Journal of Biological Engineering最新文献

{"title":"Natural diversity-guided catalytic-core chimerism engineers a rapid and inhibitor-tolerant reverse transcriptase.","authors":"Inês Fonseca Costa, Vânia Ondina Fernandes, Rita Silva Simões, Hélvio Simões, Virgínia Maria Rico Pires, Victor Diogo Alves, João Soeiro Teodoro, Pedro Bule, Carlos Mendes Godinho de Andrade Fontes","doi":"10.1186/s13036-026-00692-3","DOIUrl":"https://doi.org/10.1186/s13036-026-00692-3","url":null,"abstract":"<p><strong>Background: </strong>Reverse transcriptases (RTs) are essential components of RNA-based molecular technologies, yet their practical performance is often constrained by trade-offs between catalytic speed, thermal stability, and tolerance to inhibitory substances. Conventional optimization via mutagenesis or directed evolution often improves individual traits but struggles to integrate multiple traits within a single enzyme. Here, we applied catalytic-core recombination, guided by natural sequence diversity and diagnostic performance criteria, to recover reverse transcriptase phenotypes that would be difficult to obtain by single-trait optimization alone.</p><p><strong>Results: </strong>Starting from 1,028 sequences, 24 representative RT variants were used to generate a library of chimeras by swapping a critical 405-residue polymerase domain into a validated M-MuLV scaffold. We identified chRT V18 as the lead candidate, demonstrating consistent activity across a broad temperature range (40-70 °C). In addition, chRT V18 enabled efficient and linear cDNA synthesis in as little as 1 min at elevated temperatures, while maintaining strong resistance to clinically relevant inhibitors. Beyond RT-qPCR, chRT V18 supported high-temperature RT-LAMP at 69 °C, a regime typically incompatible with conventional RTs, enabling improved specificity and rapid amplification. When formulated into one-step RT-qPCR master mixes, chRT V18 achieved low limits of detection and full concordance with IVD-approved assays, while reducing reverse transcription time by 90%, enabling shorter time-to-result.</p><p><strong>Conclusions: </strong>Natural diversity-guided catalytic-core chimerism enabled the development of a rapid, inhibitor-tolerant reverse transcriptase with an unusual combination of broad temperature compatibility, minute-scale reverse transcription, and resilience to clinically relevant inhibitors. Beyond the properties of chRT V18 itself, the results support modular recombination of evolutionarily optimized domains as a practical engineering strategy for integrating multiple performance traits in complex enzymes.</p>","PeriodicalId":15053,"journal":{"name":"Journal of Biological Engineering","volume":" ","pages":""},"PeriodicalIF":6.5,"publicationDate":"2026-05-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147838437","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Proof-of-concept engineering of Escherichia coli expressing a bee-derived cytochrome P450 monooxygenase for thiamethoxam detoxification.","authors":"Pachara Sattayawat, Kasimaporn Promubon, Kritapas Sripinta, Warisa Chunguaongsuk, Chonlaphon Juntama, Nuttapol Noirungsee, Aussara Panya, Terd Disayathanoowat","doi":"10.1186/s13036-026-00686-1","DOIUrl":"https://doi.org/10.1186/s13036-026-00686-1","url":null,"abstract":"<p><p>Bees are unintentionally exposed to pesticides applied to control other insect pests. Although bees possess endogenous detoxification mechanisms, their efficiency may not always be sufficient. To this end, the use of probiotics that can assist in pesticide detoxification may offer an additional layer of protection. In this study, we engineered Escherichia coli BL21(DE3) to heterologously express the bee-derived cytochrome P450 monooxygenase (CYP450), CYP9Q1, from Apis mellifera as a proof-of-concept microbial platform for thiamethoxam biotransformation and future microbiome-assisted pesticide detoxification strategies. We first assessed the toxicity of thiamethoxam, a systemic insecticide, on E. coli, which showed obvious impaired growth at relatively high concentrations (5 and 10 g/L). Subsequently, we computationally modeled the 3D structure and modified the CYP450 to improve its solubility when expressed in E. coli. Molecular docking of the CYP450 and thiamethoxam was also performed to confirm their potential interaction. pRSFDuet-1 plasmid was used to carry the modified CYP450 gene for expression in E. coli and the induction condition was optimized, with 0.5 mM isopropyl 𝛽-D-1-thiogalactopyranoside (IPTG) yielding the most favorable expression level. A whole-cell detoxification assay subsequently showed reduced concentrations of thiamethoxam after 46 h of biocatalytic activity, as monitored by High-Performance Liquid Chromatography (HPLC). Liquid Chromatography-Mass Spectrometry (LC-MS) analysis further revealed the presence of desnitro thiamethoxam corresponding to m/z 247.04105 confirming enzymatic transformation of thiamethoxam. To our knowledge, this study provides the first demonstration of bee CYP9Q1 heterologous expression in E. coli for whole-cell thiamethoxam biotransformation, highlighting its potential as a platform for developing probiotic or gut microbiome-based detoxification strategies to support bee pesticide tolerance.</p>","PeriodicalId":15053,"journal":{"name":"Journal of Biological Engineering","volume":" ","pages":""},"PeriodicalIF":6.5,"publicationDate":"2026-05-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147838390","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Early bioprocess development using Komagataella phaffii: application scenarios for the microTOM mini bioreactor system.","authors":"David Wollborn, Carol J Hartley, Andreas Schulte, Juan C P Correa","doi":"10.1186/s13036-026-00691-4","DOIUrl":"https://doi.org/10.1186/s13036-026-00691-4","url":null,"abstract":"<p><p>Developing novel bioprocesses usually starts with an iterative cycle to engineer genetically modified organisms. This leads to large strain libraries that differ in their genetic makeup, showing different growth and production performance. Finding the optimal production strain requires extensive screening and is arguably one of the most important steps for large-scale productivity. For aerobic processes, it is important to ensure the cultivation conditions avoid oxygen limitation. The oxygen transfer rate (OTR) is a parameter to monitor metabolic activity. The micro-scale Transfer rate Online Measurement (microTOM) system allows online monitoring of the respiration activity in 96-individual wells of a microtiter plate. This novel late-stage beta version prototype, provided by Kuhner Shaker, was trialled with the common microbial host Komagataella phaffii. It is demonstrated how the microTOM system can be applied to rapidly determine essential parameters for bioprocess design. Using a 96-well microtiter plate format enabled high-throughput investigation of three strains in parallel. The methanol uptake and growth rate was estimated, which can be used in process models to guide scale-up and decisions regarding feeding rates in fed-batch operation mode. Further, we assessed seven media formulations with two carbon sources, the impact of filling volume on the maximum oxygen transfer capacity, eight different antifoam agents, and growth rates were estimated at five different temperatures. These findings can help select screening conditions and demonstrate how the microTOM system can accelerate bioprocess development. Until now, obtaining high-resolution OTR datasets at high-throughput, suitable for deriving critical process parameters, has not been feasible. Thus, the microTOM supports robust, data-driven bioprocess development.</p>","PeriodicalId":15053,"journal":{"name":"Journal of Biological Engineering","volume":" ","pages":""},"PeriodicalIF":6.5,"publicationDate":"2026-05-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147838301","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Mechanical isolation of stromal vascular fraction using a spiral-based device: cellular outcomes and point-of-care workflow considerations.","authors":"Marcos Sforza, Olga Ivanenko, Nazanin Biabani, Lara Sforza, Deepak M Kalaskar, Zahra Mohri, Afshin Mosahebi","doi":"10.1186/s13036-026-00688-z","DOIUrl":"https://doi.org/10.1186/s13036-026-00688-z","url":null,"abstract":"<p><strong>Background: </strong>Stromal vascular fraction (SVF) from adipose tissue represents a promising source of regenerative cells. A major trend in SVF isolation is the increasing use of dedicated devices and systems to enhance standardization and reproducibility. Real-world implementation of new technologies requires consideration of workflow-related factors alongside biological results. This study primarily aimed to characterize the novel BMC Uniq® SVF device, describe a mechanical isolation approach (MI-SVF) with this device, and evaluate the cellular outcomes of MI-SVF. Enzymatic isolation (EI-SVF) was included as a reference method for comparison of selected cellular parameters.</p><p><strong>Results: </strong>The BMC Uniq® SVF device is a spiral-based, closed system composed of two housings and three filters, designed as a sterile disposable unit. Lipoaspirate samples from 10 healthy female donors were equally divided and handled using MI-SVF (decantation, device processing, centrifugation) or EI-SVF (adding collagenase and post-digestion steps to the MI-SVF protocol). Total cell yield was significantly greater in the EI-SVF group (mean ± standard error (SE) 94.9 × 10⁴ ± 6.4 × 10⁴ versus 37.5 × 10⁴ ± 2.7 × 10⁴ for MI-SVF; p = 0.002). The flow cytometry demonstrated significantly higher relative proportions of CD90+ (10.3 ± 1.3% versus 5.4 ± 0.9%; p = 0.014) and CD73+ cells (10.7 ± 0.7% versus 6.6 ± 0.7%; p = 0.010) in MI-SVF compared with EI-SVF, with no significant differences in CD105+ percentage. There were no differences in cell viability (MI-SVF: mean 87.0%; EI-SVF: 83.9%). MI-SVF samples showed a CFU-F frequency of 5.0 ± 0.3% and consistently demonstrated adipogenic, chondrogenic, and osteogenic differentiation potential.</p><p><strong>Conclusions: </strong>The BMC Uniq® SVF device is a simple closed single-use system for mechanical SVF extraction in point-of-care applications. Fresh MI-SVF samples derived in such a way had one of the highest reported cell yields and demonstrated viability, immunophenotype, clonogenic capacity, and multilineage differentiation potential that were comparable to, and in some cases exceeded, those of previously described non-enzymatic protocols. Relative to EI-SVF, the mechanical approach showed a lower total cell count but a higher relative percentage of CD73+ and CD90+ cells. In general, this work contributes to a broader framework for assessing SVF isolation methods by integrating biological performance with practical aspects of clinical workflow.</p>","PeriodicalId":15053,"journal":{"name":"Journal of Biological Engineering","volume":" ","pages":""},"PeriodicalIF":6.5,"publicationDate":"2026-05-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147838363","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Therapeutic efficacy of mesenchymal stem cell-derived extracellular vesicles combined with injectable hydrogels in bone defect repair: a systematic review and meta-analysis of preclinical studies.","authors":"Amir Hossein Aghayan, Davood Mohammadi, Hamed Soleimani Samarkhazan, Amir Atashi, Zahra Jamalpoor","doi":"10.1186/s13036-026-00684-3","DOIUrl":"https://doi.org/10.1186/s13036-026-00684-3","url":null,"abstract":"","PeriodicalId":15053,"journal":{"name":"Journal of Biological Engineering","volume":" ","pages":""},"PeriodicalIF":6.5,"publicationDate":"2026-04-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147771901","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Trends of nucleic acid - based point-of-care diagnostics for infectious diseases.","authors":"Hossam Hatem, Mohamed Mysara, Raghda Ramadan","doi":"10.1186/s13036-026-00681-6","DOIUrl":"10.1186/s13036-026-00681-6","url":null,"abstract":"","PeriodicalId":15053,"journal":{"name":"Journal of Biological Engineering","volume":"20 1","pages":""},"PeriodicalIF":6.5,"publicationDate":"2026-04-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13130695/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147771882","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Metabolic rewiring for enhanced itaconate yield through heterotrophic CO₂ fixation in Escherichia coli.","authors":"Dae-Yeol Ye, Myeongwon Min, Ga Yeon Lim, Hyun Gyu Hwang, Gyoo Yeol Jung, Hyun Gyu Lim","doi":"10.1186/s13036-026-00687-0","DOIUrl":"https://doi.org/10.1186/s13036-026-00687-0","url":null,"abstract":"","PeriodicalId":15053,"journal":{"name":"Journal of Biological Engineering","volume":" ","pages":""},"PeriodicalIF":6.5,"publicationDate":"2026-04-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147771923","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"pH/glucose dual-responsive drug release hydrogel loaded with tannic acid and gallium ions enhances diabetic wound healing upon bacterial infection.","authors":"Le Guo, Pihong Zhang, Minghua Zhang, Pengfei Liang, Situo Zhou","doi":"10.1186/s13036-025-00581-1","DOIUrl":"https://doi.org/10.1186/s13036-025-00581-1","url":null,"abstract":"","PeriodicalId":15053,"journal":{"name":"Journal of Biological Engineering","volume":" ","pages":""},"PeriodicalIF":6.5,"publicationDate":"2026-04-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147771903","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Waste-to-sensor upcycling of polyethylene terephthalate over Ag/Zr-MOF photocatalyst for microplastic degradation and AI-assisted heavy metal detection.","authors":"Minse Kim, Jaewon Choi, Kisung Lee, Subin Lee, Jaewon Lee, Jeong-Ann Park, Kwang Suk Lim, Suk-Jin Ha, Hyun-Ouk Kim","doi":"10.1186/s13036-026-00683-4","DOIUrl":"https://doi.org/10.1186/s13036-026-00683-4","url":null,"abstract":"<p><p>This study aims to develop an integrated waste to sensor platform that couples photocatalytic PET depolymerization with on stream quantification of coexisting heavy metals. We achieve this by preparing AIM-202 through surface coordination of Ag to a water stable Zr-aspartate MOF (MIP-202), creating Ag⁰/Ag^δ+ electron sink sites that enhance charge separation and reactive oxygen species generation while preserving the parent framework. The material was characterized by FT-IR, XRD, XPS, photoluminescence, KPFM, and EPR, and its photocatalytic activity was evaluated under Xe lamp irradiation. AIM-202 (5 mg mL-1) oxidized PET with increased FT-IR oxidation indices and surface pitting, and generated soluble products that increased from 151.40 µmol (week 1) to 277.27 µmol (week 2), corresponding to 11.63% and 28.68% conversion to quantified byproducts. The PET derived fluorophore 2-hydroxyterephthalate enabled fluorescence quenching based Cu²⁺ and Fe³⁺ sensing, and physics guided models refined with lightweight machine learning provided robust concentration back calculation. The novelty lies in a closed loop design in which MOF mediated depolymerization simultaneously produces the sensing reporter and enables data assisted quantification, supporting scalable remediation and monitoring within a single integrated scheme.</p>","PeriodicalId":15053,"journal":{"name":"Journal of Biological Engineering","volume":" ","pages":""},"PeriodicalIF":6.5,"publicationDate":"2026-04-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147771895","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A continuous viral vaccine biomanufacturing platform utilizing multiple bioreactor configurations.","authors":"Justin Sargunas, Bradley Priem, Dylan Carman, Taravat Sarvari, Natalie M Nold, Vaishali Sharma, Andrew Pekosz, Caryn L Heldt, Michael Betenbaugh","doi":"10.1186/s13036-026-00680-7","DOIUrl":"https://doi.org/10.1186/s13036-026-00680-7","url":null,"abstract":"<p><p>Scalable, continuous biomanufacturing processes have grown in importance to meet demand for smaller bioreactor sizes, lowered production costs, and improved quality attribute consistency. The Sf9/recombinant baculovirus (rBV) expression system demonstrates promise for virus-like particle (VLP) vaccine and gene therapy production. Here, we present a continuous rBV platform integrating an infection plug flow reactor (PFR) between stirred tank growth (gCSTR) and production (pCSTR) bioreactors. Cell expansion in the gCSTR included a ramp-up stage followed by continuous growth, reaching a steady state of 5×10⁶ cells/mL and >90% viability. Péclet number-fit tracer studies confirmed near-ideal plug flow in the PFR, yielding a 10 h residence time and progressive infection as measured by gp64 signaling. Finally, a pCSTR with a residence time of 48 h exhibited sustained recombinant protein production. An integrated pilot cascade incorporating all reactors ran continuously for 5 days, maintaining stable CSTR cell densities and a measurable increase in infected cell diameter from 14.5 μm to 16.1 μm. Western blotting and EM of ~ 100 nm VLPs in pCSTR effluent demonstrated platform success. Digital twin mechanistic models across four distinct stages of bioreactor operation and Hill-type relationships for rBV infection kinetics predicted cell growth and death for a 7-day run, demonstrating promise for designing continuous systems in silico and building a quantitative framework for scale-up and optimization. Our multi-stage reactor configuration represents a cell host- and product-agnostic production scheme, particularly for processes prone to product heterogeneity, and paves the way towards a true end-to-end continuous platform for myriad modalities in the future.</p>","PeriodicalId":15053,"journal":{"name":"Journal of Biological Engineering","volume":" ","pages":""},"PeriodicalIF":6.5,"publicationDate":"2026-04-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147771879","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}