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Heterologous expression lab

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https://www.readbyqxmd.com/read/27906033/the-role-of-the-backbone-torsion-in-protein-folding
#1
EDITORIAL
Irina Sorokina, Arcady Mushegian
BACKGROUND: The set of forces and sequence of events that govern the transition from an unfolded polypeptide chain to a functional protein with correct spatial structure remain incompletely known, despite the importance of the problem and decades of theory development, computer simulations, and laboratory experiments. Information about the correctly folded state of most proteins is likely to be present in their sequences, and yet many proteins fail to attain native structure after overexpression in a non-native environment or upon experimental denaturation and refolding...
December 1, 2016: Biology Direct
https://www.readbyqxmd.com/read/27888410/genome-sequence-prevalence-and-quantification-of-the-first-iflavirus-identified-in-a-phytoplasma-insect-vector
#2
Simona Abbà, Luciana Galetto, Marta Vallino, Marika Rossi, Massimo Turina, Anne Sicard, Cristina Marzachì
The leafhopper Euscelidius variegatus is a natural vector of chrysanthemum yellows phytoplasma (CY) and an efficient vector of flavescence dorée phytoplasma (FD) under laboratory conditions. During a transcriptome sequencing (RNA-seq) project aimed at investigating the interactions between the insect and the two phytoplasmas, a 10,616-nucleotide-long contig with high sequence similarity to known picorna-like viruses was identified among the assembled insect transcripts. The discovery came totally unexpected, because insects from the laboratory colony did not show any evident symptom that could be related to the presence of a virus...
November 25, 2016: Archives of Virology
https://www.readbyqxmd.com/read/27830495/cloning-a-novel-endo-1-4-%C3%AE-d-glucanase-gene-from-trichoderma-virens-and-heterologous-expression-in-e-coli
#3
Rong Zeng, Qiao Hu, Xiao-Yan Yin, Hao Huang, Jia-Bao Yan, Zhi-Wei Gong, Zhong-Hua Yang
Endo-1,4-β-D-glucanase (EG), as a key constituent of cellulase taking the responsibility of cutting β-1,4 glycosidic bonds, plays the essential role in the process of degrading cellulose by cellulase. Cloning and expressing the EG gene is important to the cellulase research and application. In this work, a novel EG gene was cloned from Trichoderma virens ZY-01, which was a cellulase secreting microbe isolated by our laboratory. The DNA sequence showed that the length of the cloned EG is 1069 bp, which had 95...
December 2016: AMB Express
https://www.readbyqxmd.com/read/27816999/heterologous-expression-of-the-crassostrea-gigas-pacific-oyster-alternative-oxidase-in-the-yeast-saccharomyces-cerevisiae
#4
Aaron Robertson, Kyle Schaltz, Karina Neimanis, James F Staples, Allison E McDonald
Alternative oxidase (AOX) is a terminal oxidase within the inner mitochondrial membrane (IMM) present in many organisms where it functions in the electron transport system (ETS). AOX directly accepts electrons from ubiquinol and is therefore capable of bypassing ETS Complexes III and IV. The human genome does not contain a gene coding for AOX, so AOX expression has been suggested as a gene therapy for a range of human mitochondrial diseases caused by genetic mutations that render Complex III and/or IV dysfunctional...
November 5, 2016: Journal of Bioenergetics and Biomembranes
https://www.readbyqxmd.com/read/27745812/meningococcal-serogroup-b-specific-responses-after-vaccination-with-bivalent-rlp2086-4-year-follow-up-of-a-randomised-single-blind-placebo-controlled-phase-2-trial
#5
Helen S Marshall, Peter C Richmond, Johannes Beeslaar, Qin Jiang, Kathrin U Jansen, Maria Garcés-Sánchez, Federico Martinón-Torres, Leszek Szenborn, Jacek Wysocki, Joseph Eiden, Shannon L Harris, Thomas R Jones, Su-San Lee, John L Perez
BACKGROUND: Bivalent rLP2086 is a recombinant factor H binding protein-based vaccine approved in the USA for prevention of meningococcal serogroup B disease in 10-25-year-olds. We aimed to assess the persistence of bactericidal antibodies up to 4 years after a three-dose schedule of bivalent rLP2086. METHODS: We did this randomised, single-blind, placebo-controlled, phase 2 trial at 25 sites in Australia, Poland, and Spain. In stage 1 of the study (February, 2009-May, 2010), healthy adolescents (aged 11-18 years) were randomly assigned, via an interactive voice and web-response system with computer-generated sequential random numbers, to receive either ascending doses of vaccine (60 μg, 120 μg, and 200 μg) or placebo at months 0, 2, and 6...
October 10, 2016: Lancet Infectious Diseases
https://www.readbyqxmd.com/read/27692771/identification-of-plasmodium-falciparum-reticulocyte-binding-protein-homologue-5-interacting-protein-pfripr-as-a-highly-conserved-blood-stage-malaria-vaccine-candidate
#6
Edward H Ntege, Nobuko Arisue, Daisuke Ito, Tomoyuki Hasegawa, Nirianne M Q Palacpac, Thomas G Egwang, Toshihiro Horii, Eizo Takashima, Takafumi Tsuboi
Genetic variability in Plasmodium falciparum malaria parasites hampers current malaria vaccine development efforts. Here, we hypothesize that to address the impact of genetic variability on vaccine efficacy in clinical trials, conserved antigen targets should be selected to achieve robust host immunity across multiple falciparum strains. Therefore, suitable vaccine antigens should be assessed for levels of polymorphism and genetic diversity. Using a total of one hundred and two clinical isolates from a region of high malaria transmission in Uganda, we analyzed extent of polymorphism and genetic diversity in four recently reported novel blood-stage malaria vaccine candidate proteins: Rh5 interacting protein (PfRipr), GPI anchored micronemal antigen (PfGAMA), rhoptry-associated leucine zipper-like protein 1 (PfRALP1) and Duffy binding-like merozoite surface protein 1 (PfMSPDBL1)...
September 28, 2016: Vaccine
https://www.readbyqxmd.com/read/27609229/secretion-of-m2e-hbc-fusion-protein-by-lactobacillus-casei-using-cwh-signal-peptide
#7
Sirintra Themsakul, Namfon Suebwongsa, Baltasar Mayo, Marutpong Panya, Viraphong Lulitanond
The ability to serve as a delivery vehicle for various interesting biomolecules makes lactic acid bacteria (LAB) very useful in several applications. In the medical field, recombinant LAB expressing pathogenic antigens at different cellular locations have been used to elicit both mucosal and systemic immune responses. Expression-secretion vectors (ESVs) with a signal peptide (SP) are pivotal for protein expression and secretion. In this study, the genome sequence of Lactobacillus casei ATCC334 was explored for new SPs using bioinformatics tools...
October 2016: FEMS Microbiology Letters
https://www.readbyqxmd.com/read/27538995/roseolovirus-associated-encephalitis-in-immunocompetent-and-immunocompromised-individuals
#8
Joseph Ongrádi, Dharam V Ablashi, Tetsushi Yoshikawa, Balázs Stercz, Masao Ogata
The roseoloviruses, human herpesvirus (HHV)-6A, HHV-6B, and HHV-7, can cause severe encephalitis or encephalopathy. In immunocompetent children, primary HHV-6B infection is occasionally accompanied by diverse clinical forms of encephalitis. Roseolovirus coinfections with heterologous viruses and delayed primary HHV-7 infection in immunocompetent adults result in very severe neurological and generalized symptoms. Recovery from neurological sequelae is slow and sometimes incomplete. In immunocompromised patients with underlying hematological malignancies and transplantation, frequent single or simultaneous reactivation of roseoloviruses elicit severe, lethal organ dysfunctions, including damages in the limbic system, brain stem, and hippocampus...
August 18, 2016: Journal of Neurovirology
https://www.readbyqxmd.com/read/27486448/transfer-of-a-catabolic-pathway-for-chloromethane-in-methylobacterium-strains-highlights-different-limitations-for-growth-with-chloromethane-or-with-dichloromethane
#9
Joshua K Michener, Stéphane Vuilleumier, Françoise Bringel, Christopher J Marx
Chloromethane (CM) is an ozone-depleting gas, produced predominantly from natural sources, that provides an important carbon source for microbes capable of consuming it. CM catabolism has been difficult to study owing to the challenging genetics of its native microbial hosts. Since the pathways for CM catabolism show evidence of horizontal gene transfer, we reproduced this transfer process in the laboratory to generate new CM-catabolizing strains in tractable hosts. We demonstrate that six putative accessory genes improve CM catabolism, though heterologous expression of only one of the six is strictly necessary for growth on CM...
2016: Frontiers in Microbiology
https://www.readbyqxmd.com/read/27382786/-progress-in-heterologous-expression-of-fungal-natural-products-a-review
#10
REVIEW
Zihui Ma, Wei Li, Wenbing Yin
Fungal products are important resources for natural drugs. Availabilities of many fungal genome sequences indicate that fungi have the huge potential to produce secondary metabolites. However, many valuable fungal natural products are hard to obtain under regular lab conditions because fungi are slow to grow, difficult to genetically modify, or have low yield of desired metabolites. Therefore, developing a heterologous expression system has been an efficient approach to discover novel natural product and elucidate the complicated biosynthetic pathways...
March 4, 2016: Wei Sheng Wu Xue Bao, Acta Microbiologica Sinica
https://www.readbyqxmd.com/read/27381522/insights-into-microbial-cryptic-gene-activation-and-strain-improvement-principle-application-and-technical-aspects
#11
Kozo Ochi
As bacteria and fungi have been found to contain genes encoding enzymes that synthesize a plethora of potential secondary metabolites, interest has grown in the activation of these cryptic pathways. Homologous and heterologous expression of these cryptic secondary metabolite-biosynthetic genes, often silent under ordinary laboratory fermentation conditions, may lead to the discovery of novel secondary metabolites. This review addresses current progress in the activation of these pathways, describing methods for activating silent genes...
July 6, 2016: Journal of Antibiotics
https://www.readbyqxmd.com/read/27373940/heterologous-expression-and-purification-of-plantaricin-nc8-a-two-peptide-bacteriocin-against-salmonella-spp-from-lactobacillus-plantarum-zj316
#12
Han Jiang, Ping Li, Qing Gu
Bacteriocin, which is produced by lactic acid bacteria (LAB), has the potential to act as natural preservatives in the food industry. To develop strategies to overproduce such peptides, plantaricin NC8, a class IIb LAB bacteriocin that consists of two peptides, PLNC8α and PLNC8β, was successfully heterologously expressed in Escherichia coli BL21 (DE3). PLNC8α and PLNC8β peptides were expressed as His6-tag fusion proteins and were separated by Ni(2+) chelating affinity chromatography. To get the PLNC8α and PLNC8β peptides without extra amino acids in the N-terminus, the fusion proteins were cleaved by enterokinase and further purified using the Ni-NTA Sefinose™ Resin Kit...
November 2016: Protein Expression and Purification
https://www.readbyqxmd.com/read/27317421/the-fed-batch-principle-for-the-molecular-biology-lab-controlled-nutrient-diets-in-ready-made-media-improve-production-of-recombinant-proteins-in-escherichia-coli
#13
REVIEW
Mirja Krause, Antje Neubauer, Peter Neubauer
While the nutrient limited fed-batch technology is the standard of the cultivation of microorganisms and production of heterologous proteins in industry, despite its advantages in view of metabolic control and high cell density growth, shaken batch cultures are still the standard for protein production and expression screening in molecular biology and biochemistry laboratories. This is due to the difficulty and expenses to apply a controlled continuous glucose feed to shaken cultures. New ready-made growth media, e...
2016: Microbial Cell Factories
https://www.readbyqxmd.com/read/27303712/radicicol-mediated-inhibition-of-topoisomerase-vib-via-activity-of-the-human-malaria-parasite-plasmodium-falciparum
#14
Sureshkumar Chalapareddy, Swati Chakrabarty, Mrinal Kanti Bhattacharyya, Sunanda Bhattacharyya
Plasmodium falciparum topoisomerase VIB (TopoVIB)-TopoVIA (TopoVIB-VIA) complex can be potentially exploited as a drug target against malaria due to its absence from the human genome. Previous work in our laboratory has suggested that P. falciparum TopoVIB (PfTopoVIB) might be a target of radicicol since treatment of parasite cultures with this antibiotic is associated with upregulation of Plasmodium TopoVIB at the transcript level as well as at the protein level. Further studies demonstrated that radicicol treatment impaired mitochondrial replication of human malaria parasite P...
January 2016: MSphere
https://www.readbyqxmd.com/read/27190148/a-new-lactobacilli-in-vivo-expression-system-for-the-production-and-delivery-of-heterologous-proteins-at-mucosal-surfaces
#15
Thibault Allain, Nahla M Mansour, May M A Bahr, Rebeca Martin, Isabelle Florent, Philippe Langella, Luis G Bermúdez-Humarán
Food-grade lactic acid bacteria, such as lactobacilli, represent good candidates for the development of mucosal vectors. Indeed, they are generally recognized as safe microorganisms and some strains display beneficial effects (probiotics). In this study, we described a new lactobacilli in vivo expression (LIVE) system for the production and delivery of therapeutic molecules at mucosal surfaces. The versatility and functionality of this system was successfully validated in several lactobacilli species; furthermore, we assessed in vivo LIVE system in two different mouse models of human pathologies: (i) a model of therapy against intestinal inflammation (inflammatory bowel diseases) and (ii) a model of vaccination against dental caries...
July 2016: FEMS Microbiology Letters
https://www.readbyqxmd.com/read/27172193/a-comprehensive-toolbox-for-genome-editing-in-cultured-drosophila-melanogaster-cells
#16
Stefan Kunzelmann, Romy Böttcher, Ines Schmidts, Klaus Förstemann
Custom genome editing has become an essential element of molecular biology. In particular, the generation of fusion constructs with epitope tags or fluorescent proteins at the genomic locus facilitates the analysis of protein expression, localization, and interaction partners at physiologic levels. Following up on our initial publication, we now describe a considerably simplified, more efficient, and readily scalable experimental workflow for PCR-based genome editing in cultured Drosophila melanogaster cells...
2016: G3: Genes—Genomes—Genetics
https://www.readbyqxmd.com/read/27165322/protein-complex-production-in-alternative-prokaryotic-hosts
#17
REVIEW
Sara Gómez, Miguel López-Estepa, Francisco J Fernández, M Cristina Vega
Research for multiprotein expression in nonconventional bacterial and archaeal expression systems aims to exploit particular properties of "alternative" prokaryotic hosts that might make them more efficient than E. coli for particular applications, especially in those areas where more conventional bacterial hosts traditionally do not perform well. Currently, a wide range of products with clinical or industrial application have to be isolated from their native source, often microorganisms whose growth present numerous problems owing to very slow growth phenotypes or because they are unculturable under laboratory conditions...
2016: Advances in Experimental Medicine and Biology
https://www.readbyqxmd.com/read/27165318/complex-reconstitution-and-characterization-by-combining-co-expression-techniques-in-escherichia-coli-with-high-throughput
#18
REVIEW
Renaud Vincentelli, Christophe Romier
Single protein expression technologies have strongly benefited from the Structural Genomics initiatives that have introduced parallelization at the laboratory level. Specifically, the developments made in the wake of these initiatives have revitalized the use of Escherichia coli as major host for heterologous protein expression. In parallel to these improvements for single expression, technologies for complex reconstitution by co-expression in E. coli have been developed. Assessments of these co-expression technologies have highlighted the need for combinatorial experiments requiring automated protocols...
2016: Advances in Experimental Medicine and Biology
https://www.readbyqxmd.com/read/27155483/functional-characterization-of-an-%C3%AE-esterase-gene-involving-malathion-detoxification-in-bactrocera-dorsalis-hendel
#19
Luo-Luo Wang, Xue-Ping Lu, Li-Wei Meng, Yong Huang, Dong Wei, Hong-Bo Jiang, Guy Smagghe, Jin-Jun Wang
Extensive use of insecticides in many orchards has prompted resistance development in the oriental fruit fly, Bactrocera dorsalis (Hendel). In this study, a laboratory selected strain of B. dorsalis (MR) with a 21-fold higher resistance to malathion was used to examine the resistance mechanisms to this organophosphate insecticide. Carboxylesterase (CarE) was found to be involved in malathion resistance in B. dorsalis from the synergism bioassay by CarE-specific inhibitor triphenylphosphate (TPP). Molecular studies further identified a previously uncharacterized α-esterase gene, BdCarE2, that may function in the development of malathion resistance in B...
June 2016: Pesticide Biochemistry and Physiology
https://www.readbyqxmd.com/read/27084006/gene-amplification-on-demand-accelerates-cellobiose-utilization-in-engineered-saccharomyces-cerevisiae
#20
Eun Joong Oh, Jeffrey M Skerker, Soo Rin Kim, Na Wei, Timothy L Turner, Matthew J Maurer, Adam P Arkin, Yong-Su Jin
UNLABELLED: Efficient microbial utilization of cellulosic sugars is essential for the economic production of biofuels and chemicals. Although the yeast Saccharomyces cerevisiae is a robust microbial platform widely used in ethanol plants using sugar cane and corn starch in large-scale operations, glucose repression is one of the significant barriers to the efficient fermentation of cellulosic sugar mixtures. A recent study demonstrated that intracellular utilization of cellobiose by engineered yeast expressing a cellobiose transporter (encoded by cdt-1) and an intracellular β-glucosidase (encoded by gh1-1) can alleviate glucose repression, resulting in the simultaneous cofermentation of cellobiose and nonglucose sugars...
June 15, 2016: Applied and Environmental Microbiology
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