Biotechnology for Biofuels最新文献

{"title":"Insights into the action of phylogenetically diverse microbial expansins on the structure of cellulose microfibrils","authors":"Majid Haddad Momeni,&nbsp;Aleksi Zitting,&nbsp;Vilma Jäämuru,&nbsp;Rosaliina Turunen,&nbsp;Paavo Penttilä,&nbsp;Garry W. Buchko,&nbsp;Salla Hiltunen,&nbsp;Natalia Maiorova,&nbsp;Anu Koivula,&nbsp;Janak Sapkota,&nbsp;Kaisa Marjamaa,&nbsp;Emma R. Master","doi":"10.1186/s13068-024-02500-w","DOIUrl":"10.1186/s13068-024-02500-w","url":null,"abstract":"<div><h3>Background</h3><p>Microbial expansins (EXLXs) are non-lytic proteins homologous to plant expansins involved in plant cell wall formation. Due to their non-lytic cell wall loosening properties and potential to disaggregate cellulosic structures, there is considerable interest in exploring the ability of microbial expansins (EXLX) to assist the processing of cellulosic biomass for broader biotechnological applications. Herein, EXLXs with different modular structure and from diverse phylogenetic origin were compared in terms of ability to bind cellulosic, xylosic, and chitinous substrates, to structurally modify cellulosic fibrils, and to boost enzymatic deconstruction of hardwood pulp.</p><h3>Results</h3><p>Five heterogeneously produced EXLXs (<i>Clavibacter michiganensis; Cmi</i>EXLX2, <i>Dickeya aquatica; Daq</i>EXLX1, <i>Xanthomonas sacchari; Xsa</i>EXLX1, <i>Nothophytophthora sp.; Nsp</i>EXLX1 and <i>Phytophthora cactorum; Pca</i>EXLX1) were shown to bind xylan and hardwood pulp at pH 5.5 and <i>Cmi</i>EXLX2 (harboring a family-2 carbohydrate-binding module) also bound well to crystalline cellulose. Small-angle X-ray scattering revealed a 20–25% increase in interfibrillar distance between neighboring cellulose microfibrils following treatment with <i>Cmi</i>EXLX2, <i>Daq</i>EXLX1, or <i>Nsp</i>EXLX1. Correspondingly, combining xylanase with <i>Cmi</i>EXLX2 and <i>Daq</i>EXLX1 increased product yield from hardwood pulp by ~ 25%, while supplementing the <i>Tr</i>AA9A LPMO from <i>Trichoderma reesei</i> with <i>Cmi</i>EXLX2, <i>Daq</i>EXLX1, and <i>Nsp</i>EXLX1 increased total product yield by over 35%.</p><h3>Conclusion</h3><p>This direct comparison of diverse EXLXs revealed consistent impacts on interfibrillar spacing of cellulose microfibers and performance of carbohydrate-active enzymes predicted to act on fiber surfaces. These findings uncover new possibilities to employ EXLXs in the creation of value-added materials from cellulosic biomass.</p></div>","PeriodicalId":494,"journal":{"name":"Biotechnology for Biofuels","volume":"17 1","pages":""},"PeriodicalIF":6.3,"publicationDate":"2024-04-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://biotechnologyforbiofuels.biomedcentral.com/counter/pdf/10.1186/s13068-024-02500-w","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140639474","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Small GTPase Rab7 is involved in stress adaptation to carbon starvation to ensure the induced cellulase biosynthesis in Trichoderma reesei","authors":"Lin Liu,&nbsp;Zhixing Wang,&nbsp;Yu Fang,&nbsp;Renfei Yang,&nbsp;Yi Pu,&nbsp;Xiangfeng Meng,&nbsp;Weifeng Liu","doi":"10.1186/s13068-024-02504-6","DOIUrl":"10.1186/s13068-024-02504-6","url":null,"abstract":"<div><h3>Background</h3><p>The saprophytic filamentous fungus <i>Trichoderma reesei</i> represents one of the most prolific cellulase producers. The bulk production of lignocellulolytic enzymes by <i>T. reesei</i> not only relies on the efficient transcription of cellulase genes but also their efficient secretion after being translated. However, little attention has been paid to the functional roles of the involved secretory pathway in the high-level production of cellulases in <i>T. reesei</i>. Rab GTPases are key regulators in coordinating various vesicle trafficking associated with the eukaryotic secretory pathway. Specifically, Rab7 is a representative GTPase regulating the transition of the early endosome to the late endosome followed by its fusion to the vacuole as well as homotypic vacuole fusion. Although crosstalk between the endosomal/vacuolar pathway and the secretion pathway has been reported, the functional role of Rab7 in cellulase production in <i>T. reesei</i> remains unknown.</p><h3>Results</h3><p>A TrRab7 was identified and characterized in <i>T. reesei</i>. TrRab7 was shown to play important roles in <i>T. reesei</i> vegetative growth and vacuole morphology. Whereas knock-down of <i>Trrab7</i> significantly compromised the induced production of <i>T. reesei</i> cellulases, overexpression of the key transcriptional activator, Xyr1, restored the production of cellulases in the <i>Trrab7</i> knock-down strain (P<i>tcu</i>-<i>rab7</i>^KD) on glucose, indicating that the observed defective cellulase biosynthesis results from the compromised cellulase gene transcription. Down-regulation of <i>Trrab7</i> was also found to make <i>T. reesei</i> more sensitive to various stresses including carbon starvation. Interestingly, overexpression of Snf1, a serine/threonine protein kinase known as an energetic sensor, partially restored the cellulase production of P<i>tcu</i>-<i>rab7</i>^KD on Avicel, implicating that TrRab7 is involved in an energetic adaptation to carbon starvation which contributes to the successful cellulase gene expression when <i>T. reesei</i> is transferred from glucose to cellulose.</p><h3>Conclusions</h3><p>TrRab7 was shown to play important roles in <i>T. reesei</i> development and a stress response to carbon starvation resulting from nutrient shift. This adaptation may allow <i>T. reesei</i> to successfully initiate the inducing process leading to efficient cellulase production. The present study provides useful insights into the functional involvement of the endosomal/vacuolar pathway in <i>T. reesei</i> development and hydrolytic enzyme production.</p></div>","PeriodicalId":494,"journal":{"name":"Biotechnology for Biofuels","volume":"17 1","pages":""},"PeriodicalIF":6.3,"publicationDate":"2024-04-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://biotechnologyforbiofuels.biomedcentral.com/counter/pdf/10.1186/s13068-024-02504-6","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140625266","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Cellular ATP redistribution achieved by deleting Tgparp improves lignocellulose utilization of Trichoderma under heat stress","authors":"Tuo Li,&nbsp;Yang Liu,&nbsp;Han Zhu,&nbsp;Linhua Cao,&nbsp;Yihao Zhou,&nbsp;Dongyang Liu,&nbsp;Qirong Shen","doi":"10.1186/s13068-024-02502-8","DOIUrl":"10.1186/s13068-024-02502-8","url":null,"abstract":"<div><h3>Background</h3><p>Thermotolerance is widely acknowledged as a pivotal factor for fungal survival across diverse habitats. Heat stress induces a cascade of disruptions in various life processes, especially in the acquisition of carbon sources, while the mechanisms by which filamentous fungi adapt to heat stress and maintain carbon sources are still not fully understood.</p><h3>Results</h3><p>Using <i>Trichoderma guizhouense</i>, a representative beneficial microorganism for plants, we discover that heat stress severely inhibits the lignocellulases secretion, affecting carbon source utilization efficiency. Proteomic results at different temperatures suggest that proteins involved in the poly ADP-ribosylation pathway (<i>Tg</i>PARP and <i>Tg</i>ADPRase) may play pivotal roles in thermal adaptation and lignocellulose utilization. <i>Tg</i>PARP is induced by heat stress, while the deletion of <i>Tgparp</i> significantly improves the lignocellulose utilization capacity and lignocellulases secretion in <i>T. guizhouense</i>. Simultaneously, the absence of <i>Tgparp</i> prevents the excessive depletion of ATP and NAD⁺, enhances the protective role of mitochondrial membrane potential (MMP), and elevates the expression levels of the unfolded protein response (UPR)-related regulatory factor <i>Tgire</i>. Further investigations reveal that a stable MMP can establish energy homeostasis, allocating more ATP within the endoplasmic reticulum (ER) to reduce protein accumulation in the ER, thereby enhancing the lignocellulases secretion in <i>T. guizhouense</i> under heat stress.</p><h3>Conclusions</h3><p>Overall, these findings underscored the significance of <i>Tgparp</i> as pivotal regulators in lignocellulose utilization under heat stress and provided further insights into the molecular mechanism of filamentous fungi in utilizing lignocellulose.</p></div>","PeriodicalId":494,"journal":{"name":"Biotechnology for Biofuels","volume":"17 1","pages":""},"PeriodicalIF":6.3,"publicationDate":"2024-04-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://biotechnologyforbiofuels.biomedcentral.com/counter/pdf/10.1186/s13068-024-02502-8","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140619688","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Structural and functional analysis of the active cow rumen’s microbial community provides a catalogue of genes and microbes participating in the deconstruction of cardoon biomass","authors":"Andrea Firrincieli,&nbsp;Andrea Minuti,&nbsp;Martina Cappelletti,&nbsp;Marco Ferilli,&nbsp;Paolo Ajmone-Marsan,&nbsp;Paolo Bani,&nbsp;Maurizio Petruccioli,&nbsp;Antoine L. Harfouche","doi":"10.1186/s13068-024-02495-4","DOIUrl":"10.1186/s13068-024-02495-4","url":null,"abstract":"&lt;div&gt;&lt;h3&gt;Background&lt;/h3&gt;&lt;p&gt;Ruminal microbial communities enriched on lignocellulosic biomass have shown considerable promise for the discovery of microorganisms and enzymes involved in digesting cell wall compounds, a key bottleneck in the development of second-generation biofuels and bioproducts, enabling a circular bioeconomy. Cardoon (&lt;i&gt;Cynara cardunculus&lt;/i&gt;) is a promising inedible energy crop for current and future cellulosic biorefineries and the emerging bioenergy and bioproducts industries. The rumen microbiome can be considered an anaerobic “bioreactor”, where the resident microbiota carry out the depolymerization and hydrolysis of plant cell wall polysaccharides (PCWPs) through the catalytic action of fibrolytic enzymes. In this context, the rumen microbiota represents a potential source of microbes and fibrolytic enzymes suitable for biofuel production from feedstocks. In this study, metatranscriptomic and 16S rRNA sequencing were used to profile the microbiome and to investigate the genetic features within the microbial community adherent to the fiber fractions of the rumen content and to the residue of cardoon biomass incubated in the rumen of cannulated cows.&lt;/p&gt;&lt;h3&gt;Results&lt;/h3&gt;&lt;p&gt;The metatranscriptome of the cardoon and rumen fibre-adherent microbial communities were dissected in their functional and taxonomic components. From a functional point of view, transcripts involved in the methanogenesis from CO&lt;sub&gt;2&lt;/sub&gt; and H&lt;sub&gt;2&lt;/sub&gt;, and from methanol were over-represented in the cardoon-adherent microbial community and were affiliated with the &lt;i&gt;Methanobrevibacter&lt;/i&gt; and &lt;i&gt;Methanosphaera&lt;/i&gt; of the &lt;i&gt;Euryarchaeota&lt;/i&gt; phylum. Transcripts encoding glycoside hydrolases (GHs), carbohydrate-binding modules (CBMs), carbohydrate esterases (CEs), polysaccharide lyases (PLs), and glycoside transferases (GTs) accounted for 1.5% (6,957) of the total RNA coding transcripts and were taxonomically affiliated to major rumen fibrolytic microbes, such as &lt;i&gt;Oscillospiraceae&lt;/i&gt;, &lt;i&gt;Fibrobacteraceae&lt;/i&gt;, &lt;i&gt;Neocallimastigaceae&lt;/i&gt;, &lt;i&gt;Prevotellaceae, Lachnospiraceae,&lt;/i&gt; and &lt;i&gt;Treponemataceae&lt;/i&gt;. The comparison of the expression profile between cardoon and rumen fiber-adherent microbial communities highlighted that specific fibrolytic enzymes were potentially responsible for the breakdown of cardoon PCWPs, which was driven by specific taxa, mainly &lt;i&gt;Ruminococcus, Treponema,&lt;/i&gt; and &lt;i&gt;Neocallimastigaceae&lt;/i&gt;.&lt;/p&gt;&lt;h3&gt;Conclusions&lt;/h3&gt;&lt;p&gt;Analysis of 16S rRNA and metatranscriptomic sequencing data revealed that the cow rumen microbiome harbors a repertoire of new enzymes capable of degrading PCWPs. Our results demonstrate the feasibility of using metatranscriptomics of enriched microbial RNA as a potential approach for accelerating the discovery of novel cellulolytic enzymes that could be harnessed for biotechnology. This research contributes a relevant perspective towards degrading cellulosic biomass and providing an economical route to the","PeriodicalId":494,"journal":{"name":"Biotechnology for Biofuels","volume":"17 1","pages":""},"PeriodicalIF":6.3,"publicationDate":"2024-04-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://biotechnologyforbiofuels.biomedcentral.com/counter/pdf/10.1186/s13068-024-02495-4","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140537487","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Feedstock/pretreatment screening for bioconversion of sugar and lignin streams via deacetylated disc-refining","authors":"Darren J. Peterson,&nbsp;Changyub Paek,&nbsp;Ling Tao,&nbsp;Ryan Davis,&nbsp;Xiaowen Chen,&nbsp;Roman Brunecky,&nbsp;Matthew Fowler,&nbsp;Richard Elander","doi":"10.1186/s13068-024-02492-7","DOIUrl":"10.1186/s13068-024-02492-7","url":null,"abstract":"<div><p>Recent publications have shown the benefits of deacetylation disc-refining (DDR) as a pretreatment process to deconstruct biomass into sugars and lignin residues. Major advantages of DDR pretreatment over steam and dilute acid pretreatment are the removal of acetyl and lignin during deacetylation. DDR does not generate hydroxymethylfurfural (HMF) and furfural which are commonly produced from steam and dilute acid pretreatments. Acetate, lignin, HMF, and furfural are known inhibitors during enzymatic hydrolysis and fermentation. Another advantage of deacetylation is the production of lignin-rich black liquor, which can be upgraded to other bioproducts. Furthermore, due to the lack of sugar degradation during deacetylation, DDR has significantly less sugar loss than other pretreatment methods. Previous studies for DDR have primarily focused on corn stover, but lacked the investigative studies of other feedstocks. This study was designed to screen various DDR process conditions at pilot scale using three different feedstocks, including corn stover, poplar, and switchgrass. The impact of the pretreatment conditions was evaluated by testing hydrolysates for bioconversion to 2,3-butanediol. Pretreatment of biomass by DDR showed high-conversion-yields and 2,3-BDO fermentation production yields. Techno-economic analysis (TEA) of the pretreatment for biomass to sugar was also developed based on NREL’s Aspen Model. This study shows that the cellulose and hemicellulose in poplar was more recalcitrant than herbaceous feedstocks which ultimately drove up the sugar cost. Switchgrass was also more recalcitrant than corn stover but less than poplar.</p></div>","PeriodicalId":494,"journal":{"name":"Biotechnology for Biofuels","volume":"17 1","pages":""},"PeriodicalIF":6.3,"publicationDate":"2024-04-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://biotechnologyforbiofuels.biomedcentral.com/counter/pdf/10.1186/s13068-024-02492-7","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140351563","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Biosynthesis of fragrance 2-phenylethanol from sugars by Pseudomonas putida","authors":"Patricia Godoy,&nbsp;Zulema Udaondo,&nbsp;Estrella Duque,&nbsp;Juan L. Ramos","doi":"10.1186/s13068-024-02498-1","DOIUrl":"10.1186/s13068-024-02498-1","url":null,"abstract":"<div><h3>Background</h3><p>Petrochemicals contribute to environmental issues, with concerns ranging from energy consumption and carbon emission to pollution. In contrast, microbial biorefineries offer eco-friendly alternatives. The solvent-tolerant <i>Pseudomonas</i> <i>putida</i> DOT-T1E serves as a suitable host for producing aromatic compounds, specifically <span>l</span>-phenylalanine and its derivative, 2-phenylethanol (2-PE), which find widespread applications in various industries.</p><h3>Results</h3><p>This study focuses on enhancing 2-PE production in two <span>l</span>-phenylalanine overproducing strains of DOT-T1E, namely CM12-5 and CM12-5Δ<i>gcd</i> (<i>xylABE</i>), which grow with glucose and glucose-xylose, respectively. To synthesize 2-PE from <span>l</span>-phenylalanine, these strains were transformed with plasmid pPE-1, bearing the Ehrlich pathway genes, and it was found higher 2-PE production with glucose (about 50–60 ppm) than with xylose (&lt; 3 ppm). To understand the limiting factors, we tested the addition of phenylalanine and intermediates from the Ehrlich and shikimate pathways. The results identified intracellular <span>l</span>-phenylalanine as a key limiting factor for 2-PE production. To overcame this limitation, a chorismate mutase/prephenate dehydratase variant—insentive to feedback inhibition by aromatic amino acids—was introduced in the producing strains. This led to increased <span>l</span>-phenylalanine production and subsequently produced more 2-PE (100 ppm). Random mutagenesis of the strains also produced strains with higher <span>l</span>-phenylalanine titers and increased 2-PE production (up to 120 ppm). The improvements resulted from preventing dead-end product accumulation from shikimate and limiting the catabolism of potential pathway intermediates in the Ehrlich pathway. The study explored agricultural waste substrates, such as corn stover, sugarcane straw and corn-syrup as potential C sources. The best results were obtained using 2G substrates at 3% (between 82 and 100 ppm 2-PE), with glucose being the preferred sugar for 2-PE production among the monomeric sugars in these substrates.</p><h3>Conclusions</h3><p>The findings of this study offer strategies to enhance phenylalanine production, a key substrate for the synthesis of aromatic compounds. The ability of <i>P.</i> <i>putida</i> DOT-T1E to thrive with various C-sources and its tolerance to substrates, products, and potential toxicants in industrial wastes, are highlighted. The study identified and overcome possible bottlenecks for 2-PE production. Ultimately, the strains have potential to become efficient microbial platforms for synthesizing 2-PE from agro-industrial waste materials.</p></div>","PeriodicalId":494,"journal":{"name":"Biotechnology for Biofuels","volume":"17 1","pages":""},"PeriodicalIF":6.3,"publicationDate":"2024-04-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://biotechnologyforbiofuels.biomedcentral.com/counter/pdf/10.1186/s13068-024-02498-1","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140340215","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Regulatory mechanisms of autophagy on DHA and carotenoid accumulation in Crypthecodinium sp. SUN","authors":"Yiming Li,&nbsp;Tiantian Zhao,&nbsp;Weizheng Gao,&nbsp;Bowen Miao,&nbsp;Zhongxiang Fu,&nbsp;Zhao Zhang,&nbsp;Qingyang Li,&nbsp;Dongzhe Sun","doi":"10.1186/s13068-024-02493-6","DOIUrl":"10.1186/s13068-024-02493-6","url":null,"abstract":"<div><h3>Background</h3><p>Autophagy is a crucial process of cellular self-destruction and component reutilization that can affect the accumulation of total fatty acids (TFAs) and carotenoids in microalgae. The regulatory effects of autophagy process in a docosahexaenoic acid (DHA) and carotenoids simultaneously producing microalga, <i>Crypthecodinium</i> sp. SUN, has not been studied. Thus, the autophagy inhibitor (3-methyladenine (MA)) and activator (rapamycin) were used to regulate autophagy in <i>Crypthecodinium</i> sp. SUN.</p><h3>Results</h3><p>The inhibition of autophagy by 3-MA was verified by transmission electron microscopy, with fewer autophagy vacuoles observed. Besides, 3-MA reduced the glucose absorption and intracellular acetyl-CoA level, which resulting in the decrease of TFA and DHA levels by 15.83 and 26.73% respectively; Surprisingly, 3-MA increased intracellular reactive oxygen species level but decreased the carotenoids level. Comparative transcriptome analysis showed that the downregulation of the glycolysis, pentose phosphate pathway and tricarboxylic acid cycle may underlie the decrease of acetyl-CoA, NADPH and ATP supply for fatty acid biosynthesis; the downregulation of <i>PSY</i> and <i>HMGCR</i> may underlie the decreased carotenoids level. In addition, the class I PI3K-AKT signaling pathway may be crucial for the regulation of carbon and energy metabolism. At last, rapamycin was used to activate autophagy, which significantly enhanced the cell growth and TFA level and eventually resulted in 1.70-fold increase in DHA content.</p><h3>Conclusions</h3><p>Our findings indicate the mechanisms of autophagy in <i>Crypthecodinium</i> sp. SUN and highlight a way to manipulate cell metabolism by regulating autophagy. Overall, this study provides valuable insights to guide further research on autophagy-regulated TFA and carotenoids accumulation in <i>Crypthecodinium</i> sp. SUN.</p></div>","PeriodicalId":494,"journal":{"name":"Biotechnology for Biofuels","volume":"17 1","pages":""},"PeriodicalIF":6.3,"publicationDate":"2024-04-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://biotechnologyforbiofuels.biomedcentral.com/counter/pdf/10.1186/s13068-024-02493-6","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140340211","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A key gene, violaxanthin de-epoxidase-like 1, enhances fucoxanthin accumulation in Phaeodactylum tricornutum","authors":"Chenjie Li,&nbsp;Yufang Pan,&nbsp;Wenxiu Yin,&nbsp;Jin Liu,&nbsp;Hanhua Hu","doi":"10.1186/s13068-024-02496-3","DOIUrl":"10.1186/s13068-024-02496-3","url":null,"abstract":"<div><h3>Background</h3><p>Fucoxanthin has been widely investigated owing to its beneficial biological properties, and the model diatom <i>Phaeodactylum tricornutum</i>, possessing fucoxanthin (Fux) chlorophyll proteins as light-harvesting systems, is considered to have the potential to become a commercial cell factory for the pigment production.</p><h3>Results</h3><p>Here, we compared the pigment contents in 10 different <i>P. tricornutum</i> strains from the globe, and found that strain CCMP631 (Pt6) exhibited the highest Fux content but with a low biomass. Comparison of mRNA levels revealed that higher Fux content in Pt6 was related with the higher expression of gene <i>violaxanthin de-epoxidase-like (VDL) protein 1</i> (<i>VDL1</i>), which encodes the enzyme catalyzing the tautomerization of violaxanthin to neoxanthin in Fux biosynthesis pathway. Single nucleotide variants of <i>VDL1</i> gene and allele-specific expression in strains Pt1 (the whole genome sequenced strain CCMP632) and Pt6 were analyzed, and overexpressing of each of the 4 <i>VDL1</i> alleles, two from Pt1 and two from Pt6, in strain Pt1 leads to an increase in downstream product diadinoxanthin and channels the pigments towards Fux biosynthesis. All the 8 <i>VDL1</i> overexpression (OE) lines showed significant increases by 8.2 to 41.7% in Fux content without compromising growth, and <i>VDL1 Allele 2</i> OE lines even exhibited the higher cell density on day 8, with an increase by 24.2–28.7% in two <i>Pt1VDL1-allele 2</i> OE lines and 7.1–11.1% in two <i>Pt6VDL1-allele 2</i> OE lines, respectively.</p><h3>Conclusions</h3><p>The results reveal VDL1, localized in the plastid stroma, plays a key role in Fux over-accumulation in <i>P. tricornutum</i>. Overexpressing <i>VDL1</i>, especially <i>allele 2</i>, improved both the Fux content and growth rate, which provides a new strategy for the manipulation of Fux production in the future.</p></div>","PeriodicalId":494,"journal":{"name":"Biotechnology for Biofuels","volume":"17 1","pages":""},"PeriodicalIF":6.3,"publicationDate":"2024-04-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://biotechnologyforbiofuels.biomedcentral.com/counter/pdf/10.1186/s13068-024-02496-3","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140340197","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Development of a hybrid bio-purification process of lactic acid solutions employing an engineered E. coli strain in a membrane bioreactor","authors":"Alexandra Nastouli,&nbsp;Joseph Sweeney,&nbsp;Michael Harasek,&nbsp;Anastasios J. Karabelas,&nbsp;Sotiris I. Patsios","doi":"10.1186/s13068-024-02497-2","DOIUrl":"10.1186/s13068-024-02497-2","url":null,"abstract":"<div><h3>Background</h3><p>A potential alternative to lactic acid production through sugar fermentation is its recovery from grass silage leachate. The separation and purification of lactic acid from fermentation broths remain a key issue, as it amounts to up to 80% of its industrial production cost. In this study, a genetically engineered <i>E. coli</i> strain (A1:ldhA), that cannot catabolize lactic acid, has been used to selectively remove impurities from a synthetic medium comprising typical components (i.e., glucose and acetic acid) of green grass silage leachate. A systematic approach has been followed to provide a <i>proof-of-concept</i> for a bio-purification process of lactic acid solutions in a membrane bioreactor operating in semi-continuous mode.</p><h3>Results</h3><p>The synthetic medium composition was initially optimized in shake-flasks experiments, followed by scale-up in bench-scale bioreactor. Complete (i.e., 100%) and 60.4% removal for glucose and acetic acid, respectively, has been achieved in batch bioreactor experiments with a synthetic medium comprising 0.5 g/L glucose and 0.5 g/L acetic acid as carbon sources, and 10 g/L lactic acid; no lactic acid catabolism was observed in all batch fermentation tests. Afterwards, a hybrid biotechnological process combining semi-continuous bioreactor fermentation and ultrafiltration membrane separation (membrane bioreactor) was applied to in-situ separate purified medium from the active cells. The process was assessed under different semi-continuous operating conditions, resulting in a bacteria-free effluent and 100% glucose and acetic acid depletion, with no lactic acid catabolism, thus increasing the purity of the synthetic lactic acid solution.</p><h3>Conclusions</h3><p>The study clearly demonstrated that a bio-purification process for lactic acid employing the engineered <i>E. coli</i> strain cultivated in a membrane bioreactor is a technically feasible concept, paving the way for further technological advancement.</p></div>","PeriodicalId":494,"journal":{"name":"Biotechnology for Biofuels","volume":"17 1","pages":""},"PeriodicalIF":6.3,"publicationDate":"2024-03-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://biotechnologyforbiofuels.biomedcentral.com/counter/pdf/10.1186/s13068-024-02497-2","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140328973","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Substrate specificity mapping of fungal CAZy AA3_2 oxidoreductases","authors":"Hongbo Zhao,&nbsp;Johanna Karppi,&nbsp;Owen Mototsune,&nbsp;Daria Poshina,&nbsp;Jenny Svartström,&nbsp;Thi Truc Minh Nguyen,&nbsp;Tri Minh Vo,&nbsp;Adrian Tsang,&nbsp;Emma Master,&nbsp;Maija Tenkanen","doi":"10.1186/s13068-024-02491-8","DOIUrl":"10.1186/s13068-024-02491-8","url":null,"abstract":"<div><h3>Background</h3><p>Oxidative enzymes targeting lignocellulosic substrates are presently classified into various auxiliary activity (AA) families within the carbohydrate-active enzyme (CAZy) database. Among these, the fungal AA3 glucose–methanol–choline (GMC) oxidoreductases with varying auxiliary activities are attractive sustainable biocatalysts and important for biological function. CAZy AA3 enzymes are further subdivided into four subfamilies, with the large AA3_2 subfamily displaying diverse substrate specificities. However, limited numbers of enzymes in the AA3_2 subfamily are currently biochemically characterized, which limits the homology-based mining of new AA3_2 oxidoreductases. Importantly, novel enzyme activities may be discovered from the uncharacterized parts of this large subfamily.</p><h3>Results</h3><p>In this study, phylogenetic analyses employing a sequence similarity network (SSN) and maximum likelihood trees were used to cluster AA3_2 sequences. A total of 27 AA3_2 proteins representing different clusters were selected for recombinant production. Among them, seven new AA3_2 oxidoreductases were successfully produced, purified, and characterized. These enzymes included two glucose dehydrogenases (<i>Ta</i>GdhA and <i>Mc</i>GdhA), one glucose oxidase (<i>Ap</i>GoxA), one aryl alcohol oxidase (<i>Ps</i>AaoA), two aryl alcohol dehydrogenases (<i>As</i>AadhA and <i>As</i>AadhB), and one novel oligosaccharide (gentiobiose) dehydrogenase (<i>Ki</i>OdhA). Notably, two dehydrogenases (<i>Ta</i>GdhA and <i>Ki</i>OdhA) were found with the ability to utilize phenoxy radicals as an electron acceptor. Interestingly, phenoxy radicals were found to compete with molecular oxygen in aerobic environments when serving as an electron acceptor for two oxidases (<i>Ap</i>GoxA and <i>Ps</i>AaoA), which sheds light on their versatility. Furthermore, the molecular determinants governing their diverse enzymatic functions were discussed based on the homology model generated by AlphaFold.</p><h3>Conclusions</h3><p>The phylogenetic analyses and biochemical characterization of AA3_2s provide valuable guidance for future investigation of AA3_2 sequences and proteins. A clear correlation between enzymatic function and SSN clustering was observed. The discovery and biochemical characterization of these new AA3_2 oxidoreductases brings exciting prospects for biotechnological applications and broadens our understanding of their biological functions.</p></div>","PeriodicalId":494,"journal":{"name":"Biotechnology for Biofuels","volume":"17 1","pages":""},"PeriodicalIF":6.3,"publicationDate":"2024-03-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://biotechnologyforbiofuels.biomedcentral.com/counter/pdf/10.1186/s13068-024-02491-8","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140308301","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}