Journal of Industrial Microbiology & Biotechnology最新文献

{"title":"Discovering methylated DNA motifs in bacterial nanopore sequencing data with MIJAMP.","authors":"Alyssa K Tidwell, Evelyn Faust, Carrie A Eckert, Adam M Guss, William G Alexander","doi":"10.1093/jimb/kuaf022","DOIUrl":"10.1093/jimb/kuaf022","url":null,"abstract":"<p><p>Bacterial DNA methylation is involved in diverse cellular functions, including modulation of gene expression, DNA repair, and restriction-modification systems for defense against viruses and other foreign DNA. Restriction systems hinder efforts to engineer organisms to produce fuels and chemicals from waste and renewable feedstocks by degrading DNA during transformation. Methylome analysis allows identification of motifs within a bacterial chromosome that may be targeted by native restriction enzymes. Further expression of the corresponding methyltransferases in Escherichia coli allows plasmid DNA to be protected from restriction in the target organism, thereby drastically enhancing transformation efficiency. Nanopore sequencing can detect methylated bases, but software is needed to transform modified base coordinates into methylated motifs. Here, we develop MIJAMP (MIJAMP Is Just A  MethylBED Parser), a software package that was developed to discover methylated motifs from the output of ONT's Modkit or other data in the methylBED format. MIJAMP employs a human-driven refinement strategy that empirically validates all motifs against genome-wide methylation data, thus eliminating incorrect motifs. MIJAMP also reports methylation data on specific, user-defined motifs. Using MIJAMP, we determined the methylated motifs both in a control strain (wild-type E. coli) and in Synecococcus sp. strain PCC7002, laying the foundation for improved transformation in this organism. MIJAMP is available at https://code.ornl.gov/alexander-public/mijamp/. One Sentence Summary: Here we describe software written to discover DNA methylation motifs from nanopore sequencing data.</p>","PeriodicalId":16092,"journal":{"name":"Journal of Industrial Microbiology & Biotechnology","volume":" ","pages":""},"PeriodicalIF":3.2,"publicationDate":"2024-12-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12320774/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144690505","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Anti-Parkinsonian 4-hydroxy-2-pyridones from an endolichenic fungus, Tolypocladium sp. (strain CNC14).","authors":"Jin Won Choi, Chaesun Kwon, Jin Woo Lee, Jae-Seoun Hur, Min-Kyoo Shin, Sang Hee Shim","doi":"10.1093/jimb/kuaf027","DOIUrl":"10.1093/jimb/kuaf027","url":null,"abstract":"<p><p>4-Hydroxy-2-pyridone alkaloids have attracted considerable attention because of their intriguing structures and diverse bioactivities. In our previous study, 4-hydroxy-2-pyridone alkaloids were shown to exhibit potent activity against neuron-associated targets. To discover this class of neuroactive compounds, an array of endolichenic fungal extracts was screened by analyzing liquid chromatography-ultraviolet-mass spectrometry (LC-UV-MS) profiles. The screening yielded strain Tolypocladium sp. (strain CNC14), which produced compounds with characteristic Ultraviolet patterns for 4-hydroxy-2-pyridone alkaloids using our in-house library. Based on these findings, we conducted a chemical investigation, which led to the isolation of four new (1-4) and ten known (5-14) compounds. Their structures were elucidated via spectroscopic methods such as nuclear magnetic resonance and mass spectrometry. The stereochemistry of the new compounds (1-4) was established using rotating frame overhauser effect spectroscopy (ROESY), and the electronic circular dichrosim (ECD) was compared with the calculated data. Interestingly, the side chains of 4-hydroxy-2-pyridone in 1 and 2 were cyclized in different directions to form benzopyrano[3,4-b]pyridinol from previously reported compounds, and all the new compounds were predicted to be biosynthesized from reduced tolypyridone C (7) via the hetero-Diels-Alder reaction. Among the isolated compounds, 4 significantly protected neuronal cells against treatment with 1-methyl-4-phenylpyridinium (MPP+), a Parkinsonian neurotoxin, in an in vitro Parkinson's disease model. One-Sentence Summary: Four new neuroprotective 4-hydroxy-2-pyridone alkaloids were discovered from an endolichenic fungus Tolypocladium sp.</p>","PeriodicalId":16092,"journal":{"name":"Journal of Industrial Microbiology & Biotechnology","volume":" ","pages":""},"PeriodicalIF":3.2,"publicationDate":"2024-12-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12455187/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144957098","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A Multi Clone Kinetic Model for characterizing Chinese hamster ovary cell line variability.","authors":"Devi Sietaram, Pavlos Kotidis, Gary Finka, Alexei A Lapkin","doi":"10.1093/jimb/kuaf029","DOIUrl":"10.1093/jimb/kuaf029","url":null,"abstract":"<p><p>This paper presents the Multi Clone Kinetic Model (MCKM), a novel generalized kinetic mechanistic model for fed-batch cultivations of diverse Chinese hamster ovary (CHO) cell lines, producing different recombinant monoclonal antibodies (mAbs). Unlike traditional kinetic models requiring multiple cultures for one parameter regression, MCKM derives a complete set of 13 kinetic parameters from a single fed-batch cell line culture of 49 data points. This enables per-cell-line metabolic characterization during cell line development, as well as direct comparisons of kinetics across clones, passages, and different recombinant mAbs. To enable MCKM to be broadly applicable across many cell lines and mAbs, and to address the high-dimensional challenge of estimating 13 kinetic parameters from a small number of datapoints, the model uniquely incorporates a mechanistic growth constraint, a glucose-dependent lactate switch, and automated parameter balancing. MCKM demonstrated successful regression of 656 fed-batch culture runs of 157 unique CHO cell lines across four passage generations, recombinant for three different mAbs, achieving high accuracy in biomass and mAb titre (average ${rm{bar{R}}}_{{{{rm{X}}}_{rm{v}}}}^2$ ≈ 0.96 ± 0.07 and ${rm{bar{R}}}_{rm{P}}^2$ ≈ 0.97 ± 0.05, respectively). MCKM could facilitate automated cell line selection, identification of critical process parameters and biomarkers, guide media and feeding strategies, predict metabolite profiles, and support scale-up and quality-by-design studies, delivering overall reduction of experimental workload. One-Sentence Summary: This paper presents a novel kinetic model that derives distinct parameter sets from a single fed-batch run, enabling characterization of individual CHO clones across different mAb targets.</p>","PeriodicalId":16092,"journal":{"name":"Journal of Industrial Microbiology & Biotechnology","volume":" ","pages":""},"PeriodicalIF":3.2,"publicationDate":"2024-12-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12486387/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145113155","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Development of Modular Expression Across Phylogenetically Distinct Diazotrophs","authors":"Shawn Kulakowski, Alex Rivier, Rita Kuo, Sonya Mengel, Thomas Eng","doi":"10.1093/jimb/kuae033","DOIUrl":"https://doi.org/10.1093/jimb/kuae033","url":null,"abstract":"Diazotrophic bacteria can reduce atmospheric nitrogen into ammonia enabling bioavailability of the essential element. Many diazotrophs closely associate with plant roots increasing nitrogen availability, acting as plant growth promoters. These associations have the potential to reduce the need for costly synthetic fertilizers if they could be engineered for agricultural applications. However, despite the importance of diazotrophic bacteria, genetic tools are poorly developed in a limited number of species, in turn narrowing the crops and root microbiomes that can be targeted. Here we report optimized protocols and plasmids to manipulate phylogenetically diverse diazotrophs with the goal of enabling synthetic biology and genetic engineering. Three broad-host-range plasmids can be used across multiple diazotrophs, with the identification of one specific plasmid (containing origin of replication RK2 and a kanamycin resistance marker) showing the highest degree of compatibility across bacteria tested. We then demonstrated modular expression by testing seven promoters and eleven ribosomal binding sites using proxy fluorescent proteins. Finally, we tested four small molecule inducible systems to report expression in three diazotrophs and demonstrated genome editing in Klebsiella michiganensis M5al.","PeriodicalId":16092,"journal":{"name":"Journal of Industrial Microbiology & Biotechnology","volume":"4 1","pages":""},"PeriodicalIF":3.4,"publicationDate":"2024-09-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142205949","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Simultaneous enumeration of yeast and bacterial cells in the context of industrial bioprocesses","authors":"Carolina Teixeira Martins, Ana Paula Jacobus, Renilson Conceição, Douglas Fernandes Barbin, Helena Bolini, Andreas Karoly Gombert","doi":"10.1093/jimb/kuae029","DOIUrl":"https://doi.org/10.1093/jimb/kuae029","url":null,"abstract":"In scenarios where yeast and bacterial cells coexist, it is of interest to simultaneously quantify the concentrations of both cell types, since traditional methods used to determine these concentrations individually take more time and resources. Here, we compared different methods for quantifying the fuel ethanol Saccharomyces cerevisiae PE-2 yeast strain and cells from the probiotic Lactiplantibacillus plantarum strain in microbial suspensions. Individual suspensions were prepared, mixed in 1:1 or 100:1 yeast-to-bacteria ratios, covering the range typically encountered in sugarcane biorefineries, and analyzed using bright field microscopy, manual and automatic Spread-plate and Drop-plate counting, flow cytometry (at 1:1 and 100:1 ratios), and a Coulter Counter (at 1:1 and 100:1 ratios). We observed that for yeast cell counts in the mixture (1:1 and 100:1 ratios), flow cytometry, the Coulter Counter, and both Spread-plate options (manual and automatic CFU counting) yielded statistically similar results, while the Drop-plate and microscopy-based methods gave statistically different results. For bacterial cell quantification, the microscopy-based method, Drop-plate, and both Spread-plate plating options and flow cytometry (1:1 ratio) produced no significantly different results (p &amp;gt; .05). In contrast, the Coulter Counter (1:1 ratio) and flow cytometry (100:1 ratio) presented results statistically different (p &amp;lt; .05). Additionally, quantifying bacterial cells in a mixed suspension at a 100:1 ratio wasn't possible due to an overlap between yeast cell debris and bacterial cells. We conclude that each method has limitations, advantages, and disadvantages. One-Sentence Summary This study compares methods for simultaneously quantifying yeast and bacterial cells in a mixed sample, highlighting that in different cell proportions, some methods cannot quantify both cell types and present distinct advantages and limitations regarding time, cost, and precision.","PeriodicalId":16092,"journal":{"name":"Journal of Industrial Microbiology & Biotechnology","volume":"57 1","pages":""},"PeriodicalIF":3.4,"publicationDate":"2024-09-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142205852","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Multi-modular metabolic engineering and efflux engineering for enhanced lycopene production in recombinant Saccharomyces cerevisiae","authors":"Guangxi Huang, Jiarong Li, Jingyuan Lin, Changqing Duan, Guoliang Yan","doi":"10.1093/jimb/kuae015","DOIUrl":"https://doi.org/10.1093/jimb/kuae015","url":null,"abstract":"Lycopene has been widely used in the food industry and medical field due to its antioxidant, anti-cancer, and anti-inflammatory properties. However, achieving efficient manufacture of lycopene using chassis cells on an industrial scale remains a major challenge. Herein, we attempted to integrate multiple metabolic engineering strategies to establish an efficient and balanced lycopene biosynthetic system in Saccharomyces cerevisiae. First, the lycopene synthesis pathway was modularized to sequentially enhance the metabolic flux of the Mevalonate pathway, the acetyl-CoA supply module, and lycopene exogenous enzymatic module. The modular operation enabled the efficient conversion of acetyl-CoA to downstream pathway of lycopene synthesis, resulting in a 3.1-fold increase of lycopene yield. Second, we introduced acetate as an exogenous carbon source and utilized an acetate-repressible promoter to replace the natural ERG9 promoter. This approach not only enhanced the supply of acetyl-CoA but also concurrently diminished the flux towards the competitive ergosterol pathway. As a result, a further 42.3% increase in lycopene production was observed. Third, we optimized NADPH supply and mitigated cytotoxicity by overexpressing ABC transporters to promote lycopene efflux. The obtained strain YLY-PDR11 showed a 12.7-fold increase in extracellular lycopene level compared to the control strain. Finally, the total lycopene yield reached 343.7mg/L, which was 4.3 times higher than that of the initial strain YLY-04. Our results demonstrate that combining multi-modular metabolic engineering with efflux engineering is an effective approach to improve the production of lycopene. This strategy can also be applied to the overproduction of other desirable isoprenoid compounds with similar synthesis and storage patterns in S. cerevisiae.","PeriodicalId":16092,"journal":{"name":"Journal of Industrial Microbiology & Biotechnology","volume":"414 1","pages":""},"PeriodicalIF":3.4,"publicationDate":"2024-04-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140584408","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Application of the surface engineered recombinant Escherichia coli to the industrial battery waste solution for lithium recovery","authors":"Jaehoon Jeong, Vidhya Selvamani, Murali kannan Maruthamuthu, Kulandaisamy Arulsamy, Soon Ho Hong","doi":"10.1093/jimb/kuae012","DOIUrl":"https://doi.org/10.1093/jimb/kuae012","url":null,"abstract":"Escherichia coli were engineered to selectively adsorb and recover lithium from the environment by employing bacterial cell surface display strategy. Lithium binding peptide LBP1 was integrated to the Escherichia coli membrane protein OmpC. The effect of environmental conditions on the adsorption of lithium by recombinant strain was evaluated, and lithium particles on cellular surface was analysed by FE-SEM and XRD. To elevate the lithium adsorption, dimeric, trimeric and tetrameric repeats of the LBP1 peptide was constructed and displayed on the surface of E. coli. The constructed recombinant E. coli displaying LBP1 trimer was applied to real industrial lithium battery wastewater to recover lithium.","PeriodicalId":16092,"journal":{"name":"Journal of Industrial Microbiology & Biotechnology","volume":"35 1","pages":""},"PeriodicalIF":3.4,"publicationDate":"2024-04-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140584320","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Harnessing regulatory networks in Actinobacteria for natural product discovery","authors":"Hannah E Augustijn, Anna M Roseboom, Marnix H Medema, Gilles P van Wezel","doi":"10.1093/jimb/kuae011","DOIUrl":"https://doi.org/10.1093/jimb/kuae011","url":null,"abstract":"Microbes typically live in complex habitats where they need to rapidly adapt to continuously changing growth conditions. To do so, they produce an astonishing array of natural products with diverse structures and functions. Actinobacteria stand out for their prolific production of bioactive molecules, including antibiotics, anticancer agents, antifungals, and immunosuppressants. Attention has been directed especially towards the identification of the compounds they produce and the mining of the large diversity of biosynthetic gene clusters (BGCs) in their genomes. However, the current return on investment in random screening for bioactive compounds is low, while it is hard to predict which of the millions of BGCs should be prioritized. Moreover, many of the BGCs for yet undiscovered natural products are silent or cryptic under laboratory growth conditions. To identify ways to prioritize and activate these BGCs, knowledge regarding the way their expression is controlled is crucial. Intricate regulatory networks control global gene expression in Actinobacteria, governed by a staggering number of up to 1000 transcription factors per strain. This review highlights recent advances in experimental and computational methods for characterizing and predicting transcription factor binding sites and their applications to guide natural product discovery. We propose that regulation-guided genome mining approaches will open new avenues toward eliciting the expression of BGCs, as well as prioritizing subsets of BGCs for expression using synthetic biology approaches. One-Sentence Summary This review provides insights into advances in experimental and computational methods aimed at predicting transcription factor binding sites and their applications to guide natural product discovery.","PeriodicalId":16092,"journal":{"name":"Journal of Industrial Microbiology & Biotechnology","volume":"22 1","pages":""},"PeriodicalIF":3.4,"publicationDate":"2024-04-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140584317","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"One-pot chemoenzymatic syntheses of non-canonical amino acids.","authors":"Tsung-Han Chao, Xiangyu Wu, Hans Renata","doi":"10.1093/jimb/kuae005","DOIUrl":"10.1093/jimb/kuae005","url":null,"abstract":"<p><p>Despite their prevalent use in drug discovery and protein biochemistry, non-canonical amino acids are still challenging to synthesize through purely chemical means. In recent years, biocatalysis has emerged as a transformative paradigm for small-molecule synthesis. One strategy to further empower biocatalysis is to use it in combination with modern chemical reactions and take advantage of the strengths of each method to enable access to challenging structural motifs that were previously unattainable using each method alone. In this Mini-Review, we highlight several recent case studies that feature the synergistic use of chemical and enzymatic transformations in one pot to synthesize novel non-canonical amino acids.</p><p><strong>One-sentence summary: </strong>This Mini-Review highlights several recent case studies that feature the synergistic use of chemical and enzymatic transformations in one pot to synthesize novel non-canonical amino acids.</p>","PeriodicalId":16092,"journal":{"name":"Journal of Industrial Microbiology & Biotechnology","volume":" ","pages":""},"PeriodicalIF":3.4,"publicationDate":"2024-01-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10853765/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139564289","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Performance evaluation of a low-throughput qPCR-based Legionella assay for utility as an onsite industrial water system monitoring method.","authors":"Alexsandra Corrigan, Benjamin Niemaseck, Mackenzie Moore, Douglas McIlwaine, Jeremy Duguay","doi":"10.1093/jimb/kuae030","DOIUrl":"10.1093/jimb/kuae030","url":null,"abstract":"<p><p>Legionella is a bacterial genus found in natural aquatic environments, as well as domestic and industrial water systems. Legionella presents potential human health risks when aerosolized and inhaled by at-risk individuals and is commonly monitored at locations with likelihood of proliferation and human exposure. Legionella monitoring is widely performed using culture-based testing, which faces limitations including turnaround time and interferences. Molecular biology methodologies, including quantitative polymerase chain reaction (qPCR), are being explored to supplement or replace culture-based testing because of faster turnaround and lower detection limits, allowing for more rapid water remediation measures. In this study, three methods were compared by testing industrial water samples: culture-based testing by a certified lab, high throughput qPCR testing (HT qPCR), and field deployable low throughput qPCR testing (LT qPCR). The qPCR test methods reported more positive results than culture testing, indicating improved sensitivity and specificity. The LT qPCR test is portable with quick turnaround times, and can be leveraged for environmental surveillance, process optimization, monitoring, and onsite case investigations. The LT qPCR test had high negative predictive value and would be a useful tool for negative screening of Legionella samples from high-risk environments and/or outbreak investigations to streamline samples for culture testing.</p><p><strong>One-sentence summary: </strong>This study compared three test methods for Legionella to evaluate performance of a low throughput quantitative polymerase chain reaction (LT qPCR) test for Legionella that can be used onsite; the study found that the high throughput (HT) and LT qPCR tests used in this study gave more positive results than culture testing, and the results indicated a similar negative predictive value for the HT and LT qPCR tests, supporting that the LT qPCR method could be useful for negative screening of Legionella samples in industrial water systems onsite.</p>","PeriodicalId":16092,"journal":{"name":"Journal of Industrial Microbiology & Biotechnology","volume":" ","pages":""},"PeriodicalIF":3.2,"publicationDate":"2024-01-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11388924/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142008897","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}