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https://www.readbyqxmd.com/read/28531161/development-of-optimized-inhibitor-rnas-allowing-multisite-targeting-of-the-hcv-genome
#1
Cristina Romero-López, Thomas Lahlali, Beatriz Berzal-Herranz, Alfredo Berzal-Herranz
Engineered multivalent drugs are promising candidates for fighting infection by highly variable viruses, such as HCV. The combination into a single molecule of more than one inhibitory domain, each with its own target specificity and even a different mechanism of action, results in drugs with potentially enhanced therapeutic properties. In the present work, the anti-HCV chimeric inhibitor RNA HH363-10, which has a hammerhead catalytic domain and an aptamer RNA domain, was subjected to an in vitro selection strategy to isolate ten different optimised chimeric inhibitor RNAs...
May 22, 2017: Molecules: a Journal of Synthetic Chemistry and Natural Product Chemistry
https://www.readbyqxmd.com/read/28523913/expression-and-characterization-of-windmill-palm-tree-trachycarpus-fortunei-peroxidase-by-pichia-pastoris
#2
Boting Wen, Margaret R Baker, Hongwei Zhao, Zongjun Cui, Qing Xiao Li
Currently, commercial plant peroxidases are all native and are isolated from plants such as horseradish and soybean. No recombinant plant peroxidase products have been available on the commercial market. The gene encoding peroxidase was cloned from windmill palm tree leaves. The codon-optimized gene was transformed into Pichia pastoris for expression. The recombinant windmill palm tree peroxidase (rWPTP) expressed by P. pastoris showed high stability under pH 2-10, temperature up to 70 C, to many metallic salts and organic solvents...
May 19, 2017: Journal of Agricultural and Food Chemistry
https://www.readbyqxmd.com/read/28522266/site-specific-conjugation-of-fibroblast-growth-factor-2-fgf2-based-on-incorporation-of-alkyne-reactive-unnatural-amino-acid
#3
K W Swiderska, A Szlachcic, A Czyrek, M Zakrzewska, J Otlewski
Recent advances in site-specific protein modification include the increasingly popular incorporation of unnatural amino acid(s) using amber codon, a method developed by Schultz and coworkers. In this study, we employ this technique to introduce propargyllysine (PrK) in human fibroblast growth factor 2 (FGF2). Owing to an alkyne moiety in its side chain, PrK is compatible with Cu(I)-catalyzed azide-alkyne 1,3-dipolar cycloaddition (CuAAC). We successfully tested CuAAC-mediated conjugation of FGF2 with two compounds - a fluorophore carboxyrhodamine 110 or a cytotoxic drug monomethyl auristatin E (MMAE)...
May 5, 2017: Bioorganic & Medicinal Chemistry
https://www.readbyqxmd.com/read/28521201/validation-of-the-use-of-an-artificial-mitochondrial-reporter-dna-vector-containing-a-cytomegalovirus-promoter-for-mitochondrial-transgene-expression
#4
Yuma Yamada, Takuya Ishikawa, Hideyoshi Harashima
Mitochondria have their own gene expression system that is independent of the nuclear system, and control cellular functions in cooperation with the nucleus. While a number of useful technologies for achieving nuclear transgene expression have been reported, only a few have focused on mitochondria. In this study, we validated the utility of an artificial mitochondrial DNA vector with a virus promoter on mitochondrial transgene expression. We designed and constructed pCMV-mtLuc (CGG) that contains a CMV promotor derived from Cytomegalovirus and an artificial mitochondrial genome with a NanoLuc (Nluc) luciferase gene that records adjustments to the mitochondrial codon system...
May 10, 2017: Biomaterials
https://www.readbyqxmd.com/read/28520731/genetically-engineered-probiotic-for-the-treatment-of-phenylketonuria-pku-assessment-of-a-novel-treatment-in-vitro-and-in-the-pahenu2-mouse-model-of-pku
#5
Katherine E Durrer, Michael S Allen, Ione Hunt von Herbing
Phenylketonuria (PKU) is a genetic disease characterized by the inability to convert dietary phenylalanine to tyrosine by phenylalanine hydroxylase. Given the importance of gut microbes in digestion, a genetically engineered microbe could potentially degrade some ingested phenylalanine from the diet prior to absorption. To test this, a phenylalanine lyase gene from Anabaena variabilis (AvPAL) was codon-optimized and cloned into a shuttle vector for expression in Lactobacillus reuteri 100-23C (pHENOMMenal). Functional expression of AvPAL was determined in vitro, and subsequently tested in vivo in homozygous PAHenu2 (PKU model) mice...
2017: PloS One
https://www.readbyqxmd.com/read/28515770/high-level-expression-of-recombinant-human-growth-hormone-in-escherichia-coli-crucial-role-of-translation-initiation-region
#6
Mahsa Ghavim, Khalil Abnous, Fatemeh Arasteh, Sahar Taghavi, Maryam Sadat Nabavinia, Mona Alibolandi, Mohammad Ramezani
For high-throughput production of recombinant protein in Escherichia coli (E. coli), besides important parameters such as efficient vector with strong promoter and compatible host, other important issues including codon usage, rare codons, and GC content specially at N-terminal region should be considered. In the current study, the effect of decreasing the percentage of GC nucleotides and optimizing codon usage at N-terminal region of human growth hormone (hGH) cDNA on the level of its expression in E. coli were investigated...
April 2017: Research in Pharmaceutical Sciences
https://www.readbyqxmd.com/read/28514980/evaluation-of-benzimidazole-resistance-status-in-ascaridia-galli
#7
B Tarbiat, D S Jansson, E Tydén, J Höglund
Susceptability of Ascaridia galli to benzimidazole (BZ) was investigated using faecal egg count reduction test (FECRT), in ovo larval development test (LDT) and genetic markers (mutations at codons 167, 198 and 200 of β-tubulin gene). Six flocks (F1-F6) of a commercial laying hen farm with different number of exposure to BZ were recruited. The FECR was calculated by analyzing individual faeces (F1, F2, F4 and F5) before and 10 days after treatment. The LDT was performed on parasite eggs from pooled samples from F1 to F6 and LC50 and LC99 were calculated...
May 18, 2017: Parasitology
https://www.readbyqxmd.com/read/28502718/mitochondrial-nd5-mutation-mediated-elevated-ros-regulates-apoptotic-pathway-epigenetically-in-a-p53-dependent-manner-for-generating-pro-cancerous-phenotypes
#8
Rajnish Kumar Singh, Sunil Saini, Dileep Verma, Ponnusamy Kalaiarasan, Rameshwar N K Bamezai
We have previously observed concomitant events of mutations in mitochondrial and nuclear genes, along with elevated reactive oxygen species (ROS) and differential methylation within the promoters of nuclear genes in tumors and in vitro experiments of tumorigenesis. These observations have made it pertinent to replicate and understand the role of acquired mitochondrial condition in tuning a cell to accomplish a pro-cancerous state. Using a codon optimized vector system for exogenous over-expression and mitochondrial localization; we have characterized here the role of over-expressed wild type mtND5 and one of its non-synonymous somatic mutation, ND5:P265H...
May 11, 2017: Mitochondrion
https://www.readbyqxmd.com/read/28501472/prokaryotic-expression-of-a-codon-optimized-capsid-gene-from-duck-circovirus-and-its-application-to-an-indirect-elisa
#9
Cui Yang, Yu Xu, Renyong Jia, Pengfei Li, Lei Zhang, Mingshu Wang, Dekang Zhu, Shun Chen, Mafeng Liu, Zhongqiong Yin, Anchun Cheng
Duck circovirus (DuCV), as a causative agent of long-term immunosuppressive disease, has caused heavy damage to waterfowl breeding worldwide. In this study, the full-length Cap (capsid) gene of DuCV was expressed in Escherichia coli (E. coli) for the first time by optimizing the codons in its nuclear localization signal (NLS) regions. The recombinant protein was expressed as inclusion bodies, and the quantification of purified Cap protein could reach 0.29mgmL(-1). Moreover, an indirect ELISA method (Cap-ELISA) was established based on the recombinant Cap protein...
May 10, 2017: Journal of Virological Methods
https://www.readbyqxmd.com/read/28494467/expression-and-characterization-of-functional-recombinant-bet-v-1-0101-in-the-chloroplast-of-chlamydomonas-reinhardtii
#10
Sonja Hirschl, Claudia Ralser, Claudia Asam, Alessandro Gangitano, Sara Huber, Christof Ebner, Barbara Bohle, Martin Wolf, Peter Briza, Fatima Ferreira, Christoph Griesbeck, Michael Wallner
BACKGROUND: Allergen immunotherapy (AIT) still plays a minor role in the treatment of allergic diseases. To improve the acceptance of AIT by allergic patients, the treatment has to become more convenient and efficacious. One possibility is the oral application of allergens or derivatives thereof. Therefore, we sought to produce a recombinant allergen in the green alga Chlamydomonas reinhardtii as a novel production platform. METHODS: The major birch pollen allergen Bet v 1 was selected as candidate molecule, and a codon-optimized gene was synthesized and stably integrated into the microalga C...
May 12, 2017: International Archives of Allergy and Immunology
https://www.readbyqxmd.com/read/28489302/characterization-of-a-furan-aldehyde-tolerant-%C3%AE-xylosidase-%C3%AE-arabinosidase-obtained-through-a-synthetic-metagenomics-approach
#11
Mukil Maruthamuthu, Diego Javier Jiménez, Jan Dirk van Elsas
AIMS: The aim of the study was to characterize ten hemicellulolytic enzymes obtained from a wheat straw-degrading microbial consortium. METHODS AND RESULTS: Based on previous metagenomics analyses, ten glycosyl hydrolases were selected, codon-optimized, synthetized, cloned and expressed in Escherichia coli. Nine of the overexpressed recombinant proteins accumulated in cellular inclusion bodies, whereas one, a 37.5 kDa protein encoded by gene xylM1989, was found in the soluble fractions...
May 10, 2017: Journal of Applied Microbiology
https://www.readbyqxmd.com/read/28488117/fusion-expression-of-the-pgla-am1-with-native-structure-and-evaluation-of-its-anti-helicobacter-pylori-activity
#12
Xiaolin Zhang, Anmin Jiang, Guisheng Wang, Hao Yu, Banghua Qi, Youyi Xiong, Guoliang Zhou, Meisong Qin, Jinfeng Dou, Jianfei Wang
Helicobacter pylori (H. pylori) shows increasingly enhanced resistance to various antibiotics, and its eradication has become a major problem in medicine. The antimicrobial peptide PGLa-AM1 is a short peptide with 22 amino acids and exhibits strong antibacterial activity. In this study, we investigated whether it has anti-H. pylori activity for the further development of anti-H. pylori drugs to replace existing antibiotics. However, the natural antimicrobial peptide PGLa-AM1 shows a low yield and is difficult to separate, limiting its application...
May 10, 2017: Applied Microbiology and Biotechnology
https://www.readbyqxmd.com/read/28486986/the-link-between-adjacent-codon-pairs-and-mrna-stability
#13
Yuriko Harigaya, Roy Parker
BACKGROUND: Evidence in diverse organisms suggests that codon optimality is a major determinant of mRNA translation and degradation. Codon optimality is thought to act by modulating the efficiency of ribosome elongation. In Saccharomyces cerevisiae, a recent study has identified 17 adjacent codon pairs that mediate strong inhibition of translation elongation. However, relationships between the inhibitory codon pairs and other aspects of gene expression are unknown. RESULTS: To gain insights into how the inhibitory codon pairs may affect aspects of gene expression, we utilized existing datasets to conduct genome-scale analyses in S...
May 10, 2017: BMC Genomics
https://www.readbyqxmd.com/read/28484046/exploring-the-%C3%AE-ctd-amino-acid-residue-requirements-for-productive-interaction-between-spx-and-rna-polymerase-of-bacillus-subtilis
#14
Cierra Birch, Madison Davis, Lea Mbengi, Peter Zuber
Bacillus subtilis Spx is a global transcriptional regulator that is conserved among gram-positive bacteria, in which Spx is required for preventing oxidatively induced proteotoxicity. Upon stress induction, Spx engages RNA polymerase (RNAP) through interaction with the C-terminal domain of the rpoA-encoded RNAP α subunit (αCTD). Previous mutational analysis of rpoA revealed that substitutions of Y263 in αCTD severely impaired Spx-activated transcription. Attempts to substitute alanine for αCTD R261, R268, R289, E255, E298 and K294 were unsuccessful, suggesting that these residues are essential...
May 8, 2017: Journal of Bacteriology
https://www.readbyqxmd.com/read/28476488/improving-pertuzumab-production-by-gene-optimization-and-proper-signal-peptide-selection
#15
Amin Ramezani, Elham Mahmoudi Maymand, Mahsa Yazdanpanah-Samani, Ahmad Hosseini, Fatemeh Sadat Toghraie, Abbas Ghaderi
Using proper signal peptide and codon optimization are important factors that must be considered when designing the vector to increase protein expression in Chinese Hamster Ovary (CHO) cells. The aim of the present study is to investigate how to enhance Pertuzumab production through heavy and light chain coding gene optimization and proper signal peptide selection. First, CHO-K1 cells were transiently transfected with whole-antibody-gene-optimized, variable-regions-optimized and non-optimized constructs and then we employed five different signal peptides to improve the secretion efficiency of Pertuzumab...
May 3, 2017: Protein Expression and Purification
https://www.readbyqxmd.com/read/28475588/widespread-position-specific-conservation-of-synonymous-rare-codons-within-coding-sequences
#16
Julie L Chaney, Aaron Steele, Rory Carmichael, Anabel Rodriguez, Alicia T Specht, Kim Ngo, Jun Li, Scott Emrich, Patricia L Clark
Synonymous rare codons are considered to be sub-optimal for gene expression because they are translated more slowly than common codons. Yet surprisingly, many protein coding sequences include large clusters of synonymous rare codons. Rare codons at the 5' terminus of coding sequences have been shown to increase translational efficiency. Although a general functional role for synonymous rare codons farther within coding sequences has not yet been established, several recent reports have identified rare-to-common synonymous codon substitutions that impair folding of the encoded protein...
May 5, 2017: PLoS Computational Biology
https://www.readbyqxmd.com/read/28473715/rationally-co-targeting-divergent-pathways-in-kras-wild-type-colorectal-cancers-by-canscript-technology-reveals-tumor-dependence-on-notch-and-erbb2
#17
Nilesh Brijwani, Misti Jain, Muthu Dhandapani, Farrah Zahed, Pragnashree Mukhopadhyay, Manjusha Biswas, Deepak Khatri, Vinod D Radhakrishna, Biswanath Majumder, Padhma Radhakrishnan, Saravanan Thiyagarajan
KRAS mutation status can distinguish between metastatic colorectal carcinoma (mCRC) patients who may benefit from therapies that target the epidermal growth factor receptor (EGFR), such as cetuximab. However, patients whose tumors harbor mutant KRAS (codons 12/13, 61 and 146) are often excluded from EGFR-targeted regimens, while other patients with wild type KRAS will sometimes respond favorably to these same drugs. These conflicting observations suggest that a more robust approach to individualize therapy may enable greater frequency of positive clinical outcome for mCRC patients...
May 4, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28472670/heterologous-production-of-different-styrene-oxide-isomerases-for-the-highly-efficient-synthesis-of-phenylacetaldehyde
#18
Michel Oelschlägel, Lysann Richter, Anna Stuhr, Sarah Hofmann, Michael Schlömann
The styrene oxide isomerase (SOI, StyC) represents a key enzyme of the styrene-degrading pathway and has been discussed as promising biocatalyst during recent studies. The enzyme enables the synthesis of pure phenylacetaldehyde from styrene oxide. In this study the native as well as the corresponding codon-optimized genes of three different SOIs from Rhodococcus opacus 1CP (StyC-1CP), Sphingopyxis fribergensis Kp5.2 (StyC-Kp5.2), and Pseudomonas fluorescens ST (StyC-ST) were investigated for the expression in Escherichia coli BL21(DE3)pLysS...
May 1, 2017: Journal of Biotechnology
https://www.readbyqxmd.com/read/28470625/an-anti-amoebic-vaccine-generation-of-the-recombinant-antigen-lc3-from-entamoeba-histolytica-linked-to-mutated-exotoxin-a-pe%C3%AE-iii-via-the-pichia-pastoris-system
#19
Sandra Luz Martínez-Hernández, Daniel Cervantes-García, Martín Muñoz-Ortega, Liseth R Aldaba-Muruato, Victor M Loera-Muro, Jorge A Ascacio-Martínez, María de Jesús Loera-Arias, Roberto Montes de Oca-Luna, Javier Ventura-Juárez
OBJECTIVE: To generate an immunogenic chimeric protein containing the Entamoeba histolytica LC3 fragment fused to the retrograde delivery domains of exotoxin A of Pseudomonas aeruginosa and KDEL3 for use as an effective vaccine. RESULTS: A codon-optimized synthetic gene encoding the PEΔIII-LC3-KDEL3 fusion construct was designed for expression in Pichia pastoris. This transgene was subcloned into the plasmid pPIC9 for methanol-inducible expression. After transformation and selection of positive-transformed clones by PCR, the expression of the recombinant protein PEΔIII-LC3-KDEL3 was elicited...
May 3, 2017: Biotechnology Letters
https://www.readbyqxmd.com/read/28468662/heterologous-expression-of-abaecin-peptide-from-apis-mellifera-in-pichia-pastoris
#20
Denis Prudencio Luiz, Juliana Franco Almeida, Luiz Ricardo Goulart, Nilson Nicolau-Junior, Carlos Ueira-Vieira
BACKGROUND: Antimicrobial peptides (AMPs) are the first line of host immune defense against pathogens. Among AMPs from the honeybee Apis mellifera, abaecin is a major broad-spectrum antibacterial proline-enriched cationic peptide. RESULTS: For heterologous expression of abaecin in Pichia pastoris, we designed an ORF with HisTag, and the codon usage was optimized. The gene was chemically synthetized and cloned in the pUC57 vector. The new ORF was sub-cloned in the pPIC9 expression vector and transformed into P...
May 3, 2017: Microbial Cell Factories
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