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Journal of Biotechnology

Caroline Medeiros Dos Reis, Bárbara Vargas da Rosa, Gisele Padro da Rosa, Gabriele do Carmo, Liziane Maria Barassuol Morandini, Gustavo Andrade Ugalde, Kátia Regina Kuhn, Ademir Farias Morel, Sérgio Luiz Jahn, Raquel Cristine Kuhn
The aim of this work was the production of bioactive metabolites by submerged fermentation from the fungus Diaporthe schini, followed by their extraction, separation and characterization. Different solvents (methanol, dichloromethane and hexane) were used for the extraction of metabolites from the fermentation broth and the extracts obtained were evaluated by in vitro antibacterial and antifungal activity. The separation and characterization of the extract from the hexane extraction was performed by column chromatography and GC-MS, respectively...
February 12, 2019: Journal of Biotechnology
Wei Chen, Yujuan Li, Guangjin Wu, Lingfeng Zhao, Li Lu, Peng Wang, Jianguang Zhou, Cheng Cao, Shanhu Li
Seamless modification of the Escherichia coli genome using positive selection / negative selection is widely used in metabolic engineering and functional genome analysis. Some excellent negative selection systems have been reported, of which tetA-sacB and inducible toxins system are prominent. To expand the existing negative selection toolkit, we constructed a new negative selection marker system based on kil gene of lambda prophage. The selection stringency of kil was measured and compared with the most widely used counter-selection gene, sacB, at the lacI, ack, and dbpa loci using different E...
February 12, 2019: Journal of Biotechnology
Wu Zou, Susan M Browne, Mohamed Al-Rubeai
Growth-arrested strategies (e.g. hypothermia and hyperosmolarity) have been widely employed to enhance cell-specific productivity (qP) in mammalian cell culture bioprocess. In addition to enhanced qP, alterations in cell physiology, such as cell size and cell cycle phase, have also attracted extensive attention under growth-arrested conditions. However, to date, very few reports on associations between physiological changes in growth-inhibiting approaches have been published. In this study, we explored associations between the physiological changes of GS-CHO cells in response to adenosine monophosphate (AMP) treatment...
February 12, 2019: Journal of Biotechnology
Jana Löwe, Olga Blifernez-Klassen, Thomas Baier, Lutz Wobbe, Olaf Kruse, Harald Gröger
Microbial consortia, which degrade branched, long-chain hydrocarbons, can be regarded as a promising source of novel enzymes for the stereo- and regio-selective oxyfunctionalization of hydrocarbons. The hydrocarbon-degrading bacterium Pimelobacter sp. Bb-B was isolated from the consortium associated with the colonial hydrocarbon-excreting microalga Botryococcus braunii. Three new type II flavoprotein monooxygenases (type II FMOs) from this bacterium have been made available in recombinant form through cloning and overexpression in an E...
January 28, 2019: Journal of Biotechnology
Yusuf B Johari, Adam J Brown, Christina S Alves, Yizhou Zhou, Chapman M Wright, Scott D Estes, Rashmi Kshirsagar, David C James
Synthetic promoters are an attractive alternative for use in mammalian hosts such as CHO cells as they can be designed de novo with user-defined functionalities. In this study, we describe and validate a method for bioprocess-directed design of synthetic promoters utilizing CHO genomic sequence information. We designed promoters with two objective features, (i) constitutive high-level recombinant gene transcription, and (ii) upregulated transcription under mild hypothermia or late-stage culture. CHO genes varying in transcriptional activity were selected based on a comparative analysis of RNA-Seq transcript levels in normal and biphasic cultures in combination with estimates of mRNA half-life from published genome scale datasets...
January 28, 2019: Journal of Biotechnology
Qian Gao, Xin Wang, Shewei Hu, Nana Xu, Mingjun Jiang, Chen Ma, Jianming Yang, Sheng Xu, Kequan Chen, Pingkai Ouyang
Approaches using metabolic engineering to produce D-1, 2, 4-butanetriol (BT) from renewable biomass in microbial systems have achieved initial success. However, due to the lack of incomplete understanding of the complex branch pathway, the efficient fermentation system for BT production was difficult to develop. Here we reconstituted a cell-free system in vitro using purified enzymes to produce BT from D-xylose. The factors that influencing the efficiency of cell-free system, including enzyme concentration, reaction buffer, pH, temperature, metal ion additives and cofactors were first identified to define optimal reaction conditions and essential components for the cascade reaction...
January 28, 2019: Journal of Biotechnology
Qingtao Liu, Xuerong Jin, Fang Fang, Jianghua Li, Guocheng Du, Zhen Kang
Enzymatic degradation of urea, the precursor of carcinogenic compound ethylcarbamate in rice wine, is always attractive. In the present study, we achieved high efficient production of Bacillus paralicheniformis iron-containing urease (Bp_Urease) in B. subtilis with the food-grade expression system. After reassembly of the urease gene cluster with inserting ribosome binding site (RBS), the production was increased from 38 U/L to 187 U/L. After altering the position of ureC and co-expressing the iron transporter encoding gene ureH, the activity was further increased to 1307 U/L...
January 28, 2019: Journal of Biotechnology
Dong-Fang Xie, Jun-Xing Yang, Chang-Jiang Lv, Jia-Qi Mei, Hong-Peng Wang, Sheng Hu, Wei-Rui Zhao, Jia-Ren Cao, Jun-Liang Tu, Jun Huang, Le-He Mei
Amine transaminases are a class of efficient and industrially-desired biocatalysts for the production of chiral amines. In this study, stabilized variants of the (R)-selective amine transaminase from Aspergillus terreus (AT-ATA) were constructed by consensus mutagenesis. Using Consensus Finder (, six positions with the most prevalent amino acid (over 60% threshold) among the homologous family members were identified. Subsequently, these six residues were individually mutated to match the consensus sequence (I77 L, Q97E, H210 N, N245D, G292D, and I295 V) using site-directed mutagenesis...
January 28, 2019: Journal of Biotechnology
Jana Husarcikova, Anett Schallmey
Bacterial β-etherases and glutathione lyases are glutathione-dependent enzymes that catalyze the selective cleavage of β-O-4 aryl ether bonds found in lignin. Their glutathione (GSH) dependence is regarded as major limitation for their application in the production of aromatics from lignin polymer and oligomers, as stoichiometric glutathione amounts are required. Thus, recycling of the GSH cofactor by a NAD(P)H-dependent glutathione reductase was proposed previously. Herein, the use of a whole-cell catalyst was studied for efficient β-O-4 aryl ether bond cleavage with intracellular GSH supply and recycling...
January 28, 2019: Journal of Biotechnology
Alexandra Yu Skorokhodova, Andrey Yu Gulevich, Vladimir G Debabov
An Escherichia coli K-12 MG1655-derived strain was engineered for respiro-fermentative production of pyruvate from glucose under anoxic conditions, which is preferred for industrial-scale microbial synthesis of valuable chemicals. The pathways of anaerobic pyruvate dissimilation were blocked in the strain by the deletion of the ackA, pta, poxB, ldhA, adhE, and pflB genes. The phosphoenolpyruvate-dependent phosphotransferase system of glucose transport and phosphorylation was substituted by an alternative ATP-dependent system resulting from the overexpression of galP and glk upon deletion of ptsG...
January 26, 2019: Journal of Biotechnology
Vanina Alzogaray, Mariela Urrutia, Paula Berguer, Andrés Rossi, Vanesa Zylberman, Romina Pardo, Hernán R Bonomi, Fernando A Goldbaum
Vaccination is as one of the most beneficial biopharmaceutical interventions against pathogens due to its ability to induce adaptive immunity through targeted activation of the immune system. Each vaccine needs a tailor-made set of tests in order to monitor its quality throughout the development and manufacturing. The analysis of the conformational state of protein nanoparticles is one of the key steps in vaccine quality control. The enzyme lumazine synthase from Brucella spp. (BLS) acts as a potent oral and systemic immunogen...
January 26, 2019: Journal of Biotechnology
Vicente Gotor-Fernández, Caroline E Paul
Deep eutectic solvents (DES) are a class of neoteric solvents used in multiple applications amongst which biocatalytic processes. Due to its simple preparation, low cost and inherent biodegradable properties, its use as a non-volatile biocompatible co-solvent with both whole cells and isolated enzymes has displayed increased enzyme activity and stability translating to higher product conversions and a surprising higher enantioselectivity in a range of biotransformations. This review lays out the latest updates on the use of DES in redox biocatalytic reactions...
January 26, 2019: Journal of Biotechnology
Beata Żyszka-Haberecht, Anna Poliwoda, Jacek Lipok
Halophilic and freshwater strains of cyanobacteria representing the Oscillatoriales, Nostocales, Chroococcales, and Synechococcales orders of Cyanophyta were examined to determine (i) the resistance of their cultures when suppressed by the presence of exogenous methoxylated and methylated derivatives of 2'-hydroxychalcone, (ii) morphological changes in cells treated with the tested chalcones and, most importantly, (iii) whether these photoautotrophic microorganisms transform chalcone derivatives in a structure- or strain-dependent manner...
January 25, 2019: Journal of Biotechnology
Julia F Hyslop, Sarah L Lovelock, Allan J B Watson, Peter W Sutton, Gheorghe-Doru Roiban
N-alkylated-α-amino acids are useful building blocks for the pharmaceutical and fine chemical industries. Enantioselective methods of N-alkylated-α-amino acid synthesis are therefore highly valuable and widely investigated. While there are a variety of chemical methods for their synthesis, they often employ stoichiometric quantities of hazardous reagents such as pyrophoric metal hydrides or genotoxic alkylating agents, whereas biocatalytic routes can provide a greener and cleaner alternative to existing methods...
January 25, 2019: Journal of Biotechnology
Wei Jiang, Ruonan Yang, Peng Lin, Wenjing Hong, Baishan Fang
A bioinspired strategy for the synthesis of supramolecular and biocatalytical materials was developed base on protein-protein supramolecular interaction and genetic engineering. Formate dehydrogenase (FDH) and its functional fragments were separately fused to form a multi-function domain. The fusion proteins and functional fragments self-assembled into the expanded and controllable supramolecular interaction networks. Morphology characterization by scanning-electron microscopy showed that the assembled functional fragments and fusion proteins formed multi-dimensional (3D) and two-dimensional (2D) layer-like structures...
January 25, 2019: Journal of Biotechnology
Tanyaporn Siriwong, Bustomi Laimeheriwa, Uyun Nurul Aini, Muhammad Nur Cahyanto, Alissara Reungsang, Apilak Salakkam
The present study investigated cold hydrolysis of cassava pulp (CP) and the use of cold hydrolysis with simultaneous saccharification and fermentation (SSF) for ethanol production. Cold hydrolysis of 100 g-CP/L at 50 °C for 2 h, followed by at 30 °C for 72 h resulted in the production of 71.5 ± 1.8 g/L of reducing sugar, with a yield of 0.72 g/g-CP. A mathematical model describing the cold hydrolysis process was subsequently developed. The model proved to be applicable for other cold hydrolysis systems with satisfactory results...
January 25, 2019: Journal of Biotechnology
Hui Li, Yan Ni, Xun Cao, Xun He, Ganlu Li, Kequan Chen, Pingkai Ouyang, Jianming Yang, Weiming Tan
Metal-driven papain-surfactant nanocomposite (PA@MSNC), a novel soft nanobiocatalyst, was successfully prepared via one-pot self-assembly technique in aqueous solution for the biosynthesis of N-(benzyloxycarbonyl)-L-alanyl-L-glutamine (Z-Ala-Gln) dipeptide in deep eutectic solvents (DESs). The metal-driven self-assembly process generated PA@MSNC as nanospheres of ˜130 nm in diameter, with high protein loading and relative enzyme activity of 420 mg/g and 80% (4270 U/g protein), respectively. PA@MSNC showed high apparent substrate affinity and catalytic efficiency...
January 25, 2019: Journal of Biotechnology
Xuguo Duan, Xinyi Zhang, Zhenyan Shen, Erzheng Su, Linguo Zhao, Jianjun Pei
In this study, a combined optimization strategy, based on co-expression of molecular chaperones and supplementation of osmolytes, was used to reduce the formation of inclusion bodies and enhance the expression of the soluble form of 4-α-glucanotransferase. The 4-α-glucanotransferase yield was enhanced by co-expression with pGro7 and supplementation of trimetlylamine oxide. Subsequently, the effects of process conditions (temperature, inducer concentration, and arabinose concentration) on cell growth and 4-α-glucanotransferase production were also investigated in shake flasks...
January 25, 2019: Journal of Biotechnology
Dulce Andrade-Pavón, Jossue Ortiz-Álvarez, Eugenia Sánchez-Sandoval, Joaquín Tamariz, César Hernández-Rodríguez, J Antonio Ibarra, Lourdes Villa-Tanaca
Due to increasing resistance of Candida species to antifungal drugs, especially azoles, new drugs are needed. The proposed compounds 3 and 4 are analogous to α-asarone (2), a naturally occurring potent inhibitor of HMGR with hypolipidemic and antifungal activity. We used the recombinant enzyme 3-hydroxy-3-methylglutaryl coenzyme A reductase of Candida glabrata (CgHMGR) as a model to test the effectiveness of the test compounds. Compounds 3 and 4 demonstrated inhibitory kinetics, having lower IC50 values (42...
January 25, 2019: Journal of Biotechnology
Mateusz Wojtusik, Priscilla Vergara, Juan C Villar, Felix Garcia-Ochoa, Miguel Ladero
The stabilization effects on a novel commercial β-glucosidase preparation from Aspergillus fumigatus during saccharification of ethanol-water pretreated wheat straw were analysed in comparison to this enzyme stability during cellobiose hydrolysis. For this purpose, the kinetics of β-glucosidase residual activity during cellobiose hydrolysis from 40 till 70 °C were studied, resulting in the fitting of a first-order partial deactivation model. Furthermore, a subsequent fitting of a kinetic model including this first-order deactivation equation and a Michaelis-Menten equation with double competitive inhibition by glucose and uncompetitive inhibition by cellobiose to released glucose was successful...
January 19, 2019: Journal of Biotechnology
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