Karishma Shah , Daniel Kracher , Peter Macheroux , Silvia Wallner , André Pick , Robert Kourist
{"title":"Discovery and characterization of NADH oxidases for selective sustainable synthesis of 5-hydroxymethylfuran carboxylic acid","authors":"Karishma Shah , Daniel Kracher , Peter Macheroux , Silvia Wallner , André Pick , Robert Kourist","doi":"10.1016/j.jbiotec.2024.11.009","DOIUrl":"10.1016/j.jbiotec.2024.11.009","url":null,"abstract":"<div><div>Efficient regeneration of NAD<sup>+</sup> remains a significant challenge for oxidative biotransformations. In order to identify enzymes with higher activity and stability, a panel of NADH oxidases (Nox) was investigated in the regeneration of nicotinamide cofactors for the oxidation of hydroxymethyl furfural (HMF) to 5-hydroxymethyl-2-furancarboxylic acid (HMFCA). We present novel Nox that exhibit remarkable catalytic activities, elevated thermal and pH stabilities, and higher intrinsic flavin loadings, thus eliminating the need for external flavin addition. The kinetic analysis of the NADH oxidases indicates that <em>Ad</em>Nox, <em>Gd</em>Nox, <em>Cm</em>Nox, and <em>Lv</em>Nox exhibit V<sub>max</sub> values of 86 U/mg, 50 U/mg, 4.3 U/mg, and 23 U/mg, respectively. When these NADH oxidases were applied in a HMF oxidation reaction, LvNox demonstrated the highest HMFCA yield of 97 % in the presence of 0.1 mM NAD and 10 mM HMF. In contrast to previously reported NADH oxidases from the same family, these NADH oxidases naturally accept NADPH as a substrate. Rapid kinetics experiments identified the oxidative reaction as the rate-limiting step of the reaction. NADH oxidases achieved high atom economy, a high reaction mass efficiency and a low process mass intensity. The findings contribute significantly to the field of biocatalysis and offer potential avenues for more environmentally friendly cofactor regeneration in chemical synthesis.</div></div>","PeriodicalId":15153,"journal":{"name":"Journal of biotechnology","volume":"398 ","pages":"Pages 18-28"},"PeriodicalIF":4.1,"publicationDate":"2025-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142739743","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Andreas Maccani , Robert Pachlinger , Peter Eisenhut, Sabine Unterthurner, Susanne Heider, Christoph Repik, Stefan Reuberger, Peter Andorfer, Johannes Lengler, Bernhard Kinastberger, Dieter Seczer, Petra Gruber, Alexander Apschner, Lucia Micutkova
{"title":"Potency by design: Novel insights in transfection and purification for manufacturing of rAAV gene therapy vectors","authors":"Andreas Maccani , Robert Pachlinger , Peter Eisenhut, Sabine Unterthurner, Susanne Heider, Christoph Repik, Stefan Reuberger, Peter Andorfer, Johannes Lengler, Bernhard Kinastberger, Dieter Seczer, Petra Gruber, Alexander Apschner, Lucia Micutkova","doi":"10.1016/j.jbiotec.2024.12.007","DOIUrl":"10.1016/j.jbiotec.2024.12.007","url":null,"abstract":"<div><div>This study investigates the crucial role of transfection methods in the manufacturability and potency of recombinant adeno-associated virus (rAAV) gene therapies. By employing a novel analytical approach, multiplex digital PCR (dPCR), we evaluated the impact of different transfection reagents and conditions on the scalability and quality of rAAV. Our research demonstrates that the selection of transfection approach significantly influences not only the yield and ease of scale-up but also the potency of the final product. Importantly, later changes to transfection parameters established in the early stages of development can be challenging, potentially compromising product quality and leading to comparability issues. Leveraging multiplex dPCR has proven instrumental in guiding these early-stage decisions, ensuring a reliable manufacturing process that consistently delivers high-quality therapeutic products. Our findings highlight the importance of optimizing transfection strategies early in development to guarantee the successful production of potent and cost-effective gene therapies, ultimately impacting patient accessibility.</div></div>","PeriodicalId":15153,"journal":{"name":"Journal of biotechnology","volume":"398 ","pages":"Pages 127-132"},"PeriodicalIF":4.1,"publicationDate":"2025-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142864376","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Supenya Chittapun , Kattiya Suwanmanee , Chatchol Kongsinkaew , Soisuda Pornpukdeewattana , Yusuf Chisti , Theppanya Charoenrat
{"title":"Thermal degradation kinetics and purification of C-phycocyanin from thermophilic and mesophilic cyanobacteria","authors":"Supenya Chittapun , Kattiya Suwanmanee , Chatchol Kongsinkaew , Soisuda Pornpukdeewattana , Yusuf Chisti , Theppanya Charoenrat","doi":"10.1016/j.jbiotec.2024.11.018","DOIUrl":"10.1016/j.jbiotec.2024.11.018","url":null,"abstract":"<div><div>The natural blue colorant C-phycocyanin (C-PC) has many potential applications but its poor heat stability limits its commercial use. This study compares the production and thermal stability of C-PC from two cyanobacteria: the thermophilic <em>Thermosynechococcus</em> sp. TUBT-T01 and the mesophilic <em>Synechococcus cedrorum</em> TISTR8589. <em>Thermosynechococcus</em> sp. produced nearly 1.9-fold more C-PC than <em>S. cedrorum</em>. Batch adsorption using a chromatographic cationic ion exchange resin (Streamline Direct HST1) was used to effectively purify the C-PC. The equilibrium adsorption capacity (<em>Q</em><sub><em>eq</em></sub>) of the resin for C-PC was the highest at pH 5. At this pH, the <em>Q</em><sub><em>eq</em></sub> for the thermophilic C-PC was 5.5 ± 0.1 mg mL⁻¹ , whereas for the mesophilic C-PC it was 1.5 ± 0.2 mg mL⁻¹ . Purification increased the concentration of the thermophilic C-PC by 5.9-fold, and that of mesophilic C-PC by 4.2-fold. The purity ratios of the final products from the two cyanobacteria were similar at ∼2.2. At 60 °C and pH 7, the C-PC of <em>Thermosynechococcus</em> sp. had ∼12-times longer half-life than the mesophilic C-PC; however, the productivity of the thermophilic C-PC was comparatively low because of a low biomass productivity of <em>Thermosynechococcus</em> sp.</div></div>","PeriodicalId":15153,"journal":{"name":"Journal of biotechnology","volume":"398 ","pages":"Pages 76-86"},"PeriodicalIF":4.1,"publicationDate":"2025-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142769264","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Corrigendum to ‘Enhanced activation of signaling pathway by recombinant human adiponectin from genome-edited chickens’ [J. Biotechnol., Vol. 395 (2024) 95–99/0168–1656]","authors":"Eunhui Yoo, Hee Jung Choi, Jae Yong Han","doi":"10.1016/j.jbiotec.2024.11.019","DOIUrl":"10.1016/j.jbiotec.2024.11.019","url":null,"abstract":"","PeriodicalId":15153,"journal":{"name":"Journal of biotechnology","volume":"398 ","pages":"Page 41"},"PeriodicalIF":4.1,"publicationDate":"2025-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142785728","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Ting Wei , Hong Li , Yuyao Wang , Ming Chi , Junkang Guo , Honglei Jia , Chao Zhang
{"title":"Alleviation of cadmium toxicity and minimizing its accumulation in rice plants by methyl jasmonate: Performance and mechanisms","authors":"Ting Wei , Hong Li , Yuyao Wang , Ming Chi , Junkang Guo , Honglei Jia , Chao Zhang","doi":"10.1016/j.jbiotec.2024.12.009","DOIUrl":"10.1016/j.jbiotec.2024.12.009","url":null,"abstract":"<div><div>Heavy metal pollution is a worldwide problem that threaten agricultural production and human health. Methyl jasmonate (MeJA) is a phytohormone that could enhance plant resistance against various stresses. However, the mechanism of MeJA in cadmium (Cd) uptake, distribution, and translocation in rice plants remains elusive. In this study, we found that the Cd induced-growth inhibition was ameliorated by MeJA. Upon MeJA application, Cd content in root and shoot was decreased by 10.15 % and 36.39 %, which paralleled with less Cd<sup>2 +</sup> influx of rice roots and depressed expression of the cation transporters (<em>OsNramp1</em> and <em>OsNramp5</em>). The subcellular distribution revealed that MeJA enriched Cd distribution in cell wall, which was accompanied by increased cell wall thickness and altered cell wall polysaccharide (pectin, cellulose, hemicellulose) content, meanwhile, the Cd content in pectin, cellulose, hemicellulose was increased, the FTIR analysis implied that functional groups (especially -OH and COO-) on cell wall were involved in Cd fixation. The root to shoot translocation of Cd was hindered by exogenous MeJA, this was validated by the decreased expression of <em>OsHMA2</em> in root and declined Cd level in xylem sap. Overall, our results revealed that MeJA could act as a foliar resistance control substance to reduce Cd accumulation in rice plants. The detailed molecular mechanisms of MeJA in Cd detoxification in plants still need further investigation.</div></div>","PeriodicalId":15153,"journal":{"name":"Journal of biotechnology","volume":"398 ","pages":"Pages 133-145"},"PeriodicalIF":4.1,"publicationDate":"2025-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142894664","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Karthika Balusamy , Balaji V. Rokade , Manuel Bruch , Meg Walsh , William Casey , Federico Cerrone , Bryan Dalton , Patrick Jerome Guiry , Tanja Narančić , Kevin E. O’Connor
{"title":"Converting multiple hydrophobic aromatic plastic monomers into a single water-soluble substrate to increase bioavailability for the synthesis of polyhydroxyalkanoates by bacteria using batch, fed batch and continuous cultivation","authors":"Karthika Balusamy , Balaji V. Rokade , Manuel Bruch , Meg Walsh , William Casey , Federico Cerrone , Bryan Dalton , Patrick Jerome Guiry , Tanja Narančić , Kevin E. O’Connor","doi":"10.1016/j.jbiotec.2024.12.008","DOIUrl":"10.1016/j.jbiotec.2024.12.008","url":null,"abstract":"<div><div>We demonstrate the proof of concept of increasing the bioavailability of carbon substrates, derived from plastic waste, for their conversion to the biodegradable polymer polyhydroxyalkanoate [PHA] by bacteria and test various approaches to PHA accumulation through batch, fed batch and continuous culture. Styrene, ethylbenzene, and toluene are produced from the pyrolysis of mixed plastic waste (Kaminsky, 2021; Miandad et al., 2017), but they are volatile and poorly soluble in water making them difficult to work with in aqueous fermentation systems. By chemically converting these aromatic compounds to benzoic acid, and subsequently to its sodium salt, we increased the solubility and reduced the volatility of the substrate supplied to <em>Pseudomonas putida</em> CA-3 to accumulate polyhydroxyalkanoates. 1 L scale batch, fed batch, and continuous fermentations were carried out; the fed batch fermentation resulted in the maximum volumetric PHA productivity of 61.67 ± 7.34 mg L<sup>−1</sup> h<sup>−1</sup>; while batch and continuous, at a dilution rate, d = 0.2 h<sup>−1</sup>, fermentations resulted in 13.30 ± 0.01 and 4.06 ± 0.01 mg L<sup>−1</sup> h<sup>−1</sup> of PHA respectively.</div></div>","PeriodicalId":15153,"journal":{"name":"Journal of biotechnology","volume":"398 ","pages":"Pages 193-201"},"PeriodicalIF":4.1,"publicationDate":"2025-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142894667","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Guoli Wang , Xiqin Liang , Zhenke Wu, Bengui Fan, Jun Wang, Qiusheng Zheng, Defang Li, Tianyue An
{"title":"Improved biosynthesis of tyrosol by epigenetic modification-based regulation and metabolic engineering in Saccharomyces cerevisiae","authors":"Guoli Wang , Xiqin Liang , Zhenke Wu, Bengui Fan, Jun Wang, Qiusheng Zheng, Defang Li, Tianyue An","doi":"10.1016/j.jbiotec.2024.12.013","DOIUrl":"10.1016/j.jbiotec.2024.12.013","url":null,"abstract":"<div><div>Aromatic amino acids and their derivatives are high value chemicals widely used in food, pharmaceutical and feed industries. Current preparation methods for aromatic amino acid products are fraught with limitations. In this study, the efficient biosynthesis of aromatic amino acid compound tyrosol was investigated by epigenetic modification-based regulation and optimization of the biosynthetic pathway of aromatic amino acids. The production of tyrosol was significantly improved by the overexpression of m<sup>6</sup>A modification writer Ime4 and reader Pho92, and the positive regulator Gcr2. Introduction of Bbxfpk and deletion of Gpp1 further improved tyrosol production. Then the feedback inhibition of the shikimate pathway was relieved by the mutants Aro4<sup>K229L</sup> and Aro7<sup>G141S</sup>. The final tyrosol producing engineered strain was constructed by the deletion of <em>PHA2</em>, replacement of the native promoter of <em>ARO10</em> with the strong promoter PGK1p, and introduction of tyrosine decarboxylase PcAAS. In the background of m<sup>6</sup>A modification regulation, this strain ultimately produced 954.69 ± 43.72 mg/L of tyrosol, promoted by 61.7-fold in shake-flask fermentation.</div></div>","PeriodicalId":15153,"journal":{"name":"Journal of biotechnology","volume":"398 ","pages":"Pages 175-182"},"PeriodicalIF":4.1,"publicationDate":"2025-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142921807","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Marcelo M. Ruas-Filho , Ana Maria Mazotto , Anderson S. Pinheiro
{"title":"High-yield soluble production of recombinant β-keratin from Gallus gallus feathers using an experimental design approach","authors":"Marcelo M. Ruas-Filho , Ana Maria Mazotto , Anderson S. Pinheiro","doi":"10.1016/j.jbiotec.2025.01.002","DOIUrl":"10.1016/j.jbiotec.2025.01.002","url":null,"abstract":"<div><div>The search for new non-animal textile materials has increased yearly as environmental awareness and veganism continue to spread, driving the development of greener fabrics. Concurrently, β-keratin, a fibrous, resistant, and insoluble protein shows great potential for producing innovative biomaterials. However, β-keratin is naturally abundant in animal feathers. Therefore, the recombinant production of β-keratin from G<em>allus gallus</em> feathers was proposed using a strategy of parallel expression in different vectors. Statistical tools of experimental design were employed to improve the production of soluble biosynthetic keratin. It was shown that β-keratins fused to His<sub>6</sub>MBP had better performance regarding soluble expression. In addition, the optimized regions for the values of induction temperature, induction time, and induction absorbance were obtained. As a result, a yield of 185.3 ± 1.4 mg/L of soluble His<sub>6</sub>MBP-Chr2.FK4 was achieved, representing the highest yield reported to date.</div></div>","PeriodicalId":15153,"journal":{"name":"Journal of biotechnology","volume":"398 ","pages":"Pages 215-223"},"PeriodicalIF":4.1,"publicationDate":"2025-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142970739","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Bone marrow-derived dendritic cells play a role in attenuating inflammation on Bothrops jararacussu venom muscle damage","authors":"N.M. Nery , A.A. Ferreira e Ferreira , H.M. Santana , S.N. Serrath , V.P. Reis , M.V. Paloschi , M.D.S. Silva , J.G.S. Magalhães , L.F. Cruz , T.Y. Shibayama , S.S. Setubal. , J.P. Zuliani","doi":"10.1016/j.jbiotec.2024.11.014","DOIUrl":"10.1016/j.jbiotec.2024.11.014","url":null,"abstract":"<div><div>The immune system is regulated by dendritic cells (DCs), which are highly specialized cells for presenting antigens. They are thought of as natural sentinels that start the immune response triggered by naive T cells against invasive infections. DCs participate in the initial stage of muscle damage in conjunction with monocytes, macrophages, and myogenic cells. The goal of this study was to determine whether DCs might mitigate tissue damage and aid in the regeneration of the gastrocnemius muscle following envenomation with <em>Bothrops jararacussu</em> venom (BjV). Mature bone marrow dendritic cells (BMDCs) were used to treat mice in an experimental envenomation model with BjV by activation with lipopolysaccharide (LPS). BMDCs were injected into the gastrocnemius muscle at the same site of the BjV injury, in a single dose, 3 h after envenomation, and envenoming effects were observed at different periods for 7 days. In both untreated (NT) and treated (T) groups tissue necrosis, leukocyte influx, and hemorrhage at the injury site were observed. Results showed an increase in serum and tissue CK as well as IL-6, TNF-α, and IL-1β release in the first hours after envenoming. In contrast, after treatment with BMDCs results obtained demonstrated an attenuated local effect with a small leukocyte influx, decreased or non-existent necrosis and hemorrhage, as well as a reduction in both serum and tissue CK levels as well as cytokine release and, consequently, the onset of a moderate regenerative process. The present study's findings concluded that BjV causes a severe inflammatory reaction at the site of injury and that treating envenoming with BMDCs in the muscle was crucial for minimizing damage to the muscle and the inflammatory reaction and promoting the early onset of the tissue repair process.</div></div>","PeriodicalId":15153,"journal":{"name":"Journal of biotechnology","volume":"398 ","pages":"Pages 29-40"},"PeriodicalIF":4.1,"publicationDate":"2025-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142769260","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Qiaolei Zhu , Bingyu Zhao , Nairu Ji , Yunping Zhu , Xinyu Shi
{"title":"Effects of ORF14 gene on melanin expression, fermentation conditions and properties of melanin production in modified strains","authors":"Qiaolei Zhu , Bingyu Zhao , Nairu Ji , Yunping Zhu , Xinyu Shi","doi":"10.1016/j.jbiotec.2025.01.001","DOIUrl":"10.1016/j.jbiotec.2025.01.001","url":null,"abstract":"<div><div>Melanin with antioxidant and antibacterial properties can be used in food, cosmetics, biotechnology, and other fields, but its insolubility become a main challenge hindering for its application. In this study, water-soluble melanin produced by the novel species <em>Streptomyces vilmorinianum</em> YP1 was characterized using scanning electron microscopy (SEM), UV<img>visible spectroscopy (with an absorption peak at 220 nm), and Fourier transform infrared (FTIR) spectroscopy. The glycosyltransferase gene <em>ORF14</em> was knocked out, which improved the production of water-soluble melanin by inhibiting competitive pathway. In order to further enhance production of melanin, Plackett<img>Burman and response surface methodology statistical design was employed to screen for key factors and determine the optimal combination. The maximum melanin production (4.00 g/L) was obtained under the conditions: amylodextrine concentration of 40 g/L, soya peptone concentration of 7 g/L, tryptone concentration of 5 g/L, NaCl concentration of 5.4 g/L, pH of 6.7 and temperature of 36 °C for 180 h. The physicochemical properties and bioactivity of melanin were further investigated, revealing that melanin had a good stability across a pH range of 4–12, antioxidant (with a survival rate of over 85 %), and resistance to reducing agents (with a survival rate of over 99 %). The results underscored that <em>S. vilmorinianum</em> YP1 is a promising candidate for water-soluble melanin production.</div></div>","PeriodicalId":15153,"journal":{"name":"Journal of biotechnology","volume":"398 ","pages":"Pages 224-234"},"PeriodicalIF":4.1,"publicationDate":"2025-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142970131","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}