Metabolic Engineering Communications最新文献

{"title":"Itaconic acid production is regulated by LaeA in Aspergillus pseudoterreus","authors":"Kyle R. Pomraning ,&nbsp;Ziyu Dai ,&nbsp;Nathalie Munoz ,&nbsp;Young-Mo Kim ,&nbsp;Yuqian Gao ,&nbsp;Shuang Deng ,&nbsp;Teresa Lemmon ,&nbsp;Marie S. Swita ,&nbsp;Jeremy D. Zucker ,&nbsp;Joonhoon Kim ,&nbsp;Stephen J. Mondo ,&nbsp;Ellen Panisko ,&nbsp;Meagan C. Burnet ,&nbsp;Bobbie-Jo M. Webb-Robertson ,&nbsp;Beth Hofstad ,&nbsp;Scott E. Baker ,&nbsp;Kristin E. Burnum-Johnson ,&nbsp;Jon K. Magnuson ,&nbsp;for the Agile BioFoundry","doi":"10.1016/j.mec.2022.e00203","DOIUrl":"10.1016/j.mec.2022.e00203","url":null,"abstract":"<div><p>The global regulator LaeA controls secondary metabolism in diverse Aspergillus species. Here we explored its role in regulation of itaconic acid production in <em>Aspergillus pseudoterreus</em>. To understand its role in regulating metabolism, we deleted and overexpressed <em>laeA,</em> and assessed the transcriptome, proteome, and secreted metabolome prior to and during initiation of phosphate limitation induced itaconic acid production. We found that secondary metabolite clusters, including the itaconic acid biosynthetic gene cluster, are regulated by <em>laeA</em> and that <em>laeA</em> is required for high yield production of itaconic acid. Overexpression of LaeA improves itaconic acid yield at the expense of biomass by increasing the expression of key biosynthetic pathway enzymes and attenuating the expression of genes involved in phosphate acquisition and scavenging. Increased yield was observed in optimized conditions as well as conditions containing excess nutrients that may be present in inexpensive sugar containing feedstocks such as excess phosphate or complex nutrient sources. This suggests that global regulators of metabolism may be useful targets for engineering metabolic flux that is robust to environmental heterogeneity.</p></div>","PeriodicalId":18695,"journal":{"name":"Metabolic Engineering Communications","volume":"15 ","pages":"Article e00203"},"PeriodicalIF":5.2,"publicationDate":"2022-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/4e/ad/main.PMC9440423.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40352282","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":"Development of a dedicated Golden Gate Assembly Platform (RtGGA) for Rhodotorula toruloides","authors":"Nemailla Bonturi ,&nbsp;Marina Julio Pinheiro ,&nbsp;Paola Monteiro de Oliveira ,&nbsp;Eka Rusadze ,&nbsp;Tobias Eichinger ,&nbsp;Gintare Liudžiūtė ,&nbsp;Juliano Sabedotti De Biaggi ,&nbsp;Age Brauer ,&nbsp;Maido Remm ,&nbsp;Everson Alves Miranda ,&nbsp;Rodrigo Ledesma-Amaro ,&nbsp;Petri-Jaan Lahtvee","doi":"10.1016/j.mec.2022.e00200","DOIUrl":"10.1016/j.mec.2022.e00200","url":null,"abstract":"<div><p><em>Rhodotorula toruloides</em> is a potential chassis for microbial cell factories as this yeast can metabolise different substrates into a diverse range of natural products, but the lack of efficient synthetic biology tools hinders its applicability. In this study, the modular, versatile and efficient Golden Gate DNA assembly system (RtGGA) was adapted to the first basidiomycete, an oleaginous yeast <em>R. toruloides</em>. <em>R. toruloides</em> CCT 0783 was sequenced, and used for the GGA design. The DNA fragments were assembled with predesigned 4-nt overhangs and a library of standardized parts was created containing promoters, genes, terminators, insertional regions, and resistance genes. The library was combined to create cassettes for the characterization of promoters strength and to overexpress the carotenoid production pathway. A variety of reagents, plasmids, and strategies were used and the RtGGA proved to be robust. The RtGGA was used to build three versions of the carotenoid overexpression cassette by using different promoter combinations. The cassettes were transformed into <em>R. toruloides</em> and the three new strains were characterized. Total carotenoid concentration increased by 41%. The dedicated GGA platform fills a gap in the advanced genome engineering toolkit for <em>R. toruloides</em>, enabling the efficient design of complex metabolic pathways.</p></div>","PeriodicalId":18695,"journal":{"name":"Metabolic Engineering Communications","volume":"15 ","pages":"Article e00200"},"PeriodicalIF":5.2,"publicationDate":"2022-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S2214030122000098/pdfft?md5=a2fe424d543e5912a0eacd9cd6daa4d6&pid=1-s2.0-S2214030122000098-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"46506584","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":"Tuning a high performing multiplexed-CRISPRi Pseudomonas putida strain to further enhance indigoidine production","authors":"Jeffrey J. Czajka ,&nbsp;Deepanwita Banerjee ,&nbsp;Thomas Eng ,&nbsp;Javier Menasalvas ,&nbsp;Chunsheng Yan ,&nbsp;Nathalie Munoz Munoz ,&nbsp;Brenton C. Poirier ,&nbsp;Young-Mo Kim ,&nbsp;Scott E. Baker ,&nbsp;Yinjie J. Tang ,&nbsp;Aindrila Mukhopadhyay","doi":"10.1016/j.mec.2022.e00206","DOIUrl":"10.1016/j.mec.2022.e00206","url":null,"abstract":"<div><p>In this study, a 14-gene edited <em>Pseudomonas putida</em> KT2440 strain for heterologous indigoidine production was examined using three distinct omic datasets. Transcriptomic data indicated that CRISPR/dCpf1-interference (CRISPRi) mediated multiplex repression caused global gene expression changes, implying potential undesirable changes in metabolic flux. ¹³C-metabolic flux analysis (¹³C-MFA) revealed that the core <em>P. putida</em> flux network after CRISPRi repression was conserved, with moderate reduction of TCA cycle and pyruvate shunt activity along with glyoxylate shunt activation during glucose catabolism. Metabolomic results identified a change in intracellular TCA metabolites and extracellular metabolite secretion profiles (sugars and succinate overflow) in the engineered strains. These omic analyses guided further strain engineering, with a random mutagenesis screen first identifying an optimal ribosome binding site (RBS) for Cpf1 that enabled stronger product-substrate pairing (1.6–fold increase). Then, deletion strains were constructed with excision of the PHA operon (Δ<em>phaAZC-IID</em>) resulting in a 2.2–fold increase in indigoidine titer over the optimized Cpf1-RBS construct at the end of the growth phase (∼6 h). The maximum indigoidine titer (at 72 h) in the Δ<em>phaAZC-IID</em> strain had a 1.5–fold and 1.8–fold increase compared to the optimized Cpf1-RBS construct and the original strain, respectively. Overall, this study demonstrated that integration of omic data types is essential for understanding responses to complex metabolic engineering designs and directly quantified the effect of such modifications on central metabolism.</p></div>","PeriodicalId":18695,"journal":{"name":"Metabolic Engineering Communications","volume":"15 ","pages":"Article e00206"},"PeriodicalIF":5.2,"publicationDate":"2022-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/15/32/main.PMC9494242.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"33482926","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":"RNA polymerase II-driven CRISPR-Cas9 system for efficient non-growth-biased metabolic engineering of Kluyveromyces marxianus","authors":"Danielle Bever ,&nbsp;Ian Wheeldon ,&nbsp;Nancy Da Silva","doi":"10.1016/j.mec.2022.e00208","DOIUrl":"https://doi.org/10.1016/j.mec.2022.e00208","url":null,"abstract":"<div><p>The thermotolerant yeast <em>Kluyveromyces marxianus</em> has gained significant attention in recent years as a promising microbial candidate for industrial biomanufacturing. Despite several contributions to the expanding molecular toolbox for gene expression and metabolic engineering of <em>K. marxianus</em>, there remains a need for a more efficient and versatile genome editing platform. To address this, we developed a CRISPR-based editing system that enables high efficiency marker-less gene disruptions and integrations using only 40 bp homology arms in NHEJ functional and non-functional <em>K. marxianus</em> strains. The use of a strong RNA polymerase II promoter allows efficient expression of gRNAs flanked by the self-cleaving RNA structures, tRNA and HDV ribozyme, from a single plasmid co-expressing a codon optimized Cas9. Implementing this system resulted in nearly 100% efficiency of gene disruptions in both NHEJ-functional and NHEJ-deficient <em>K. marxianus</em> strains, with donor integration efficiencies reaching 50% and 100% in the two strains, respectively. The high gRNA targeting performance also proved instrumental for selection of engineered strains with lower growth rate but improved polyketide biosynthesis by avoiding an extended outgrowth period, a common method used to enrich for edited cells but that fails to recover advantageous mutants with even slightly impaired fitness. Finally, we provide the first demonstration of simultaneous, markerless integrations at multiple loci in <em>K. marxianus</em> using a 2.6 kb and a 7.6 kb donor, achieving a dual integration efficiency of 25.5% in a NHEJ-deficient strain. These results highlight both the ease of use and general robustness of this system for rapid and flexible metabolic engineering in this non-conventional yeast.</p></div>","PeriodicalId":18695,"journal":{"name":"Metabolic Engineering Communications","volume":"15 ","pages":"Article e00208"},"PeriodicalIF":5.2,"publicationDate":"2022-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S2214030122000177/pdfft?md5=f6be45d36815b9de356945d06bf05598&pid=1-s2.0-S2214030122000177-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"91591437","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":"Genome-scale reconstruction and metabolic modelling of the fast-growing thermophile Geobacillus sp. LC300","authors":"Emil Ljungqvist,&nbsp;Martin Gustavsson","doi":"10.1016/j.mec.2022.e00212","DOIUrl":"https://doi.org/10.1016/j.mec.2022.e00212","url":null,"abstract":"<div><p>Thermophilic microorganisms show high potential for use as biorefinery cell factories. Their high growth temperatures provide fast conversion rates, lower risk of contaminations, and facilitated purification of volatile products. To date, only a few thermophilic species have been utilized for microbial production purposes, and the development of production strains is impeded by the lack of metabolic engineering tools. In this study, we constructed a genome-scale metabolic model, an important part of the metabolic engineering pipeline, of the fast-growing thermophile <em>Geobacillus</em> sp. LC300. The model (iGEL604) contains 604 genes, 1249 reactions and 1311 metabolites, and the reaction reversibility is based on thermodynamics at the optimum growth temperature. The growth phenotype is analyzed by batch cultivations on two carbon sources, further closing balances in carbon and degree-of-reduction. The predictive ability of the model is benchmarked against experimentally determined growth characteristics and internal flux distributions, showing high similarity to experimental phenotypes.</p></div>","PeriodicalId":18695,"journal":{"name":"Metabolic Engineering Communications","volume":"15 ","pages":"Article e00212"},"PeriodicalIF":5.2,"publicationDate":"2022-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S2214030122000219/pdfft?md5=084bfef2d5e8e9c0bac22632181a9aff&pid=1-s2.0-S2214030122000219-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"91640474","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":"Isobutanol production by combined in vivo and in vitro metabolic engineering","authors":"Mamta Gupta ,&nbsp;Matthew Wong ,&nbsp;Kamran Jawed ,&nbsp;Kamil Gedeon ,&nbsp;Hannah Barrett ,&nbsp;Marcelo Bassalo ,&nbsp;Clifford Morrison ,&nbsp;Danish Eqbal ,&nbsp;Syed Shams Yazdani ,&nbsp;Ryan T. Gill ,&nbsp;Jiaqi Huang ,&nbsp;Marc Douaisi ,&nbsp;Jonathan Dordick ,&nbsp;Georges Belfort ,&nbsp;Mattheos A.G. Koffas","doi":"10.1016/j.mec.2022.e00210","DOIUrl":"https://doi.org/10.1016/j.mec.2022.e00210","url":null,"abstract":"<div><p>The production of the biofuel, isobutanol, in <em>E. coli</em> faces limitations due to alcohol toxicity, product inhibition, product recovery, and long-term industrial feasibility. Here we demonstrate an approach of combining both <em>in vivo</em> with <em>in vitro</em> metabolic engineering to produce isobutanol. The <em>in vivo</em> production of α-ketoisovalerate (KIV) was conducted through CRISPR mediated integration of the KIV pathway in bicistronic design (BCD) in <em>E. coli</em> and inhibition of competitive valine pathway using CRISPRi technology. The subsequent <em>in vitro</em> conversion to isobutanol was carried out with engineered enzymes for 2-ketoacid decarboxylase (KIVD) and alcohol dehydrogenase (ADH). For the <em>in vivo</em> production of KIV and subsequent <em>in vitro</em> production of isobutanol, this two-step serial approach resulted in yields of 56% and 93%, productivities of 0.62 and 0.074 g L⁻¹ h⁻¹, and titers of 5.6 and 1.78 g L⁻¹, respectively. Thus, this combined biosynthetic system can be used as a modular approach for producing important metabolites, like isobutanol, without the limitations associated with <em>in vivo</em> production using a consolidated bioprocess.</p></div>","PeriodicalId":18695,"journal":{"name":"Metabolic Engineering Communications","volume":"15 ","pages":"Article e00210"},"PeriodicalIF":5.2,"publicationDate":"2022-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S2214030122000190/pdfft?md5=b291a3d0e60096f33c80f32a13d96aa2&pid=1-s2.0-S2214030122000190-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"91640475","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":"Metabolomics and modelling approaches for systems metabolic engineering","authors":"Jasmeet Kaur Khanijou ,&nbsp;Hanna Kulyk ,&nbsp;Cécilia Bergès ,&nbsp;Leng Wei Khoo ,&nbsp;Pnelope Ng ,&nbsp;Hock Chuan Yeo ,&nbsp;Mohamed Helmy ,&nbsp;Floriant Bellvert ,&nbsp;Wee Chew ,&nbsp;Kumar Selvarajoo","doi":"10.1016/j.mec.2022.e00209","DOIUrl":"10.1016/j.mec.2022.e00209","url":null,"abstract":"<div><p>Metabolic engineering involves the manipulation of microbes to produce desirable compounds through genetic engineering or synthetic biology approaches. Metabolomics involves the quantitation of intracellular and extracellular metabolites, where mass spectrometry and nuclear magnetic resonance based analytical instrumentation are often used. Here, the experimental designs, sample preparations, metabolite quenching and extraction are essential to the quantitative metabolomics workflow. The resultant metabolomics data can then be used with computational modelling approaches, such as kinetic and constraint-based modelling, to better understand underlying mechanisms and bottlenecks in the synthesis of desired compounds, thereby accelerating research through systems metabolic engineering. Constraint-based models, such as genome scale models, have been used successfully to enhance the yield of desired compounds from engineered microbes, however, unlike kinetic or dynamic models, constraint-based models do not incorporate regulatory effects. Nevertheless, the lack of time-series metabolomic data generation has hindered the usefulness of dynamic models till today. In this review, we show that improvements in automation, dynamic real-time analysis and high throughput workflows can drive the generation of more quality data for dynamic models through time-series metabolomics data generation. Spatial metabolomics also has the potential to be used as a complementary approach to conventional metabolomics, as it provides information on the localization of metabolites. However, more effort must be undertaken to identify metabolites from spatial metabolomics data derived through imaging mass spectrometry, where machine learning approaches could prove useful. On the other hand, single-cell metabolomics has also seen rapid growth, where understanding cell-cell heterogeneity can provide more insights into efficient metabolic engineering of microbes. Moving forward, with potential improvements in automation, dynamic real-time analysis, high throughput workflows, and spatial metabolomics, more data can be produced and studied using machine learning algorithms, in conjunction with dynamic models, to generate qualitative and quantitative predictions to advance metabolic engineering efforts.</p></div>","PeriodicalId":18695,"journal":{"name":"Metabolic Engineering Communications","volume":"15 ","pages":"Article e00209"},"PeriodicalIF":5.2,"publicationDate":"2022-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/2e/c7/main.PMC9587336.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40679625","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 and engineering of the xylose-inducible xylP promoter for use in mold fungal species","authors":"Annie Yap ,&nbsp;Irene Glarcher ,&nbsp;Matthias Misslinger,&nbsp;Hubertus Haas","doi":"10.1016/j.mec.2022.e00214","DOIUrl":"https://doi.org/10.1016/j.mec.2022.e00214","url":null,"abstract":"<div><p>Conditional promoters allowing both induction and silencing of gene expression are indispensable for basic and applied research. The <em>xylP</em> promoter (p<em>xylP</em>) from <em>Penicillium chrysogenum</em> was demonstrated to function in various mold species including <em>Aspergillus fumigatus</em>. p<em>xylP</em> allows high induction by xylan or its degradation product xylose with low basal activity in the absence of an inducer. Here we structurally characterized and engineered p<em>xylP</em> in <em>A. fumigatus</em> to optimize its application. Mutational analysis demonstrated the importance of the putative TATA-box and a pyrimidine-rich region in the core promoter, both copies of a largely duplicated 91-bp sequence (91bpDS), as well as putative binding sites for the transcription factor XlnR and a GATA motif within the 91bpDS. In agreement, p<em>xylP</em> activity was found to depend on XlnR, while glucose repression appeared to be indirect. Truncation of the originally used 1643-bp promoter fragment to 725 bp largely preserved the promoter activity and the regulatory pattern. Integration of a third 91bpDS significantly increased promoter activity particularly under low inducer concentrations. Truncation of p<em>xylP</em> to 199 bp demonstrated that the upstream region including the 91bpDSs mediates not only inducer-dependent activation but also repression in the absence of inducer. Remarkably, the 1579-bp p<em>xylP</em> was found to act bi-bidirectionally with a similar regulatory pattern by driving expression of the upstream-located arabinofuranosidase gene. The latter opens the possibility of dual bidirectional use of p<em>xylP</em>. Comparison with a doxycycline-inducible TetOn system revealed a significantly higher dynamic range of p<em>xylP</em>. Taken together, this study identified functional elements of p<em>xylP</em> and opened new methodological opportunities for its application.</p></div>","PeriodicalId":18695,"journal":{"name":"Metabolic Engineering Communications","volume":"15 ","pages":"Article e00214"},"PeriodicalIF":5.2,"publicationDate":"2022-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S2214030122000232/pdfft?md5=43a616ce871d93dcd430b7cbcb1a7779&pid=1-s2.0-S2214030122000232-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"89988962","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":"Biosynthesis of value-added bioproducts from hemicellulose of biomass through microbial metabolic engineering","authors":"Biao Geng ,&nbsp;Xiaojing Jia ,&nbsp;Xiaowei Peng ,&nbsp;Yejun Han","doi":"10.1016/j.mec.2022.e00211","DOIUrl":"10.1016/j.mec.2022.e00211","url":null,"abstract":"<div><p>Hemicellulose is the second most abundant carbohydrate in lignocellulosic biomass and has extensive applications. In conventional biomass refinery, hemicellulose is easily converted to unwanted by-products in pretreatment and therefore can't be fully utilized. The present study aims to summarize the most recent development of lignocellulosic polysaccharide degradation and fully convert it to value-added bioproducts through microbial and enzymatic catalysis. Firstly, bioprocess and microbial metabolic engineering for enhanced utilization of lignocellulosic carbohydrates were discussed. The bioprocess for degradation and conversion of natural lignocellulose to monosaccharides and organic acids using anaerobic thermophilic bacteria and thermostable glycoside hydrolases were summarized. Xylose transmembrane transporting systems in natural microorganisms and the latest strategies for promoting the transporting capacity by metabolic engineering were summarized. The carbon catabolite repression effect restricting xylose utilization in microorganisms, and metabolic engineering strategies developed for co-utilization of glucose and xylose were discussed. Secondly, the metabolic pathways of xylose catabolism in microorganisms were comparatively analyzed. Microbial metabolic engineering for converting xylose to value-added bioproducts based on redox pathways, non-redox pathways, pentose phosphate pathway, and improving inhibitors resistance were summarized. Thirdly, strategies for degrading lignocellulosic polysaccharides and fully converting hemicellulose to value-added bioproducts through microbial metabolic engineering were proposed.</p></div>","PeriodicalId":18695,"journal":{"name":"Metabolic Engineering Communications","volume":"15 ","pages":"Article e00211"},"PeriodicalIF":5.2,"publicationDate":"2022-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9597109/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40443804","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":"Metabolic engineering of Pseudomonas taiwanensis VLB120 for rhamnolipid biosynthesis from biomass-derived aromatics","authors":"Vaishnavi Sivapuratharasan ,&nbsp;Christoph Lenzen ,&nbsp;Carina Michel ,&nbsp;Anantha Barathi Muthukrishnan ,&nbsp;Guhan Jayaraman ,&nbsp;Lars M. Blank","doi":"10.1016/j.mec.2022.e00202","DOIUrl":"https://doi.org/10.1016/j.mec.2022.e00202","url":null,"abstract":"<div><p>Lignin is a ubiquitously available and sustainable feedstock that is underused as its depolymerization yields a range of aromatic monomers that are challenging substrates for microbes. In this study, we investigated the growth of <em>Pseudomonas taiwanensis</em> VLB120 on biomass-derived aromatics, namely, 4-coumarate, ferulate, 4-hydroxybenzoate, and vanillate. The wild type strain was not able to grow on 4-coumarate and ferulate. After integration of catabolic genes for breakdown of 4-coumarate and ferulate, the metabolically engineered strain was able to grow on these aromatics. Further, the specific growth rate of the strain was enhanced up to 3-fold using adaptive laboratory evolution, resulting in increased tolerance towards 4-coumarate and ferulate. Whole-genome sequencing highlighted several different mutations mainly in two genes. The first gene was <em>actP</em>, coding for a cation/acetate symporter, and the other gene was <em>paaA</em> coding for a phenyl acetyl-CoA oxygenase. The evolved strain was further engineered for rhamnolipid production. Among the biomass-derived aromatics investigated, 4-coumarate and ferulate were promising substrates for product synthesis. With 4-coumarate as the sole carbon source, a yield of 0.27 (Cmol_rhl/Cmol_4-coumarate) was achieved, corresponding to 28% of the theoretical yield. Ferulate enabled a yield of about 0.22 (Cmol_rhl/Cmol_ferulate), representing 42% of the theoretical yield. Overall, this study demonstrates the use of biomass-derived aromatics as novel carbon sources for rhamnolipid biosynthesis.</p></div>","PeriodicalId":18695,"journal":{"name":"Metabolic Engineering Communications","volume":"15 ","pages":"Article e00202"},"PeriodicalIF":5.2,"publicationDate":"2022-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S2214030122000116/pdfft?md5=29575c540fc6b7e0e6bf633a82266d60&pid=1-s2.0-S2214030122000116-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"89989006","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}