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Biotechnology Progress

Fabian Feidl, Simone Garbellini, Sebastian Vogg, Michael Sokolov, Jonathan Souquet, Hervé Broly, Alessandro Butté, Massimo Morbidelli
On-line monitoring tools for downstream chromatographic processing (DSP) of biotherapeutics can enable fast actions to correct for disturbances in the upstream, gain process understanding and eventually lead to process optimization. While UV/Vis spectroscopy is mostly assessing the protein's amino acid composition and the application of Fourier Transform Infrared spectroscopy is limited due to strong water interactions, Raman spectroscopy is able to assess the secondary and tertiary protein structure without significant water interactions...
May 17, 2019: Biotechnology Progress
Yanhong Chang, Shuangming Tong, Hui Luo, Zijia Liu, Bo Qin, Linlin Zhu, Hongxu Sun, Huimin Yu, Zhongyao Shen
In a stirred tank reactor (STR), during catalysis with immobilized cephalosporin C acylase (CCA), the microenvironmental pH dropped to 7.2 in a non-buffered system (with the pH maintained at 8.5 by adding alkali) due to the existence of diffusional resistance. Moreover, the immobilized CCA only catalyzed five batch reactions, suggesting that the sharp pH gradient impaired the enzyme stability. To buffer the protons produced in the hydrolysis of cephalosporin C by CCA, phosphate and bicarbonate buffers were introduced...
May 17, 2019: Biotechnology Progress
E-Ching Ong, Pauline Smidt, Jeffrey T McGrew
In order to avoid the metabolic burden of protein expression during cell growth, and to avoid potential toxicity of recombinant proteins, microbial expression systems typically utilize regulated expression vectors. In contrast, constitutive expression vectors have usually been utilized for isolation of protein expressing mammalian cell lines. In mammalian systems, inducible expression vectors are typically utilized for only those proteins that are toxic when over-expressed. We developed a tetracycline regulated expression system in CHO cells, and show that cell pools selected in the uninduced state recover faster than those selected in the induced state even though the proteins showed no apparent toxicity or expression instability...
May 15, 2019: Biotechnology Progress
Paloma Andrade M Nascimento, Flávio Pereira Picheli, André Moreni Lopes, Jorge Fernando Brandão Pereira, Valéria de Carvalho Santos-Ebinuma
Lipases are well-known biocatalysts used in several industrial processes/applications. Thus, as with other enzymes, changes in their surrounding environment and/or their thermodynamic parameters can induce structural changes that can increase, decrease, or even inhibit their catalytic activity. The use of ionic compounds as solvents or additives is a common approach for adjusting reaction conditions and, consequently, for controlling the biocatalytic activity of enzymes. Herein, to elucidate the effects of ionic compounds on the structure of lipase, the stability and enzymatic activity of lipase from Aspergillus niger in aqueous solutions (at 0...
May 14, 2019: Biotechnology Progress
Sotirios-Spyridon Vamvakas, John Kapolos, Lambros Farmakis, Gregoria Koskorellou, Fotios Genneos
The genetic modification of yeast strains can be used as an approach for the improvement of ethanol fermentation. msn2 transcription factor is implicated in yeast stress response and its activation is controlled by Proteine Kinase A (PKA). PKA activation inhibiting the translocation of msn2 to the nucleus. An in silico analysis of msn2 protein sequence revealed serine residue at position 625 as a potent target of PKA. The substitution of this serine residue with alanine has as a result the increase of susceptibility of the cells to ethanol challenge reducing IC50 from 3% v/v to 2...
May 14, 2019: Biotechnology Progress
Kao-Pang Hsu, Shih-I Tan, Chen-Yaw Chiu, Yu-Kaung Chang, I-Son Ng
Carbonic anhydrase (CA, EC is an ancient enzyme with zinc ion as its active site, which catalyzes the chemical reaction of carbon dioxide (CO2 ) to react with water and form bicarbonate ions. Due to its high catalytic efficiency on CO2 assimilation, CA is expected to use for carbon sequestration in industry. However, the protein expression level, thermostability and high throughput screening of an active CA are still with difficulty. In this study, the CA from Sulfurihydrogenibium yellowstonense (denoted as SyCA) was selected for over-expressed in E...
May 10, 2019: Biotechnology Progress
Shyam B Mehta, Srishty Subramanian, Rowena D'Mello, Charlene Brisbane, Shouvik Roy
Monoclonal antibodies may be prone to self-association leading to formation of dimers, trimers or other high molecular weight species (HMWs) during bio-processing. In order to implement appropriate manufacturing control strategies during bio-processing, it is important to understand various real life bio-processing conditions where such self-associations may manifest. One such case study is presented here of increase in dimer content for a monoclonal antibody during scale-up bio-processing and the approach taken to understand the under-lying mechanism...
May 7, 2019: Biotechnology Progress
Parisa Farrokh, Mojgan Sheikhpour, Alibakhsh Kasaeian, Hassan Asadi, Roya Bavandi
Cyanobacteria are photosynthetic microorganisms which can be found in various environmental habitats. These photosynthetic bacteria are considered as promising feedstock for the production of the third and the fourth generation of biofuels. The main subject of this review is highlighting the significant aspects of the biofuel production from cyanobacteria. The most recent investigations about the extraction or separation of the bio-oil from cyanobacteria are also adduced in the present review. Moreover, the genetic engineering of cyanobacteria for improving biofuel production and the impact of bioinformatics studies on the designing better-engineered strains are mentioned...
May 7, 2019: Biotechnology Progress
Jinsung Hong, Jacob Demirji, Daniel Blackstock, James Lee, Tracey Dinh, Alvenne Goh, Frank Arnold, Joe Horwitz
An alternating tangential flow (ATF) perfusion-based transient gene expression (TGE) bioprocess has been developed using human embryonic kidney (HEK) 293 cells to produce H1-ss-np, a promising candidate for a universal influenza vaccine. Two major adjustments were taken to improve the process: 1) eliminate the interference of microbubbles during gene transfection; and 2) utilize an ATF perfusion system for a prolonged culture period. As a result, a closed-operation 9-days ATF perfusion-based TGE bioprocess was developed...
May 3, 2019: Biotechnology Progress
Steffen Kreye, Rainer Stahn, Karina Nawrath, Vicky Goralczyk, Barney Zoro, Steffen Goletz
Continuous upstream processing in mammalian cell culture for recombinant protein production holds promise to increase product yield and quality. To facilitate the design and optimization of large-scale perfusion cultures, suitable scale-down mimics are needed which allow high-throughput experiments to be performed with minimal raw material requirements. Automated microbioreactors are available that mimic batch and fed-batch processes effectively but these have not yet been adapted for perfusion cell culture...
May 3, 2019: Biotechnology Progress
Poliana Linzmeyer, Heloisa Ramlow, Ozair Souza, Noeli Sellin, Cintia Marangoni
In the present work, we carried out recovery of ethanol from a fermentation broth of banana pseudostem by pervaporation (PV) as a lower-energy-cost alternative to traditional separation processes such as distillation. Since real fermentation systems generally contain by-products, it was investigated the effects of different components from the fermentation broth of banana pseudostem on PV performance for ethanol recovery through commercial flat sheet polydimethylsiloxane (PDMS) membrane. The experiments were compared to a binary solution (ethanol/water) to determine differences in the results due to the presence of fermentation by-products...
May 3, 2019: Biotechnology Progress
Fatma Rmili, Neila Achouri, Nabil Smichi, Najeh Krayem, Ahmed Bayoudh, Youssef Gargouri, Mohamed Chamkha, Ahmed Fendri
A mesophilic bacterial culture, producing an extracellular alkaline lipase, was isolated from the gas-washing wastewaters generated from the Sfax phosphate plant of the Tunisian Chemical Group and identified as Staphylococcus capitis strain. The lipase, named SCL, has been purified to homogeneity from the culture medium. The purified enzyme molecular weight was around 45 kDa. Specific activities about 3900 U/mg and 500 U/mg were measured using tributyrin and olive oil emulsion as substrates, respectively at 37°C and pH 8...
May 3, 2019: Biotechnology Progress
J A Elegbede, A Lateef, M A Azeez, T B Asafa, T A Yekeen, I C Oladipo, S H Abbas, L S Beukes, E B Gueguim-Kana
The search for biocompatible nanoparticles with vast applicability has impacted on exploration of various biomaterials for the synthesis of mono and bimetallic nanoparticles. Xylanase is widely regarded as an industrially important enzyme but its potential in nanotechnological applications are yet to be fully explored. The current study investigates the exploit of xylanases of Aspergillus niger L3 (NE) and Trichoderma longibrachiatum L2 (TE) produced through valorization of corn-cob, to synthesize silver-gold alloy nanoparticles (Ag-AuNPs)...
May 3, 2019: Biotechnology Progress
Muhammad Aasim, Muhammad H Khan, Inam U Rahman, Noor Shad Bibi, Waqar Ali, Nadir Z Khan, Abid A Khan
Hydrophobic interaction chromatography, an important and effective purification strategy, is generally used for the purification of variety of biomolecules. A basic understanding of the protein interaction behaviour is required to effectively separate these biomolecules. A colloidal type extended Derjaguin, Landau, Verwey, and Overbeek calculations were utilized to study the interactions behavior of model proteins to commercially available hydrophobic chromatographic materials i.e. toyopearl phenyl 650-C and toyopearl butyl 650-C...
April 26, 2019: Biotechnology Progress
Hongling Liu, Xihui Wang, Shaojie Yang, Ruiming Wang, Tengfei Wang
Trehalose is a non-reducing disaccharide synthesized by trehalose synthase (TreS), which catalyzes the reversible interconversion of maltose and trehalose. We aimed to enhance the catalytic conversion of maltose to trehalose by saturation mutagenesis, and constructed a self-inducible TreS expression system by generating a robust Bacillus subtilis recombinant. We found that the conversion yield and enzymatic activity of TreS was enhanced by saturation mutations, especially by the combination of V407M and K490L mutations...
April 25, 2019: Biotechnology Progress
Lasse Jannis Frey, David Vorländer, Detlev Rasch, Hendrik Ostsieker, Bernhard Müller, Moritz Schulze, René Schenkendorf, Torsten Mayr, Jan-Hendrik Grosch, Rainer Krull
Micro-bioreactors (MBRs) have become an indispensable part for modern bioprocess development enabling automated experiments in parallel while reducing material cost. Novel developments aim to further intensify the advantages as dimensions are being reduced. However, one factor hindering the scale-down of cultivation systems is to provide adequate mixing and mass transfer. Here, vertical oscillation is demonstrated as an effective method for mixing of MBRs with a reaction volume of 20 μL providing adequate mass transfer...
April 25, 2019: Biotechnology Progress
Swarnim Ranjan, Wai Keen Chung, Min Zhu, David Robbins, Steven M Cramer
This work focused on the development of a combined experimental and computational tool set to study protein-mAb interactions. A model protein library was first screened using cross interaction chromatography to identify proteins with the strongest retention. Fluorescence polarization was then employed to study the interactions and thermodynamics of the selected proteins - lactoferrin, pyruvate kinase and ribonuclease B with the mAb. Binding affinities of lactoferrin and pyruvate kinase to the mAb were seen to be relatively salt insensitive in the range examined...
April 24, 2019: Biotechnology Progress
T V Surendra, Selvaraj Mohana Roopan, Maksudur Rahman Khan
The rare earth metal oxide nanoparticles such as gadolinium oxide nanoparticles (Gd2 O3 NPs) have been synthesized by green synthesis process using methanolic extract of Moringa oleifera (M. oleifera) peel. In this process, the Gd2 O3 NPs formation was observed at 280-300 nm in UV-Vis spectroscopy. The XRD pattern of the synthesized Gd2 O3 NPs was exactly matched with JCPDS No 3-065-3181which confirms the crystalline nature of Gd2 O3 NPs. Also, EDX analysis was stated that Gd and O elements were present as 70...
April 24, 2019: Biotechnology Progress
Paul S Kelly, Noemi Dorival-García, Samantha Paré, Sara Carillo, Christine Ta, Antonio Alarcon Miguez, Orla Coleman, Emma Harper, Maeve Shannon, Michael Henry, Lisa Connolly, Martin Clynes, Paula Meleady, Jonathan Bones, Niall Barron
Single-use technologies, in particular disposable bioreactor bags, have become integral within the biopharmaceutical community. However, safety concerns arose upon the identification of toxic leachable compounds derived from the plastic materials. Although the leachable bis(2,4-di-tert-butylphenyl)-phosphate (bDtBPP) has been previously shown to inhibit CHO cell growth, it is critical to determine if other compounds like this are still present in subsequent generations of films for industrial application. This study compares the performance of CHO cells, CHO-K1 and CHO- DP12, cultured in media conditioned in an older single-use bioreactor (SUB) film (F-1) and a newer generation film (F-2) from the same vendor...
April 24, 2019: Biotechnology Progress
Gorawit Yusakul, Seiichi Sakamoto, Hiroyuki Tanaka, Satoshi Morimoto
The assembly between heavy and light chains is a critical step of immunoglobulin (Ig) and fragment antigen-binding (Fab) antibody expression and of their binding activity. The genes encoding Fab were obtained from hybridoma cells secreting monoclonal antibody (MAb, IgG2b) against adenylate cyclase activator forskolin (FOR). The subclass of the first constant domain of heavy chain (CH 1) of IgG2b was modified to IgG1 via overlap extension polymerase chain reaction (OE-PCR) and expressed via Escherichia coli bacterial system...
April 22, 2019: Biotechnology Progress
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