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Recombinant expression

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https://www.readbyqxmd.com/read/28735081/anti-inflammatory-activity-of-cecropin-a2-from-musca-domestica
#1
Rui-Yang Wei, Jie Bai, Meng-Fei Zhao, Bin Xu, Wen-Jia Li, Feng-Xian Wei, Yan-Yan Xi, Shao-Yu Li
This study aimed to investigate the anti-inflammatory activity of Musca domestica cecropin-A2 (Mdc-A2) toward Staphylococcus aureus (S. aureus) to learn more about their immunological functions. RAW264.7 cells were transfected with recombinant lentiviruses introduce pLEX-Mdc-A2into the RAW264.7 cell line (RAW-Mdc-A2). The RAW264.7 cell line with empty pLEX (RAW-pLEX) was produced in the same manner as a negative control. Real-time quantitative reverse transcription PCR (RT-PCR) was performed to analyze the mRNA expression of TNF-a, IL-1β, NFκB-1 and NFκB-2 in S...
July 19, 2017: Microbial Pathogenesis
https://www.readbyqxmd.com/read/28735080/recombinant-truncated-e-protein-as-a-new-vaccine-candidate-against-nontypeable-h-influenzae-its-expression-and-immunogenic-evaluation
#2
Ava Behrouzi, Saeid Bouzari, Farzam Vaziri, Abolfazl Fateh, Parviz Afrough, Atefeh Davoudi Vijeh Motlagh, Seyed Davar Siadat
Protein E (PE) is a conserved entity observed in both nontypeable Haemophilus influenzae (NTHi) and encapsulated H. influenzae. This is a small surface lipoprotein, consisting of only 160 amino acids, involved in the adhesion of H. influenzae to various types of epithelial cells. A 384-bp-long fragment from NTHi PE was cloned into the prokaryotic expression vector pBAD-gIIIA. The recombinant protein was expressed with arabinose and then purified by affinity purification on an Ni-NTA agarose matrix. BALB/c mice were immunized by subcutaneous injection with purified recombinant truncated PE mixed with an alum adjuvant...
July 19, 2017: Microbial Pathogenesis
https://www.readbyqxmd.com/read/28734978/insights-into-the-functional-properties-of-the-marneral-oxidase-cyp71a16-from-arabidopsis-thaliana
#3
Sarah Kranz-Finger, Osama Mahmoud, Esther Ricklefs, Nikolas Ditz, Patrick J Bakkes, Vlada B Urlacher
The Arabidopsis thaliana gene encoding CYP71A16 is part of the gene cluster for the biosynthesis and modification of the triterpenoid marneral. Previous investigations of A. thaliana have revealed that CYP71A16 oxidizes marneral, while it also can accept marnerol as substrate. The aim of the present study was to investigate functional properties of CYP71A16 in vitro. For this purpose, heterologous expression of a N-terminally modified version of CYP71A16 was established in Escherichia coli, which yielded up to 50mg·L(-1) recombinant enzyme...
July 19, 2017: Biochimica et Biophysica Acta
https://www.readbyqxmd.com/read/28734839/recombinant-expression-of-ixolaris-a-kunitz-type-inhibitor-from-the-tick-salivary-gland-for-nmr-studies
#4
V S De Paula, F H S Silva, I M B Francischetti, R Q Monteiro, A P Valente
Ixolaris is an anticoagulant protein identified in the tick saliva of Ixodes scapularis. Ixolaris contains 2 Kunitz like domains and binds to Factor Xa or Factor X as a scaffold for inhibition of the Tissue Factor (TF)/Factor VIIa (FVIIa). In contrast to tissue factor pathway inhibitor (TFPI), however, Ixolaris does not bind to the active site cleft of FXa. Instead, complex formation is mediated by the FXa heparin-binding exosite. Due to its potent and long-lasting antithrombotic activity, Ixolaris is a promising agent for anticoagulant therapy...
July 19, 2017: Protein Expression and Purification
https://www.readbyqxmd.com/read/28734838/expression-purification-and-characterization-of-sll1981-protein-from-cyanobacterium-synechocystis-sp-pcc6803
#5
Xiaoqin Wang, Guofeng Lei, Xiaoyu Wu, Fei Wang, Chongde Lai, Zhimin Li
The sll1981 protein from cyanobacterium Synechocystis sp. PCC6803 had been reported to exhibit acetolactate synthase (ALS) and L-myo-inositol-1-phosphate synthase (MIPS) activities previously. Based on amino acids sequences alignment, sll1981 protein was postulated to function as α-ketoglutarate decarboxylase (α-KGD), which played important role in completing cyanobacterial tricarboxylic acid (TCA) cycle. However the detailed enzymatic kinetics of sll1981 as ALS, MIPS and α-KGD were not determined yet. In this study, the recombinant sll1981 protein was purified from supernatant of E...
July 19, 2017: Protein Expression and Purification
https://www.readbyqxmd.com/read/28734180/bottom-up-assembly-of-salivary-gland-microtissues-for-assessing-myoepithelial-cell-function
#6
Tugba Ozdemir, Padma Pradeepa Srinivasan, Daniel R Zakheim, Daniel A Harrington, Robert L Witt, Mary C Farach-Carson, Xinqiao Jia, Swati Pradhan-Bhatt
Myoepithelial cells are flat, stellate cells present in exocrine tissues including the salivary glands. While myoepithelial cells have been studied extensively in mammary and lacrimal gland tissues, less is known of the function of myoepithelial cells derived from human salivary glands. Several groups have isolated tumorigenic myoepithelial cells from cancer specimens, however, only one report has demonstrated isolation of normal human salivary myoepithelial cells needed for use in salivary gland tissue engineering applications...
July 14, 2017: Biomaterials
https://www.readbyqxmd.com/read/28734106/insecticide-resistance-in-trialeurodes-vaporariorum-populations-and-novel-diagnostics-for-kdr-mutations
#7
Despoina Evr Kapantaidaki, Eldem Sadikoglou, Dimitra Tsakireli, Vasileios Kampanis, Marianna Stavrakaki, Corinna Schorn, Aris Ilias, Maria Riga, George Tsiamis, Ralf Nauen, George Skavdis, John Vontas, Anastasia Tsagkarakou
BACKGROUND: Neonicotinoids, pyrethroids and ketoenols, are currently used for the control of Trialeurodes vaporariorum (Hemiptera: Aleyrodidae). In this study, insecticide resistance status and mechanisms were investigated by classical bioassays and molecular techniques. RESULTS: Dose response bioassays were performed on 19 Greek populations, among the 35 different whitefly populations used for the whole analysis. Resistance factors scaled up to 207-, 4657- and 59- fold, for imidacloprid, bifenthrin and spiromesifen, respectively...
July 22, 2017: Pest Management Science
https://www.readbyqxmd.com/read/28734049/development-of-aptamers-against-unpurified-proteins
#8
Shinichi Goto, Kaori Tsukakoshi, Kazunori Ikebukuro
SELEX (Systematic Evolution of Ligands by EXponential enrichment) has been widely used for the generation of aptamers against target proteins. However, its requirement for pure target proteins remains a major problem in aptamer selection, as procedures for protein purification from crude bio-samples are not only complicated but also time and labour consuming. This is because native proteins can be found in a large number of diverse forms because of posttranslational modifications and their complicated molecular conformations...
July 22, 2017: Biotechnology and Bioengineering
https://www.readbyqxmd.com/read/28734047/preselection-of-recombinant-gene-integration-sites-enabling-high-transcription-rates-in-cho-cells-using-alternate-start-codons-and-recombinase-mediated-cassette-exchange
#9
Martina Baumann, Elisabeth Gludovacz, Natalie Sealover, Scott Bahr, Henry George, Nan Lin, Kevin Kayser, Nicole Borth
Site-specific Recombinase Mediated Cassette Exchange (RMCE) enables the transfer of the gene of interest (GOI) into pre-selected genomic locations with defined expression properties. For the generation of recombinant production cell lines this has the advantage that screening for high transcription rates at the genome integration site would be required only once, with the possibility to reuse the selected site for new products. Here we describe a strategy that aims at the selection of transcriptionally active genome integration sites in Chinese Hamster Ovary (CHO) cells by using alternate start codons in the surface reporter protein CD4, in combination with FACS sorting for high expressers...
July 22, 2017: Biotechnology and Bioengineering
https://www.readbyqxmd.com/read/28733938/molecular-cloning-heterologous-expression-and-functional-characterization-of-a-cellulolytic-enzyme-cel-prii-from-buffalo-rumen-metagenome
#10
Ravi K Shah, Amrutlal K Patel, Deepti M Davla, Ishan K Parikh, Ramalingam B Subramanian, Kamlesh C Patel, Subhash J Jakhesara, Chaitanya G Joshi
A cellulase encoding gene, Cel PRII, was identified from Mehsani buffalo rumen metagenome, and cloned and expressed in Escherichia coli BL21(DE3)pLysS. The 1170 bp full length gene encodes a 389 residue polypeptide (Cel PRII) containing a catalytic domain belonging to glycosyl hydrolase (GH) 5 family. The fusion protein consisting of the Cel PRII, thioredoxin tag and 6x Histidine tag with predicted molecular weight of 63 kDa when recovered from inclusion bodies under denaturing conditions, exhibited cellulolytic activity against carboxymethyl cellulose (CMC)...
August 2017: 3 Biotech
https://www.readbyqxmd.com/read/28733403/transformation-of-schizosaccharomyces-japonicus
#11
Keita Aoki, Hironori Niki
This protocol describes the use of electroporation to transform Schizosaccharomyces japonicus with plasmids or linear DNA. Plasmids are helpful for the complementation testing of mutations and for the expression of specific genes. Linear DNA fragments integrated into chromosomal DNA by homologous recombination are useful for gene deletion or to fuse a gene with a tag sequence (e.g., encoding a fluorescent protein). To introduce DNA into S. japonicus, electroporation methods are recommended because S. japonicus is sensitive to lithium acetate (LiOAc)...
July 21, 2017: Cold Spring Harbor Protocols
https://www.readbyqxmd.com/read/28732288/lncrna-hotair-alleviates-rheumatoid-arthritis-by-targeting-mir-138-and-inactivating-nf-%C3%AE%C2%BAb-pathway
#12
Hong-Ju Zhang, Qiao-Feng Wei, Shu-Jun Wang, Hong-Jie Zhang, Xiu-Ying Zhang, Qin Geng, Yan-Hui Cui, Xiu-Hua Wang
Rheumatoid arthritis (RA) is a chronic and autoimmune-mediated inflammatory disease. We aimed to investigate the regulation of lncRNA HOTAIR in LPS-treated chondrocytes and RA mouse. Our results showed that HOTAIR expression was significantly reduced in LPS-treated chondrocytes. The HOTAIR was then over-expressed in chondrocytes by transfecting recombinant lentivirus carrying sequences encoding HOTAIR. The LPS-induced reduction of cell proliferation rate and production of two inflammatory factors interleukin (IL)-17, IL-23 were markedly inhibited...
July 18, 2017: International Immunopharmacology
https://www.readbyqxmd.com/read/28732140/fibroblast-growth-factor-2-fgf2-is-present-in-human-spermatozoa-and-is-related-with-sperm-motility-the-use-of-recombinant-fgf2-to-improve-motile-sperm-recovery
#13
D J Garbarino Azúa, L Saucedo, S Giordana, M L Magri, M G Buffone, F Neuspiller, M H Vazquez-Levin, C I Marín-Briggiler
Fibroblast growth factors (FGFs) and their receptors (FGFRs) regulate several functions of somatic cells. In a previous work, we reported FGFR expression in human spermatozoa and their involvement in motility. This study aimed to evaluate the presence and localization of fibroblast growth factor 2 (FGF2) in human spermatozoa, to determine the relationship of FGF2 levels with conventional semen parameters and to assess the effect of recombinant FGF2 (rFGF2) on sperm recovery in a selection procedure. Western immunoblotting analysis using an antibody against FGF2 revealed an 18-kDa band in sperm protein extracts...
July 21, 2017: Andrology
https://www.readbyqxmd.com/read/28732046/qualitative-differences-in-cellular-immunogenicity-elicited-by-hepatitis-c-virus-t-cell-vaccines-employing-prime-boost-regimens
#14
Wendy G Tan, Iryna Zubkova, Alla Kachko, Frances Wells, Heiko Adler, Gerd Sutter, Marian E Major
T-cell based vaccines have been considered as attractive candidates for prevention of hepatitis C virus (HCV) infections. In this study we compared the magnitude and phenotypic characteristics of CD8+ T-cells induced by three commonly used viral vectors, Adenovirus-5 (Ad5), Vaccinia virus (VV) and Modified Vaccinia Ankara (MVA) expressing the HCV NS3/4A protein. C57/BL6 mice were primed with DNA expressing NS3/4A and boosted with each of the viral vectors in individual groups of mice. We then tracked the vaccine-induced CD8+ T-cell responses using pentamer binding and cytokine production analysis...
2017: PloS One
https://www.readbyqxmd.com/read/28732030/identification-of-odorant-binding-proteins-and-chemosensory-proteins-in-microplitis-mediator-as-well-as-functional-characterization-of-chemosensory-protein-3
#15
Yong Peng, Shan-Ning Wang, Ke-Ming Li, Jing-Tao Liu, Yao Zheng, Shuang Shan, Ye-Qing Yang, Rui-Jun Li, Yong-Jun Zhang, Yu-Yuan Guo
Odorant binding proteins (OBPs) and chemosensory proteins (CSPs) play important roles in transporting semiochemicals through the sensillar lymph to olfactory receptors in insect antennae. In the present study, twenty OBPs and three CSPs were identified from the antennal transcriptome of Microplitis mediator. Ten OBPs (MmedOBP11-20) and two CSPs (MmedCSP2-3) were newly identified. The expression patterns of these new genes in olfactory and non-olfactory tissues were investigated by real-time quantitative PCR (qPCR) measurement...
2017: PloS One
https://www.readbyqxmd.com/read/28731580/rewiring-the-glucose-transportation-and-central-metabolic-pathways-for-overproduction-of-n-acetylglucosamine-in-bacillus-subtilis
#16
Yang Gu, Jieying Deng, Yanfeng Liu, Jianghua Li, Hyun-Dong Shin, Guocheng Du, Jian Chen, Long Liu
N-acetylglucosamine (GlcNAc) is an important amino sugar extensively used in the healthcare field. in a previous study, we constructed the recombinant Bacillus subtilis strain BSGN6-PxylA -glmS-pP43NMK-GNA1 (BN0-GNA1) for microbial production of GlcNAc by pathway design and modular optimization. Here, we further improved the production of GlcNAc by rewiring both the glucose transportation and central metabolic pathways. First, the phosphotransferase system (PTS) was blocked by deletion of three genes, yyzE (encoding the PTS system transporter subunit iiA YyzE), ypqE (encoding the PTS system transporter subunit iiA YpqE), and ptsG (encoding the PTS system glucose-specific EiiCBA component), resulting in 47...
July 21, 2017: Biotechnology Journal
https://www.readbyqxmd.com/read/28731168/osteoclast-regulation-of-osteoblasts-via-rank%C3%A2-rankl-reverse-signal-transduction-in%C3%A2-vitro
#17
Shiqian Zhang, Xiaoyu Wang, Guojun Li, Yang Chong, Jie Zhang, Xinzhen Guo, Baoxin Li, Zhenggang Bi
The treatment of osteoporosis typically inhibits the activity of osteoclasts, which subsequently results in the suppression of bone formation and maintenance, however the underlying mechanism remains to be elucidated. The receptor activator of nuclear factor κ‑B ligand (RANKL)‑receptor activator of nuclear factor κ‑B (RANK) signaling axis is important in the osteoblast regulation of osteoclasts. RANKL surface‑bound molecules expressed on T cells stimulate a reverse signaling transduction in order to regulate the T cells, therefore the present study hypothesized that RANKL expressed on osteoblasts may transfer reverse signals to regulate osteoblasts...
July 20, 2017: Molecular Medicine Reports
https://www.readbyqxmd.com/read/28731026/the-novel-gene-ha117-promotes-in-vitro-and-in-vivo-drug-resistance-in-mouse-colon-tumor-cells
#18
Y X Guo, Y H Xu, G H Zheng, X Q Jin
In this study, we characterized the role of HA117, a novel gene we previously identified, in drug resistance in vitro and in vivo. Briefly, CT26 cells expressing HA117 were obtained by infection with a recombinant adenovirus, and tested for drug sensitivity by methyl thiazolyl tetrazolium assay. In addition, the effect of HA117 on drug sensitivity was assessed in CT26 colon tumors transplanted into nude mice. In vitro, expression of HA117 increased the resistance of CT26 cells to 5-fluorouracil 2.75 times (P<0...
July 21, 2017: Cancer Gene Therapy
https://www.readbyqxmd.com/read/28730788/topoisomerase-i-deregulation-in-laryngeal-squamous-cell-carcinomas-based-on-tissue-microarray-analysis
#19
Theodoros A Papadas, Evangelos Tsiambas, Nicholas S Mastronikolis, Andreas Karameris, Stylianos N Mastronikolis, Athanasios T Papadas, Panagiotis P Fotiades, Vasileios Ragos
PURPOSE: Topoisomerases (types: I/IIa-b/IIIa-b) represent a super-family of nucleic enzymes involved in the DNA replication, transcription, recombination, and also chromosome topological formation. Topoisomerase's I (Topo I- gene location: 20q12) aberrant expression is a frequent genetic event in a variety of solid malignancies. Topo I inhibition promotes cell death due to DNA damage and for this reason it is a target for specific targeted chemotherapy (camptothecin, topotecan, irinotecan)...
May 2017: Journal of B.U.ON.: Official Journal of the Balkan Union of Oncology
https://www.readbyqxmd.com/read/28730763/dna-mismatch-repair-deficiency-in-lung-and-oral-cavity-carcinomas-the-role-of-histogenetic-origin
#20
Konstantina N Spetsotaki, Evangelos Tsiambas, Athanasios Stamatelopoulos, Panagiotis P Fotiades, Ioannis Kastanioudakis, Pericles Tomos, Vasileios Ragos
DNA mismatch repair system (DNA MMR) is a crucial genetic mechanism for DNA homeostasis in prokaryotic and eukaryotic cells. During DNA replication and also recombination, point intra-nucleotide errors including base deletion, insertion, and mis-incorporation happen. These raised abnormalities in the newly synthesized DNA strand could affect negatively the stability of the molecule and the function of the corresponding genes. DNA MMR proteins prevent these errors by recognizing and repairing them, securing directly the normal anatomy of the DNA double strand and indirectly the expression of the genes...
May 2017: Journal of B.U.ON.: Official Journal of the Balkan Union of Oncology
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