William Y Oyadomari, Elizangela A Carvalho, Gabriel E Machado, Ana Júlia O Machado, Gabriel S Santos, Marcelo Marcondes, Vitor Oliveira","doi":"10.3390/biotech14010016","DOIUrl":"https://doi.org/10.3390/biotech14010016","url":null,"abstract":"<p><p>We describe a method tailored to the in-cell selection of protease inhibitors. In this method, a target protease is co-expressed with a selective substrate, the product of which kills host cells. Therefore, the method can be applied to identify potential inhibitors based on cell host survival when inhibition of the target protease occurs. The TEV protease was chosen for this proof-of-concept experiment. The genetically encoded selective substrate is a single polypeptide chain composed of three parts: (1) a ccdB protein, which can cause host cell death when it accumulates inside the cell; (2) a protease cleavage sequence that can be changed according to the target protease, in this case the TEV substrate ENLYFQ↓G (↓-predicted cleavage site); and (3) the ssrA sequence (AANDENYALAA), which drives the polypeptide to degradation by the ClpX/ClpP complex inside host <i>E. coli</i> cells. In our experiment, co-expression of the active TEV protease and this selective substrate (ccdB-ENLYFQG-ssrA) caused the death of a significant host cell population, while control assays with an inactive mutant TEV Asp81Asn did not. Details of the methodology used are given, providing the basis for the application of similar systems for other proteases of interest.</p>","PeriodicalId":34490,"journal":{"name":"BioTech","volume":"14 1","pages":""},"PeriodicalIF":2.7,"publicationDate":"2025-03-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11940102/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144027444","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}