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Biotechnology Journal

Anamaria Todea, Diana Aparaschivei, Valentin Badea, Carmen G Boeriu, Francisc Peter
evelopments of past years placed the bio-based polyesters as competitive substitutes for fossil-based polymers. Moreover, enzymatic polymerization using lipase catalysts has become an important green alternative to chemical polymerization for the synthesis of polyesters with biomedical applications, as several drawbacks related to the presence of traces of metal catalysts, toxicity and higher temperatures could be avoided. Copolymerization of ε-caprolactone (CL) with four hydroxy-fatty acids (HFA) from renewable sources, e...
March 15, 2018: Biotechnology Journal
Luhong He, Jairav Xolo Desai, Jinxin Gao, Laurie B Hazeltine, Zhirui Lian, John N Calley, Christopher C Frye
Oxidation of monoclonal antibodies (mAb) is a common chemical modification with potential impact on a therapeutic protein's activity and immunogenicity. In a previous study, it was found that tryptophan oxidation (Trp-ox) levels of two mAb produced in Chinese hamster ovary (CHO) cells were significantly lowered by modifying cell culture medium/feed. In this study, transcriptome analysis by RNA-Seq was applied to further elucidate the underlying mechanism of those changes in lowering the Trp-ox levels. Cell samples from the 5L fed-batch conditions were harvested and subjected to RNA-Seq analysis...
March 15, 2018: Biotechnology Journal
Rita B Santos, Balakumaran Chandrasekar, Manoj K Mandal, Farnusch Kaschani, Markus Kaiser, Leonard Both, Renier Al van der Hoorn, Andreas Schiermeyer, Rita Abranches
Medicago truncatula is an established model for studying legume biology. More recently, it has also been exploited as a Molecular Farming platform for the production of recombinant proteins, with the successful expression of fungal and human proteins in plants and cell suspension cultures of this species. One of the challenges that now must be overcome is the degradation of final products during production and downstream processing stages. In the Medicago truncatula genome, there are more than 400 putative protease-encoding genes, but to date, the proteolytic content of Medicago cell cultures has not been studied...
March 12, 2018: Biotechnology Journal
Amr S Ali, Ravali Raju, Somak Ray, Rashmi Kshirsagar, Alan Gilbert, Li Zang, Barry L Karger
As the demand for biological therapeutic proteins rises, there is an increasing need for robust and highly efficient bioprocesses, specifically, maximizing protein production by controlling the cellular nutritional and metabolic needs. A comprehensive lipidomics analysis has been performed, for the first time, over the time course of CHO cells producing an IgG1 monoclonal antibody (mAb) with fed batch 5 L bioreactors. The dynamic nature and importance of the CHO lipidome, especially on cellular growth and specific productivity, is demonstrated...
March 9, 2018: Biotechnology Journal
Min Liu, Zhiyu Chen, Sisi Huang, Shenghui Chu, Nipatha Issaro, Haishan Tian, Haiyan Hu, Chao Jiang
The drug development of FGF7 has been restricted by its toxicity to the host, low expression, poor stability, and easy degradation. Recent studies have shown that Halo-tag-flanked recombinant human FGF7 can solve the problem of toxicity; however, its biological activity is unknown.This study aimed to explore the activity of Halo-rhFGF7 and rhFGF7 on acute liver injury in vitro and in vivo. The rhFGF7 was expressed with a N-terminal Halo-tag, followed by a tobacco etch virus (TEV) protease cleavage site, in Escherichia coli BL21 (DE3) pLysS in this study...
March 5, 2018: Biotechnology Journal
Yang Ji, Yuan Lu, Yishu Yan, Xinxin Liu, Nan Su, Chong Zhang, Shengli Bi, Xin-Hui Xing
The Ebola hemorrhagic fever caused by Ebola virus is an extremely dangerous disease, and effective therapeutic agents are still lacking. Platforms for the efficient production of vaccines are crucial to ensure quick response against an Ebola virus outbreak. Ebolavirus glycoprotein (EbolaGP) on the virion surface is responsible for membrane binding and virus entry, thus becoming the key target for vaccine development. However, heterologous expression of this protein still faces engineering challenges such as low production levels and insoluble aggregation...
March 3, 2018: Biotechnology Journal
James M Wagner, Eden V Williams, Hal S Alper
Yarrowia lipolytica is a non-conventional yeast of interest to the biotechnology industry. However, the physiology, metabolism, and genetic regulation of Y. lipolytica diverge significantly from more well-studied and characterized yeasts such as Saccharomyces cerevisiae. To develop additional genetic tools for this industrially relevant host, we adapt the piggyBac transposon system to enable efficient generation of genome-wide insertional mutagenesis libraries and introduction of scarless, footprint-free genomic modifications in Y...
March 1, 2018: Biotechnology Journal
Feifan Yu, Veronica Alesand, Per-Åke Nygren
Protein fragment complementation assays (PCA) rely on a proximity-driven reconstitution of a split reporter protein activity, typically via interaction between bait and prey units separately fused to the reporter protein halves. The PCA principle can also be formatted for use in immunossays for analyte detection, e.g. via the use of small immunoglobulin binding proteins (IgBp) as fusion partners to split-reporter protein fragments for conversion of pairs of antibodies into split-protein half-probes. However, the non-covalent binding between IgBp and antibodies is not ideal for development of robust assays...
February 27, 2018: Biotechnology Journal
Jaehyun Cha, Inchan Kwon
Protein immobilization has been widely used for laboratory experiments and industrial processes. Preparation of a recombinant protein for immobilization usually requires laborious and expensive purification steps. Here we report a novel purification-free, target-selective immobilization technique of a protein from cell lysates. Purification steps were skipped by immobilizing a target protein containing a clickable non-natural amino acid (p-azidophenylalanine) in cell lysates onto alkyne-functionalized solid supports via bioorthogonal azide-alkyne cycloaddition...
February 27, 2018: Biotechnology Journal
Feng Cheng, Xiao-Ling Tang, Tsvetan Kardashliev
The molecular mechanisms that cells use to sense changes in the intra- and extracellular environment are increasingly utilized in synthetic biology to build genetic reporter constructs for various applications. Although in nature sensing can be RNA-mediated, most existing genetically-encoded biosensors are based on transcription factors (TF) and cognate DNA sequences. Here, we discuss recent advances in the integration of TF-based biosensors in metabolic and protein engineering screens whereas distinction is made between production-driven and competitive screening systems for enzyme evolution under physiological conditions...
February 27, 2018: Biotechnology Journal
Hyun Aaron Kim, Hyun Ju Kim, Min Ju Lee, Jihoon Park, Ah Reum Choi, Haeyoung Jeong, Kwang-Hwan Jung, Pil Kim, Sang Jun Lee
We reported that the phototrophic metabolism via plasmid-originated Gloeobacter rhodopsin(GR)-expression is improved in Escherichia coli ET5 harboring pKJ606-GR by a genomic point mutation (dgcQC1082A ) encoding a transmembrane cell signaling protein (Microb. Cell Fact. 16:111, 2017). Another evolved descendant was isolated from the chemostat, and the genome variation of the strain named ET8 harboring pKJ606-GR was investigated in this study. Whole genome sequencing analysis identified a single point mutation (C3831976A) located in the non-coding upstream region of kdtA and an IS4 insertional mutation at galUG706 without any mutations in the plasmid...
February 22, 2018: Biotechnology Journal
Alois Jungbauer
New approaches for vaccination must be developed in order to meet the grand challenges for emerging infectious diseases. Exosomes now enter vaccine development and these are strategies are meeting these global challenges, as demonstrated by Anticoli et al, in this issue of Biotechnology Journal [1] Using exosome vaccines has been now been demonstrated in-vivo for several viruses such as Ebola Virus VP24, VP40 and NP, Influenza Virus NP, Crimean-Congo Hemorrhagic Fever NP, West Nile Virus NS3, and Hepatitis C Virus NS3...
February 22, 2018: Biotechnology Journal
Marina Lotti, Jürgen Pleiss, Francisco Valero, Pau Ferrer
Lipase-catalyzed transesterification of triglycerides and alcohols to obtain biodiesel is an environmentally friendly and sustainable route for fuels production since, besides proceeding in mild reaction conditions, it allows for the use of low-cost feedstocks that contain water and free fatty acids, for example non-edible oils and waste oils. This review article reports recent advances in the field and focus in particular on a major issue in the enzymatic process, the inactivation of most lipases caused by methanol, the preferred acyl acceptor used for alcoholysis...
February 20, 2018: Biotechnology Journal
Yu Liu, Xin Zhang
Proper regulation of protein homeostasis (proteostasis) is essential to maintain cellular fitness. Proteome stress causes imbalance of the proteostasis, leading to various diseases represented by neurodegenerative diseases, cancers, and metabolic disorders. The biosensor community recently embarked on the development of proteome stress sensors to report on the integrity of proteostasis in live cells. While most of these sensors are based on metastable mutants of specific client proteins, a recent sensor takes advantage of the specific association of heat shock protein 27 with protein aggregates and exhibits a diffusive to punctate fluorescent change in cells that are subjected to stress conditions...
February 20, 2018: Biotechnology Journal
Xuanwei Ding, Dong Liu, George Booth, Wei Gao, Yuan Lu
As mimicking natural virus structures, virus-like particles (VLPs) have evolved to become a widely accepted technology used for humans which are safe, highly efficacious and profitable. Several remarkable advantages have been achieved to revolutionize the molecule delivery for diverse applications in nanotechnology, biotechnology, and medicine. Here, we review the rational structure design, manufacturing process, functionalization strategy, and emerging applications of VLPs. The situation and challenges in the VLP engineering, the key development orientation, and future applications have been discussed...
February 17, 2018: Biotechnology Journal
Jiazhang Lian, Mohammad HamediRad, Huimin Zhao
Thanks to its ease of use, modularity, and scalability, the clustered regularly interspaced short palindromic repeats (CRISPR) system has been increasingly used in the design and engineering of Saccharomyces cerevisiae, one of the most popular hosts for industrial biotechnology. This review summarizes the recent development of this disruptive technology for metabolic engineering applications, including CRISPR-mediated gene knock-out and knock-in as well as transcriptional activation and interference. More importantly, multi-functional CRISPR systems that combine both gain- and loss-of-function modulations for combinatorial metabolic engineering are highlighted...
February 13, 2018: Biotechnology Journal
Iris Delrue, Qiubao Pan, Anna K Baczmanska, Bram W Callens, Lia Lm Verdoodt
Choosing a potent selection antibiotic (SA), is a crucial success factor when creating stably transfected cell lines using an antibiotic selection marker. The selection capacity of this antibiotic is defined as its ability to kill sensitive, untransfected parental cells, while leaving resistant, transfected cells unharmed. Currently, no procedure has been described to determine this selection capacity. Therefore, we developed a protocol to obtain a numerical value, called the "selectivity factor" (SF), that defines the selection capacity of SAs...
February 13, 2018: Biotechnology Journal
Han Liu, Guochen Fang, Hui Wu, Zhimin Li, Qin Ye
L-cysteine is an amino acid with important physiological functions and has a wide range of applications in medicine, food, animal feed and cosmetics industry. In this study, the L-cysteine synthesis in E. coli was divided into four modules: the transport module, sulfur module, precursor module, and degradation module. The engineered strain LH03 (overexpression of the feedback-insensitive cysE and the exporter ydeD in JM109) accumulated 45.8 mg/L of L-cysteine in 48 hours with yield of 0.4% g/g glucose. Further modifications of strains and culture conditions which based on the rational metabolic engineering and modular strategy improved the L-cysteine biosynthesis significantly...
February 6, 2018: Biotechnology Journal
Allison G McAtee Pereira, Jason L Walther, Myles Hollenbach, Jamey D Young
13 C metabolic flux analysis (MFA) provides a rigorous approach to quantify intracellular metabolism of industrial cell lines. In this study, 13 C MFA was used to characterize the metabolic response of Chinese hamster ovary (CHO) cells to a novel medium variant designed to reduce ammonia production. Ammonia inhibits growth and viability of CHO cell cultures, alters glycosylation of recombinant proteins, and enhances product degradation. Ammonia production was reduced by manipulating the amino acid composition of the culture medium; specifically, glutamine, glutamate, asparagine, aspartate, and serine levels were adjusted...
February 6, 2018: Biotechnology Journal
Mihris Ibnu Saleem Naduthodi, Maria J Barbosa, John van der Oost
The carbon footprint caused by unsustainable development and its environmental and economic impact has become a major concern in the past few decades. Photosynthetic microbes such as microalgae and cyanobacteria are capable of accumulating value-added compounds from carbon dioxide, and have been regarded as environmentally friendly alternatives to reduce the usage of fossil fuels, thereby contributing to reducing the carbon footprint. This light-driven generation of green chemicals and biofuels has triggered the research for metabolic engineering of these photosynthetic microbes...
February 3, 2018: Biotechnology Journal
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