Engineering Microbiology最新文献

{"title":"Recombineering enables genome mining of novel siderophores in a non-model Burkholderiales strain","authors":"Xingyan Wang,&nbsp;Haibo Zhou,&nbsp;Xiangmei Ren,&nbsp;Hanna Chen,&nbsp;Lin Zhong,&nbsp;Xianping Bai,&nbsp;Xiaoying Bian","doi":"10.1016/j.engmic.2023.100106","DOIUrl":"https://doi.org/10.1016/j.engmic.2023.100106","url":null,"abstract":"<div><p>Iron is essential for bacterial survival, and most bacteria capture iron by producing siderophores. <em>Burkholderiales</em> bacteria produce various types of bioactive secondary metabolites, such as ornibactin and malleobactin siderophores. In this study, the genome analysis of <em>Burkholderiales</em> genomes showed a putative novel siderophore gene cluster <em>crb</em>, which is highly similar to the ornibactin and malleobactin gene clusters but does not have <em>pvdF</em>, a gene encoding a formyltransferase for N-<em>δ</em>‑hydroxy-ornithine formylation. Establishing the bacteriophage recombinase Redγ-Redαβ7029 mediated genome editing system in a non-model <em>Burkholderiales</em> strain <em>Paraburkholderia caribensis</em> CICC 10960 allowed the rapid identification of the products of <em>crb</em> gene cluster, caribactins A-F (<strong>1–6</strong>). Caribactins contain a special amino acid residue N-<em>δ</em>‑hydroxy-N-<em>δ</em>-acetylornithine (haOrn), which differs from the counterpart N-<em>δ</em>‑hydroxy-N-<em>δ</em>-formylornithine (hfOrn) in ornibactin and malleobactin, owing to the absence of <em>pvdF</em>. Gene inactivation showed that the acetylation of hOrn is catalyzed by CrbK, whose homologs probably not be involved in the biosynthesis of ornibactin and malleobactin, showing possible evolutionary clues of these siderophore biosynthetic pathways from different genera. Caribactins promote biofilm production and enhance swarming and swimming abilities, suggesting that they may play crucial roles in biofilm formation. This study also revealed that recombineering has the capability to mine novel secondary metabolites from non-model <em>Burkholderiales</em> species.</p></div>","PeriodicalId":100478,"journal":{"name":"Engineering Microbiology","volume":"3 3","pages":"Article 100106"},"PeriodicalIF":0.0,"publicationDate":"2023-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"49891302","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Recent advances in the direct cloning of large natural product biosynthetic gene clusters","authors":"Jiaying Wan,&nbsp;Nan Ma,&nbsp;Hua Yuan","doi":"10.1016/j.engmic.2023.100085","DOIUrl":"https://doi.org/10.1016/j.engmic.2023.100085","url":null,"abstract":"<div><p>Large-scale genome-mining analyses have revealed that microbes potentially harbor a huge reservoir of uncharacterized natural product (NP) biosynthetic gene clusters (<em>BGCs</em>), and this has spurred a renaissance of novel drug discovery. However, the majority of these <em>BGCs</em> are often poorly or not at all expressed in their native hosts under laboratory conditions, and thus are regarded as silent/orphan <em>BGCs</em>. Currently, connecting silent <em>BGCs</em> to their corresponding NPs quickly and on a large scale is particularly challenging because of the lack of universal strategies and enabling technologies. Generally, the heterologous host-based genome mining strategy is believed to be a suitable alternative to the native host-based approach for prioritization of the vast and ever-increasing number of uncharacterized <em>BGCs</em>. In the last ten years, a variety of methods have been reported for the direct cloning of <em>BGCs</em> of interest, which is the first and rate-limiting step in the heterologous expression strategy. Essentially, each method requires that the following three issues be resolved: 1) how to prepare genomic <em>DNA</em>; 2) how to digest the bilateral boundaries for release of the target <em>BGC</em>; and 3) how to assemble the <em>BGC</em> and the capture vector. Here, we summarize recent reports regarding how to directly capture a <em>BGC</em> of interest and briefly discuss the advantages and disadvantages of each method, with an emphasis on the notion that direct cloning is very beneficial for accelerating genome mining research and large-scale drug discovery.</p></div>","PeriodicalId":100478,"journal":{"name":"Engineering Microbiology","volume":"3 3","pages":"Article 100085"},"PeriodicalIF":0.0,"publicationDate":"2023-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"49891304","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Genome-scale CRISPRi screening: A powerful tool in engineering microbiology","authors":"Letian Sun ,&nbsp;Ping Zheng ,&nbsp;Jibin Sun ,&nbsp;Volker F. Wendisch ,&nbsp;Yu Wang","doi":"10.1016/j.engmic.2023.100089","DOIUrl":"https://doi.org/10.1016/j.engmic.2023.100089","url":null,"abstract":"<div><p>Deciphering gene function is fundamental to engineering of microbiology. The clustered regularly interspaced short palindromic repeats (CRISPR) system has been adapted for gene repression across a range of hosts, creating a versatile tool called CRISPR interference (CRISPRi) that enables genome-scale analysis of gene function. This approach has yielded significant advances in the design of genome-scale CRISPRi libraries, as well as in applications of CRISPRi screening in medical and industrial microbiology. This review provides an overview of the recent progress made in pooled and arrayed CRISPRi screening in microorganisms and highlights representative studies that have employed this method. Additionally, the challenges associated with CRISPRi screening are discussed, and potential solutions for optimizing this strategy are proposed.</p></div>","PeriodicalId":100478,"journal":{"name":"Engineering Microbiology","volume":"3 3","pages":"Article 100089"},"PeriodicalIF":0.0,"publicationDate":"2023-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"49891306","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Profiling proteomic responses to hexokinase-II depletion in terpene-producing Saccharomyces cerevisiae","authors":"Zeyu Lu ,&nbsp;Qianyi Shen ,&nbsp;Lian Liu ,&nbsp;Gert Talbo ,&nbsp;Robert Speight ,&nbsp;Matt Trau ,&nbsp;Geoff Dumsday ,&nbsp;Christopher B. Howard ,&nbsp;Claudia E. Vickers ,&nbsp;Bingyin Peng","doi":"10.1016/j.engmic.2023.100079","DOIUrl":"https://doi.org/10.1016/j.engmic.2023.100079","url":null,"abstract":"<div><p>Hexokinase II (Hxk2) is a master protein in glucose-mediated transcriptional repression signaling pathway. Degrading Hxk2 through an auxin-inducible protein degradation previously doubled sesquiterpene (nerolidol) production at gram-per-liter levels in <em>Saccharomyces cerevisiae</em>. Global transcriptomics/proteomics profiles in Hxk2-deficient background are important to understanding genetic and molecular mechanisms for improved nerolidol production and guiding further strain optimization. Here, proteomic responses to Hxk2 depletion are investigated in the yeast strains harboring a <em>GAL</em> promoters-controlled nerolidol synthetic pathway, at the exponential and ethanol growth phases and in <em>GAL80</em>-wildtype and <em>gal80Δ</em> backgrounds. Carbon metabolic pathways and amino acid metabolic pathways show diversified responses to Hxk2 depletion and growth on ethanol, including upregulation of alternative carbon catabolism and respiration as well as downregulation of amino acid synthesis. De-repression of <em>GAL</em> genes may contribute to improved nerolidol production in Hxk2-depleted strains. Seventeen transcription factors associated with upregulated genes are enriched. Validating Ash1-mediated repression on the <em>RIM4</em> promoter shows the variation on the regulatory effects of different Ash1-binding sites and the synergistic effect of Ash1 and Hxk2-mediated repression. Further validation of individual promoters shows that <em>HXT1</em> promoter activities are glucose-dependent in <em>hxk2Δ</em> background, but much weaker than those in <em>HXK2</em>-wildtype background. In summary, inactivating <em>HXK2</em> may relieve glucose repression on respiration and <em>GAL</em> promoters for improved bioproduction under aerobic conditions in <em>S. cerevisiae</em>. The proteomics profiles provide a better genetics overview for a better metabolic engineering design in <em>Hxk2-deficient</em> backgrounds.</p></div>","PeriodicalId":100478,"journal":{"name":"Engineering Microbiology","volume":"3 3","pages":"Article 100079"},"PeriodicalIF":0.0,"publicationDate":"2023-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"49891366","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Metabolic engineering strategies for microbial utilization of methanol","authors":"Yamei Gan ,&nbsp;Xin Meng ,&nbsp;Cong Gao ,&nbsp;Wei Song ,&nbsp;Liming Liu ,&nbsp;Xiulai Chen","doi":"10.1016/j.engmic.2023.100081","DOIUrl":"https://doi.org/10.1016/j.engmic.2023.100081","url":null,"abstract":"<div><p>The increasing shortage of fossil resources and environmental pollution has renewed interest in the synthesis of value-added biochemicals from methanol. However, most of native or synthetic methylotrophs are unable to assimilate methanol at a sufficient rate to produce biochemicals. Thus, the performance of methylotrophs still needs to be optimized to meet the demands of industrial applications. In this review, we provide an in-depth discussion on the properties of natural and synthetic methylotrophs, and summarize the natural and synthetic methanol assimilation pathways. Further, we discuss metabolic engineering strategies for enabling microbial utilization of methanol for the bioproduction of value-added chemicals. Finally, we highlight the potential of microbial engineering for methanol assimilation and offer guidance for achieving a low-carbon footprint for the biosynthesis of chemicals.</p></div>","PeriodicalId":100478,"journal":{"name":"Engineering Microbiology","volume":"3 3","pages":"Article 100081"},"PeriodicalIF":0.0,"publicationDate":"2023-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"49891303","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The emerging role of recombineering in microbiology","authors":"Ruijuan Li ,&nbsp;Aiying Li ,&nbsp;Youming Zhang ,&nbsp;Jun Fu","doi":"10.1016/j.engmic.2023.100097","DOIUrl":"https://doi.org/10.1016/j.engmic.2023.100097","url":null,"abstract":"<div><p>Recombineering is a valuable technique for generating recombinant DNA <em>in vivo</em>, primarily in bacterial cells, and is based on homologous recombination using phage-encoded homologous recombinases, such as Redαβγ from the lambda phage and RecET from the Rac prophage. The recombineering technique can efficiently mediate homologous recombination using short homologous arms (∼50 bp) and is unlimited by the size of the DNA molecules or positions of restriction sites. In this review, we summarize characteristics of recombinases, mechanism of recombineering, and advances in recombineering for DNA manipulation in <em>Escherichia coli</em> and other bacteria. Furthermore, the broad applications of recombineering for mining new bioactive microbial natural products, and for viral mutagenesis, phage genome engineering, and understanding bacterial metabolism are also reviewed.</p></div>","PeriodicalId":100478,"journal":{"name":"Engineering Microbiology","volume":"3 3","pages":"Article 100097"},"PeriodicalIF":0.0,"publicationDate":"2023-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"49891307","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Catalog of operational taxonomic units and unified amplicon sequencing data for the microbiomes of medicinal plant roots","authors":"Meng Wang ,&nbsp;Ming Lei ,&nbsp;Hailun He","doi":"10.1016/j.engmic.2023.100087","DOIUrl":"https://doi.org/10.1016/j.engmic.2023.100087","url":null,"abstract":"<div><p>China has a rich history of cultivating medicinal plants, whose root microbial communities closely interact with the medicinal plants, thereby influencing their growth, health, and medicinal properties. Currently, researchers widely use 16S rRNA gene amplicon sequencing to study these root microbial communities. However, publicly available sequence datasets often lack essential sample information or contain errors, impeding the reuse of the datasets in the future. In this study, we aimed to create a united, reliable, and readily usable source of 16S rRNA gene sequences for medicinal plant root microbiomes. We compiled a catalog of 1392 microbiome samples for 58 medicinal plants from 58 studies, and manually provided essential sample information based on the experimental setup described in the associated papers. We then processed the sequences using a custom pipeline, generating a united catalog of operational taxonomic units (OTUs) and conducting taxonomic classification. We also predicted the ecological functions of the communities for each sample. Finally, we used this dataset, to compare the rhizosphere bacterial communities of <em>Pseudostellaria heterophylla</em> from Fujian and Guizhou Provinces, revealing significant differences in the community composition of the same plant from different geographic locations. By providing a comprehensive and united catalog of amplicon sequences and OTUs for medicinal plant root bacterial communities, this study offers an invaluable resource for future comparative studies and data mining.</p></div>","PeriodicalId":100478,"journal":{"name":"Engineering Microbiology","volume":"3 3","pages":"Article 100087"},"PeriodicalIF":0.0,"publicationDate":"2023-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"49891367","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Engineering of Saccharomyces cerevisiae for co-fermentation of glucose and xylose: Current state and perspectives","authors":"Yali Qiu ,&nbsp;Meiling Wu ,&nbsp;Haodong Bao ,&nbsp;Weifeng Liu ,&nbsp;Yu Shen","doi":"10.1016/j.engmic.2023.100084","DOIUrl":"https://doi.org/10.1016/j.engmic.2023.100084","url":null,"abstract":"<div><p>The use of non-food lignocellulosic biomass to produce ethanol fits into the strategy of a global circular economy with low dependence on fossil energy resources. Xylose is the second most abundant sugar in lignocellulosic hydrolysate, and its utilization in fermentation is a key issue in making the full use of raw plant materials for ethanol production and reduce production costs. <em>Saccharomyces cerevisiae</em> is the best ethanol producer but the organism is not a native xylose user. In recent years, great efforts have been made in the construction of xylose utilizing <em>S. cerevisiae</em> strains by metabolic and evolutionary engineering approaches. In addition, managing global transcriptional regulation works provides an effective means to increase the xylose utilization capacity of recombinant strains. Here we review the common strategies and research advances in the research field in order to facilitate the researches in xylose metabolism and xylose-based fermentation.</p></div>","PeriodicalId":100478,"journal":{"name":"Engineering Microbiology","volume":"3 3","pages":"Article 100084"},"PeriodicalIF":0.0,"publicationDate":"2023-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"49891305","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A recombineering system for Bacillus subtilis based on the native phage recombinase pair YqaJ/YqaK","authors":"Qingshu Liu ,&nbsp;Ruijuan Li ,&nbsp;Hongbo Shi ,&nbsp;Runyu Yang ,&nbsp;Qiyao Shen ,&nbsp;Qingwen Cui ,&nbsp;Xiuling Wang ,&nbsp;Aiying Li ,&nbsp;Youming Zhang ,&nbsp;Jun Fu","doi":"10.1016/j.engmic.2023.100099","DOIUrl":"https://doi.org/10.1016/j.engmic.2023.100099","url":null,"abstract":"<div><p><em>Bacillus subtilis</em> plays an important role in fundamental and applied research, and it has been widely used as a cell factory for the production of enzymes, antimicrobial materials, and chemicals for agriculture, medicine, and industry. However, genetic manipulation tools for <em>B. subtilis</em> have low efficiency. In this work, our goal was to develop a simple recombineering system for <em>B. subtilis</em>. We showed that genome editing can be achieved in <em>B. subtiliis</em> 1A751 through co-expression of YqaJ/YqaK, a native phage recombinase pair found in <em>B. subtilis</em> 168, and the competence master regulator ComK using a double-stranded DNA substrate with short homology arms (100 bp) and a phosphorothioate modification at the 5′-end. Efficient gene knockouts and large DNA insertions were achieved using this new recombineering system in <em>B. subtilis</em> 1A751. As far as we know, this is the first recombineering system using the native phage recombinase pair YqaJ/YqaK in <em>B. subtilis</em>. In conclusion, this new recombineering system provides a simple and fast tool for genetic manipulation of <em>B. subtilis</em>, and it will promote studies of genome function, construction of production strains, and genome mining in <em>B. subtilis</em>.</p></div>","PeriodicalId":100478,"journal":{"name":"Engineering Microbiology","volume":"3 3","pages":"Article 100099"},"PeriodicalIF":0.0,"publicationDate":"2023-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"49891368","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The lasso structure, biosynthesis, bioactivities and potential applications of Microcin J25: A novel antibacterial agent with unique mechanisms","authors":"Qingchun Ji ,&nbsp;Bixia Zhou ,&nbsp;Tong Shen ,&nbsp;Tianyue Jiang ,&nbsp;Cheng Cheng ,&nbsp;Bingfang He","doi":"10.1016/j.engmic.2023.100096","DOIUrl":"https://doi.org/10.1016/j.engmic.2023.100096","url":null,"abstract":"<div><p>The overuse and misuse of traditional antimicrobial drugs have led to their weakened effectiveness and the emergence of pathogenic bacterial resistance. Consequently, there has been growing interest in alternative options such as antimicrobial peptides (AMPs) in the pharmaceutical industry. Microcin J25 (MccJ25) has gained significant attention for its potent inhibitory effect on a diverse range of pathogens. Its unique rotaxane structure provides exceptional stability against extreme thermal, pH, and protease degradation, including chymotrypsin, trypsin, and pepsin. Given its remarkable stability and diverse bioactivity, we aim to provide an overview of the physicochemical properties, the mechanism underlying its antimicrobial activity, and the critical functional residues of MccJ25. Additionally, we have summarized the latest strategies for the heterologous expression of MccJ25, and its potential medical use and other applications.</p></div>","PeriodicalId":100478,"journal":{"name":"Engineering Microbiology","volume":"3 3","pages":"Article 100096"},"PeriodicalIF":0.0,"publicationDate":"2023-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"49891308","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}