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Heterologous expression lab

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https://www.readbyqxmd.com/read/29922971/surface-display-of-heterologous-%C3%AE-galactosidase-in-food-grade-recombinant-lactococcus-lactis
#1
Supeng Yin, Hongbin Zhu, Mengyu Shen, Gang Li, Shuguang Lu, Yan Zhao, Shuai Le, Yinling Tan, Yizhi Peng, Fuquan Hu, Jing Wang
β-Galactosidase is an essential enzyme for the metabolism of lactose in human beings and has an important role in the treatment of lactose intolerance (LI). β-Galactosidase expressed by intestinal microflora, such as lactic acid bacteria (LAB), also alleviates LI. A promising approach to LI management is to exploit a food-grade LAB delivery system that can inhabit the human intestine and overproduce β-galactosidase. In this study, we constructed a food-grade β-galactosidase surface display delivery system and then integrated into the chromosome of Lactococcus lactis (L...
June 19, 2018: Current Microbiology
https://www.readbyqxmd.com/read/29910943/self-resistance-guided-genome-mining-uncovers-new-topoisomerase-inhibitors-from-myxobacteria
#2
Fabian Panter, Daniel Krug, Sascha Baumann, Rolf Müller
There is astounding discrepancy between the genome-inscribed production capacity and the set of known secondary metabolite classes from many microorganisms as detected under laboratory cultivation conditions. Genome-mining techniques are meant to fill this gap, but in order to favor discovery of structurally novel as well as bioactive compounds it is crucial to amend genomics-based strategies with selective filtering principles. In this study, we followed a self-resistance guided approach aiming at the discovery of inhibitors of topoisomerase, known as valid target in both cancer and antibiotic therapy...
June 7, 2018: Chemical Science
https://www.readbyqxmd.com/read/29795673/uncovering-production-of-specialized-metabolites-by-streptomyces-argillaceus-activation-of-cryptic-biosynthesis-gene-clusters-using-nutritional-and-genetic-approaches
#3
Adriana Becerril, Susana Álvarez, Alfredo F Braña, Sergio Rico, Margarita Díaz, Ramón I Santamaría, José A Salas, Carmen Méndez
Sequencing of Streptomyces genomes has revealed they harbor a high number of biosynthesis gene cluster (BGC), which uncovered their enormous potentiality to encode specialized metabolites. However, these metabolites are not usually produced under standard laboratory conditions. In this manuscript we report the activation of BGCs for antimycins, carotenoids, germicidins and desferrioxamine compounds in Streptomyces argillaceus, and the identification of the encoded compounds. This was achieved by following different strategies, including changing the growth conditions, heterologous expression of the cluster and inactivating the adpAa or overexpressing the abrC3 global regulatory genes...
2018: PloS One
https://www.readbyqxmd.com/read/29785668/hac1p-homologues-from-higher-eukaryotes-can-improve-the-secretion-of-heterologous-proteins-in-the-yeast-pichia-pastoris
#4
Olufemi Emmanuel Bankefa, Meiyu Wang, Taicheng Zhu, Yin Li
OBJECTIVES: To systematically explore the effects of overexpressing Hac1p homologues from different sources on protein secretion in Pichia pastoris system. RESULTS: Effects of Hac1p homologues encompassing P. pastoris (PpHac1p), S. cerevisiae (ScHac1p), Trichoderma reesei (TrHac1p) and Homo sapiens (HsXbp1), on secretion of three reporter proteins-β-galactosidase, β-mannanase and glucose oxidase were investigated. No individual Hac1p was optimal for all the enzymes...
May 21, 2018: Biotechnology Letters
https://www.readbyqxmd.com/read/29785587/a-study-on-the-use-of-strain-specific-and-homologous-promoters-for-heterologous-expression-in-industrial-saccharomyces-cerevisiae-strains
#5
Daniel Pereira de Paiva, Tiago Benoliel Rocha, Marciano Regis Rubini, André Moraes Nicola, Viviane Castelo Branco Reis, Fernando Araripe Gonçalves Torres, Lidia Maria Pepe de Moraes
Polymorphism is well known in Saccharomyces cerevisiae strains used for different industrial applications, however little is known about its effects on promoter efficiency. In order to test this, five different promoters derived from an industrial and a laboratory (S288c) strain were used to drive the expression of eGFP reporter gene in both cells. The ADH1 promoter (P ADH1 ) in particular, which showed more polymorphism among the promoters analyzed, also exhibited the highest differences in intracellular fluorescence production...
May 21, 2018: AMB Express
https://www.readbyqxmd.com/read/29769376/-lactobacillus-mucosal-vaccine-vectors-immune-responses-against-bacterial-and-viral-antigens
#6
REVIEW
Jonathan S LeCureux, Gregg A Dean
Lactic acid bacteria (LAB) have been utilized since the 1990s for therapeutic heterologous gene expression. The ability of LAB to elicit an immune response against expressed foreign antigens has led to their exploration as potential mucosal vaccine candidates. LAB vaccine vectors offer many attractive advantages: simple, noninvasive administration (usually oral or intranasal), the acceptance and stability of genetic modifications, relatively low cost, and the highest level of safety possible. Experimentation using LAB of the genus Lactobacillus has become popular in recent years due to their ability to elicit strong systemic and mucosal immune responses...
June 27, 2018: MSphere
https://www.readbyqxmd.com/read/29718215/large-inserts-for-big-data-artificial-chromosomes-in-the-genomic-era
#7
Arianna Tocchetti, Stefano Donadio, Margherita Sosio
The exponential increase in available microbial genome sequences coupled with predictive bioinformatic tools is underscoring the genetic capacity of bacteria to produce an unexpected large number of specialized bioactive compounds. Since most of the biosynthetic gene clusters (BGCs) present in microbial genomes are cryptic, i.e. not expressed under laboratory conditions, a variety of cloning systems and vectors have been devised to harbor DNA fragments large enough to carry entire BGCs and to allow their transfer in suitable heterologous hosts...
May 1, 2018: FEMS Microbiology Letters
https://www.readbyqxmd.com/read/29660472/enhanced-cellobiose-fermentation-by-engineered-saccharomyces-cerevisiae-expressing-a-mutant-cellodextrin-facilitator-and-cellobiose-phosphorylase
#8
Heejin Kim, Eun Joong Oh, Stephan Thomas Lane, Won-Heong Lee, Jamie H D Cate, Yong-Su Jin
To efficiently ferment intermediate cellodextrins released during cellulose hydrolysis, Saccharomyces cerevisiae has been engineered by introduction of a heterologous cellodextrin utilizing pathway consisting of a cellodextrin transporter and either an intracellular β-glucosidase or a cellobiose phosphorylase. Among two types of cellodextrin transporters, the passive facilitator CDT-2 has not enabled better cellobiose fermentation than the active transporter CDT-1, which suggests that the CDT-2 might be engineered to provide energetic benefits over the active transporter in cellobiose fermentation...
June 10, 2018: Journal of Biotechnology
https://www.readbyqxmd.com/read/29621349/signature-pathway-expression-of-xylose-utilization-in-the-genetically-engineered-industrial-yeast-saccharomyces-cerevisiae
#9
Quanzhou Feng, Z Lewis Liu, Scott A Weber, Shizhong Li
Haploid laboratory strains of Saccharomyces cerevisiae are commonly used for genetic engineering to enable their xylose utilization but little is known about the industrial yeast which is often recognized as diploid and as well as haploid and tetraploid. Here we report three unique signature pathway expression patterns and gene interactions in the centre metabolic pathways that signify xylose utilization of genetically engineered industrial yeast S. cerevisiae NRRL Y-50463, a diploid yeast. Quantitative expression analysis revealed outstanding high levels of constitutive expression of YXI, a synthesized yeast codon-optimized xylose isomerase gene integrated into chromosome XV of strain Y-50463...
2018: PloS One
https://www.readbyqxmd.com/read/29545204/baculovirus-driven-protein-expression-in-insect-cells-a-benchmarking-study
#10
Peggy Stolt-Bergner, Christian Benda, Tim Bergbrede, Hüseyin Besir, Patrick H N Celie, Cindy Chang, David Drechsel, Ariane Fischer, Arie Geerlof, Barbara Giabbai, Joop van den Heuvel, Georg Huber, Wolfgang Knecht, Anita Lehner, Regis Lemaitre, Kristina Nordén, Gwynn Pardee, Ines Racke, Kim Remans, Astrid Sander, Judith Scholz, Magda Stadnik, Paola Storici, Daniel Weinbruch, Isabel Zaror, Linda H L Lua, Sabine Suppmann
Baculovirus-insect cell expression system has become one of the most widely used eukaryotic expression systems for heterologous protein production in many laboratories. The availability of robust insect cell lines, serum-free media, a range of vectors and commercially-packaged kits have supported the demand for maximizing the exploitation of the baculovirus-insect cell expression system. Naturally, this resulted in varied strategies adopted by different laboratories to optimize protein production. Most laboratories have preference in using either the E...
March 12, 2018: Journal of Structural Biology
https://www.readbyqxmd.com/read/29540599/development-of-a-broadly-accessible-venezuelan-equine-encephalitis-virus-replicon-particle-vaccine-platform
#11
Sudhakar Agnihothram, Vineet D Menachery, Boyd L Yount, Lisa C Lindesmith, Trevor Scobey, Alan Whitmore, Alexandra Schäfer, Mark T Heise, Ralph S Baric
Zoonotic viruses circulate as swarms in animal reservoirs and can emerge into human populations, causing epidemics that adversely affect public health. Portable, safe, and effective vaccine platforms are needed in the context of these outbreak and emergence situations. In this work, we report the generation and characterization of an alphavirus replicon vaccine platform based on a non-select agent, attenuated Venezuelan equine encephalitis (VEE) virus vaccine, strain 3526 (VRP 3526). Using both noroviruses and coronaviruses as model systems, we demonstrate the utility of the VRP 3526 platform in the generation of recombinant proteins, production of virus-like particles, and in vivo efficacy as a vaccine against emergent viruses...
June 1, 2018: Journal of Virology
https://www.readbyqxmd.com/read/29535210/rapid-production-of-pure-recombinant-actin-isoforms-in-pichia-pastoris
#12
Tomoyuki Hatano, Salvatore Alioto, Emanuele Roscioli, Saravanan Palani, Scott T Clarke, Anton Kamnev, Juan Ramon Hernandez-Fernaud, Lavanya Sivashanmugam, Bernardo Chapa-Y-Lazo, Alexandra M E Jones, Robert C Robinson, Karuna Sampath, Masanori Mishima, Andrew D McAinsh, Bruce L Goode, Mohan K Balasubramanian
Actins are major eukaryotic cytoskeletal proteins, and they are involved in many important cell functions, including cell division, cell polarity, wound healing and muscle contraction. Despite obvious drawbacks, muscle actin, which is easily purified, is used extensively for biochemical studies of the non-muscle actin cytoskeleton. Here, we report a rapid and cost-effective method to purify heterologous actins expressed in the yeast Pichia pastoris Actin is expressed as a fusion with the actin-binding protein thymosin β4 and purified by means of an affinity tag introduced in the fusion...
April 23, 2018: Journal of Cell Science
https://www.readbyqxmd.com/read/29455388/heterologous-expression-of-rtshyal-1-the-first-recombinant-hyaluronidase-of-scorpion-venom-produced-in-pichia-pastoris-system
#13
Fernanda Gobbi Amorim, Johara Boldrini-França, Karla de Castro Figueiredo Bordon, Iara Aimê Cardoso, Edwin De Pauw, Loïc Quinton, Simone Kashima, Eliane Candiani Arantes
In general, hyaluronidases have a broad potential application on medicine and esthetics fields. Hyaluronidases from animal venoms cleave hyaluronan present in the extracellular matrix, acting as spreading factors of toxins into the tissues of the victim. However, the in-depth characterization of hyaluronidase from animal venoms has been neglected due to its instability and low concentration in the venom, which hamper its isolation. Thus, heterologous expression of hyaluronidase acts as a biotechnological tool in the obtainment of enough amounts of the enzyme for structural and functional studies...
April 2018: Applied Microbiology and Biotechnology
https://www.readbyqxmd.com/read/29438403/release-of-transcriptional-repression-through-the-hcr-promoter-region-confers-uniform-expression-of-hwp1-on-surfaces-of-candida-albicans-germ-tubes
#14
Samin Kim, Quoc Bao Nguyen, Michael J Wolyniak, Gregory Frechette, Christian R Lehman, Brandon K Fox, Paula Sundstrom
The mechanisms that fungi use to co-regulate subsets of genes specifically associated with morphogenic states represent a basic unsolved problem in fungal biology. Candida albicans is an important model of fungal differentiation both for rapid interconversion between yeast and hyphal growth forms and for white/opaque switching mechanisms. The Sundstrom lab is interested in mechanisms regulating hypha-specific expression of adhesin genes that are critical for C. albicans hyphal growth phenotypes and pathogenicity...
2018: PloS One
https://www.readbyqxmd.com/read/29434326/culture-independent-discovery-of-the-malacidins-as-calcium-dependent-antibiotics-with-activity-against-multidrug-resistant-gram-positive-pathogens
#15
Bradley M Hover, Seong-Hwan Kim, Micah Katz, Zachary Charlop-Powers, Jeremy G Owen, Melinda A Ternei, Jeffrey Maniko, Andreia B Estrela, Henrik Molina, Steven Park, David S Perlin, Sean F Brady
Despite the wide availability of antibiotics, infectious diseases remain a leading cause of death worldwide 1 . In the absence of new therapies, mortality rates due to untreatable infections are predicted to rise more than tenfold by 2050. Natural products (NPs) made by cultured bacteria have been a major source of clinically useful antibiotics. In spite of decades of productivity, the use of bacteria in the search for new antibiotics was largely abandoned due to high rediscovery rates2,3 . As only a fraction of bacterial diversity is regularly cultivated in the laboratory and just a fraction of the chemistries encoded by cultured bacteria are detected in fermentation experiments, most bacterial NPs remain hidden in the global microbiome...
April 2018: Nature Microbiology
https://www.readbyqxmd.com/read/29380150/dynamic-metabolic-control-towards-precision-engineering-of-metabolism
#16
Di Liu, Ahmad A Mannan, Yichao Han, Diego A Oyarzún, Fuzhong Zhang
Advances in metabolic engineering have led to the synthesis of a wide variety of valuable chemicals in microorganisms. The key to commercializing these processes is the improvement of titer, productivity, yield, and robustness. Traditional approaches to enhancing production use the "push-pull-block" strategy that modulates enzyme expression under static control. However, strains are often optimized for specific laboratory set-up and are sensitive to environmental fluctuations. Exposure to sub-optimal growth conditions during large-scale fermentation often reduces their production capacity...
January 29, 2018: Journal of Industrial Microbiology & Biotechnology
https://www.readbyqxmd.com/read/29371619/immunization-with-transgenic-rodent-malaria-parasites-expressing-pfs25-induces-potent-transmission-blocking-activity
#17
K A Sala, F Angrisano, D F Da, I J Taylor, T S Churcher, A M Blagborough
An anti-malarial transmission blocking vaccine (TBV) would be an important tool for disease control or elimination, though current candidates have failed to induce high efficacy in clinical studies. The ookinete surface protein P25 is a primary target for TBV development, but heterologous expression of P25 with appropriate conformation is problematic and a pre-requisite for achieving functional titers. A potential alternative to recombinant/sub-unit vaccine is immunization with a non-pathogenic, whole-parasite vaccine...
January 25, 2018: Scientific Reports
https://www.readbyqxmd.com/read/29286407/generic-protocol-for-optimization-of-heterologous-protein-production-using-automated-microbioreactor-technology
#18
Johannes Hemmerich, Lars Freier, Wolfgang Wiechert, Eric von Lieres, Marco Oldiges
A core business in industrial biotechnology using microbial production cell factories is the iterative process of strain engineering and optimization of bioprocess conditions. One important aspect is the improvement of cultivation medium to provide an optimal environment for microbial formation of the product of interest. It is well accepted that the media composition can dramatically influence overall bioprocess performance. Nutrition medium optimization is known to improve recombinant protein production with microbial systems and thus, this is a rewarding step in bioprocess development...
December 15, 2017: Journal of Visualized Experiments: JoVE
https://www.readbyqxmd.com/read/29198849/oxalate-degrading-enzyme-recombined-lactic-acid-bacteria-strains-reduce-hyperoxaluria
#19
Chenming Zhao, Huan Yang, Xiaojing Zhu, Yang Li, Ning Wang, Shanfu Han, Hua Xu, Zhiqiang Chen, Zhangqun Ye
OBJECTIVE: To develop recombinant lactic acid bacteria (LAB) strains that express oxalate-degrading enzymes through biotechnology-based approach for the treatment of hyperoxaluria by oral administration. MATERIAL AND METHODS: The coding gene of oxalate decarboxylase (ODC) and oxalate oxidase (OxO) was transformed into Lactococcus lactis MG1363. The oxalate degradation ability in vitro was evaluated in media with high concentration of oxalate. Hyperoxaluria rat models through high oxalate diet were given recombinant LAB through oral administration...
March 2018: Urology
https://www.readbyqxmd.com/read/29090064/-lactococcus-lactis-a-new-strategy-for-vaccination
#20
REVIEW
Maryam Azizpour, Seyyed Davood Hosseini, Parvaneh Jafari, Neda Akbary
Needle free vaccines have a several advantages and very attractive way for vaccination. In a body, mucosal surfaces provide a universal entry portal for all the known and emerging infectious pathogenic microbes. Therefore, it seems, vaccination strategies need to be reorganized for vaccines that are hindering the entry capability of pathogenic microbes through mucosal surfaces. Lactic acid Bacteria (LAB) are widely used in the food industry and at the present, used as delivery vehicles for biological investigations...
October 2017: Avicenna Journal of Medical Biotechnology
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