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Polymerase substrate specificity

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https://www.readbyqxmd.com/read/28107985/enhanced-sensitivity-for-detection-of-plasmodium-falciparum-gametocytes-by-magnetic-nanoparticles-combined-with-enzyme-substrate-system
#1
Tienrat Tangchaikeeree, Piamsiri Sawaisorn, Sangdao Somsri, Duangporn Polpanich, Chaturong Putaporntip, Pramuan Tangboriboonrat, Rachanee Udomsangpetch, Kulachart Jangpatarapongsa
The highly sensitive and specific detection of Pfg377 gene of Plasmodium falciparum gametocyte using Magnetic Nanoparticles PCR Enzyme-Linked Gene Assay (MELGA) was successfully developed. The MELGA included amplification of the Pfg377 gene by polymerase chain reaction (PCR) using magnetic nanoparticles (MNPs)-conjugated forward primer and biotinylated reverse primer, followed by post-analytical process using horseradish peroxidase (HRP)-conjugated streptavidin (SA). The complexes of MELGA product were incubated with the peroxidase substrate and hydrogen peroxide to produce the signal for colorimetric measurement...
March 1, 2017: Talanta
https://www.readbyqxmd.com/read/28095779/poly-adp-ribose-polymerase-inhibitors-activate-the-p53-signaling-pathway-in-neural-stem-progenitor-cells
#2
Akiko Okuda, Suguru Kurokawa, Masanori Takehashi, Aika Maeda, Katsuya Fukuda, Yukari Kubo, Hyuma Nogusa, Tomoka Takatani-Nakase, Shujiro Okuda, Kunihiro Ueda, Seigo Tanaka
BACKGROUND: Poly(ADP-ribose) polymerase 1 (PARP-1), which catalyzes poly(ADP-ribosyl)ation of proteins by using NAD(+) as a substrate, plays a key role in several nuclear events, including DNA repair, replication, and transcription. Recently, PARP-1 was reported to participate in the somatic cell reprogramming process. Previously, we revealed a role for PARP-1 in the induction of neural apoptosis in a cellular model of cerebral ischemia and suggested the possible use of PARP inhibitors as a new therapeutic intervention...
January 17, 2017: BMC Neuroscience
https://www.readbyqxmd.com/read/28057773/inverse-remodelling-of-k2p3-1-k-channel-expression-and-action-potential-duration-in-left-ventricular-dysfunction-and-atrial-fibrillation-implications-for-patient-specific-antiarrhythmic-drug-therapy
#3
Constanze Schmidt, Felix Wiedmann, Xiao-Bo Zhou, Jordi Heijman, Niels Voigt, Antonius Ratte, Siegfried Lang, Stefan M Kallenberger, Chiara Campana, Alexander Weymann, Raffaele De Simone, Gabor Szabo, Arjang Ruhparwar, Klaus Kallenbach, Matthias Karck, Joachim R Ehrlich, István Baczkó, Martin Borggrefe, Ursula Ravens, Dobromir Dobrev, Hugo A Katus, Dierk Thomas
AIMS: Atrial fibrillation (AF) prevalence increases with advanced stages of left ventricular (LV) dysfunction. Remote proarrhythmic effects of ventricular dysfunction on atrial electrophysiology remain incompletely understood. We hypothesized that repolarizing K2P3.1 K(+ )channels, previously implicated in AF pathophysiology, may contribute to shaping the atrial action potential (AP), forming a specific electrical substrate with LV dysfunction that might represent a target for personalized antiarrhythmic therapy...
January 4, 2017: European Heart Journal
https://www.readbyqxmd.com/read/28056947/triptolide-induces-hepatotoxicity-via-inhibition-of-cyp450s-in-rat-liver-microsomes
#4
Yan Lu, Tong Xie, Yajie Zhang, Fuqiong Zhou, Jie Ruan, Weina Zhu, Huaxu Zhu, Zhe Feng, Xueping Zhou
BACKGROUND: Triptolide (TP), an active constituent of Tripterygium wilfordii, possesses numerous pharmacological activities. However, its effects on cytochrome P450 enzymes (CYP450s) in rats remain unexplored. METHODS: In this study, the effects of triptolide on the six main CYP450 isoforms (1A2, 2C9, 2C19, 2D6, 2E1, and 3A) were investigated both in vivo and in vitro. We monitored the body weight, survival proportions, liver index, changes in pathology, and biochemical index upon TP administration, in vivo...
January 5, 2017: BMC Complementary and Alternative Medicine
https://www.readbyqxmd.com/read/28041959/substrate-selectivity-of-dengue-and-zika-virus-ns5-polymerase-towards-2-modified-nucleotide-analogues
#5
Supanee Potisopon, François Ferron, Véronique Fattorini, Barbara Selisko, Bruno Canard
In targeting the essential viral RNA-dependent RNA-polymerase (RdRp), nucleotide analogues play a major role in antiviral therapies. In the Flaviviridae family, the hepatitis C virus (HCV) can be eradicated from chronically infected patients using a combination of drugs which generally include the 2'-modified uridine analogue Sofosbuvir, delivered as nucleotide prodrug. Dengue and Zika viruses are emerging flaviviruses whose RdRp is closely related to that of HCV, yet no nucleoside drug has been clinically approved for these acute infections...
December 30, 2016: Antiviral Research
https://www.readbyqxmd.com/read/28005463/cdk-regulation-of-transcription-by-rnap-ii-not-over-til-it-s-over
#6
Robert P Fisher
Transcription by RNA polymerase (RNAP) II is regulated at multiple steps by phosphorylation, catalyzed mainly by members of the cyclin-dependent kinase (CDK) family. The CDKs involved in transcription have overlapping substrate specificities, but play largely non-redundant roles in coordinating gene expression. Novel functions and targets of CDKs have recently emerged at the end of the transcription cycle, when the primary transcript is cleaved, and in most cases polyadenylated, and transcription is terminated by the action of the "torpedo" exonuclease Xrn2, which is a CDK substrate...
December 22, 2016: Transcription
https://www.readbyqxmd.com/read/28000329/polymerase-mediated-site-specific-incorporation-of-a-synthetic-fluorescent-isomorphic-g-surrogate-into-rna
#7
Yao Li, Andrea Fin, Lisa McCoy, Yitzhak Tor
An enzyme-mediated approach for the assembly of singly modified RNA constructs in which specific G residues are replaced with (th) G, an emissive isomorphic G surrogate, is reported. Transcription in the presence of (th) G and native nucleoside triphosphates enforces initiation with the unnatural analogue, yielding 5'-end modified transcripts that can be mono-phosphorylated and ligated to provide longer site-specifically modified RNA constructs. The scope of this unprecedented enzymatic approach to non-canonical purine-containing RNAs is explored via the assembly of several altered hammerhead (HH) ribozymes and a singly modified HH substrate...
January 24, 2017: Angewandte Chemie
https://www.readbyqxmd.com/read/27982035/global-repositioning-of-transcription-start-sites-in-a-plant-fermenting-bacterium
#8
Magali Boutard, Laurence Ettwiller, Tristan Cerisy, Adriana Alberti, Karine Labadie, Marcel Salanoubat, Ira Schildkraut, Andrew C Tolonen
Bacteria respond to their environment by regulating mRNA synthesis, often by altering the genomic sites at which RNA polymerase initiates transcription. Here, we investigate genome-wide changes in transcription start site (TSS) usage by Clostridium phytofermentans, a model bacterium for fermentation of lignocellulosic biomass. We quantify expression of nearly 10,000 TSS at single base resolution by Capp-Switch sequencing, which combines capture of synthetically capped 5' mRNA fragments with template-switching reverse transcription...
December 16, 2016: Nature Communications
https://www.readbyqxmd.com/read/27966711/development-of-a-point-of-care-diagnostic-for-influenza-detection-with-antiviral-treatment-effectiveness-indication
#9
Richard C Murdock, Karen M Gallegos, Joshua A Hagen, Nancy Kelley-Loughnane, Alison A Weiss, Ian Papautsky
Currently, diagnosis of influenza is performed either through tedious polymerase chain reaction (PCR) or through rapid antigen detection assays. The rapid antigen detection assays available today are highly specific but not very sensitive, and most importantly, lack the ability to show if the strain of influenza detected is susceptible to antiviral agents, such as Tamiflu and Relenza. The ability to rapidly determine if a patient has an infectious disease and what type of treatment the infection will respond to, would significantly reduce the treatment decision time, shorten the impact of symptoms, and minimize transfer to others...
January 17, 2017: Lab on a Chip
https://www.readbyqxmd.com/read/27913570/revisiting-the-human-polypeptide-galnac-t1-and-t13-paralogs
#10
María Florencia Festari, Felipe Trajtenberg, Nora Berois, Sergio Pantano, Leslie Revoredo, Yun Kong, Patricia Solari-Saquieres, Yoshiki Narimatsu, Teresa Freire, Sylvie Bay, Carlos Robello, Jean Bénard, Thomas A Gerken, Henrik Clausen, Eduardo Osinaga
Polypeptide GalNAc-transferases (GalNAc-Ts) constitute a family of 20 human glycosyltransferases (comprising 9 subfamilies), which initiate mucin-type O-glycosylation. The O-glycoproteome is thought to be differentially regulated via the different substrate specificities and expression patterns of each GalNAc-T isoforms. Here, we present a comprehensive in vitro analysis of the peptide substrate specificity of GalNAc-T13, showing that it essentially overlaps with the ubiquitous expressed GalNAc-T1 isoform found in the same subfamily as T13...
January 2017: Glycobiology
https://www.readbyqxmd.com/read/27898685/high-resolution-phenotypic-landscape-of-the-rna-polymerase-ii-trigger-loop
#11
Chenxi Qiu, Olivia C Erinne, Jui M Dave, Ping Cui, Huiyan Jin, Nandhini Muthukrishnan, Leung K Tang, Sabareesh Ganesh Babu, Kenny C Lam, Paul J Vandeventer, Ralf Strohner, Jan Van den Brulle, Sing-Hoi Sze, Craig D Kaplan
The active sites of multisubunit RNA polymerases have a "trigger loop" (TL) that multitasks in substrate selection, catalysis, and translocation. To dissect the Saccharomyces cerevisiae RNA polymerase II TL at individual-residue resolution, we quantitatively phenotyped nearly all TL single variants en masse. Three mutant classes, revealed by phenotypes linked to transcription defects or various stresses, have distinct distributions among TL residues. We find that mutations disrupting an intra-TL hydrophobic pocket, proposed to provide a mechanism for substrate-triggered TL folding through destabilization of a catalytically inactive TL state, confer phenotypes consistent with pocket disruption and increased catalysis...
November 2016: PLoS Genetics
https://www.readbyqxmd.com/read/27885504/plant-dicer-like-proteins-double-stranded-rna-cleaving-enzymes-for-small-rna-biogenesis
#12
Akihito Fukudome, Toshiyuki Fukuhara
Dicer, a double-stranded RNA (dsRNA)-specific endoribonuclease, plays an essential role in triggering both transcriptional and post-transcriptional gene silencing in eukaryotes by cleaving dsRNAs or single-stranded RNAs bearing stem-loop structures such as microRNA precursor transcripts into 21- to 24-nt small RNAs. Unlike animals, plants have evolved to utilize at least four Dicer-like (DCL) proteins. Extensive genetic studies have revealed that each DCL protein participates in a specific gene silencing pathway, with some redundancy...
January 2017: Journal of Plant Research
https://www.readbyqxmd.com/read/27881682/stabilization-of-luciferase-from-renilla-reniformis-using-random-mutations
#13
Megumi Shigehisa, Norie Amaba, Shigeki Arai, Chisato Higashi, Ryo Kawanabe, Ayano Matsunaga, Fina Amreta Laksmi, Masao Tokunaga, Matsujiro Ishibashi
We expressed luciferase (RLuc) from Renilla reniformis in Escherichia coli RLuc was purified using a Ni-NTA column and subsequently characterized. It was unstable in acidic solutions and at 30°C. To increase the stability of RLuc, the Rluc gene was randomly mutated using error-prone polymerase chain reaction. E. coli harboring the mutated gene was screened by detecting luminescence on a plate containing the substrate coelenterazine at 34°C. Three mutants, i.e. N264SS287P, N178D and F116LI137V, were obtained...
January 2017: Protein Engineering, Design & Selection: PEDS
https://www.readbyqxmd.com/read/27833949/the-emerging-picture-of-cdk9-p-tefb-more-than-20-years-of-advances-since-pitalre
#14
REVIEW
Nikolas Ferreira Dos Santos Paparidis, Maxwell Castro Durvale, Fernanda Canduri
CDK9 is a prominent member of the transcriptional CDKs subfamily, a group of kinases whose function is to control the primary steps of mRNA synthesis and processing by eukaryotic RNA polymerase II. As a cyclin-dependent kinase, CDK9 activation in vivo depends upon its association with T-type cyclins to assemble the positive transcription elongation factor (P-TEFb). Although CDK9/P-TEFb phosphorylates the C-terminal domain of RNAP II in the same positions targeted by CDK7 (TFIIH) and CDK8 (Mediator), the former does not participate in the transcription initiation, but rather plays a unique role by driving the polymerase to productive elongation...
November 11, 2016: Molecular BioSystems
https://www.readbyqxmd.com/read/27817742/parg-inhibitors-and-functional-parg-inhibition-models
#15
Yuka Sasaki, Miyuki Hozumi, Hiroaki Fujimori, Yasufumi Murakami, Fumiaki Koizumi, Kengo Inoue, Mitsuko Masutani
Poly(ADP-ribose) polymerases (PARPs) family proteins catalyze poly(ADP-ribosylation) (PARylation) by conjugating ADP-ribose residues repeatedly on amino acid residues using nicotinamide adenine dinucleotide as a substrate. The inhibitors of PARP widely block DNA repair processes and are currently examined in clinical trials of cancer therapy. Poly(ADP-ribose) glycohydrolase (PARG) is the main nuclear enzyme, which digests poly(ADP-ribose) into ADP-ribose. PARG inhibitor could also be considered as a chemotherapeutic agent for cancer, because of its involvement in DNA repair...
2016: Current Protein & Peptide Science
https://www.readbyqxmd.com/read/27808482/crystal-structure-of-ralstonia-eutropha-polyhydroxyalkanoate-synthase-c-terminal-domain-and-reaction-mechanisms
#16
Jieun Kim, Yeo-Jin Kim, So Young Choi, Sang Yup Lee, Kyung-Jin Kim
Polyhydroxyalkanoates (PHAs) are natural polyesters synthesized by numerous microorganisms as energy and reducing power storage materials, and have attracted much attention as substitutes for petroleum-based plastics. Here, we report the first crystal structure of Ralstonia eutropha PHA synthase at 1.8 Å resolution and structure-based mechanisms for PHA polymerization. RePhaC1 contains two distinct domains, the N-terminal (RePhaC1ND ) and C-terminal domains (RePhaC1CD ), and exists as a dimer. RePhaC1CD catalyzes polymerization via non-processive ping-pong mechanism using a Cys-His-Asp catalytic triad...
November 3, 2016: Biotechnology Journal
https://www.readbyqxmd.com/read/27754888/regulation-of-transcription-initiation-by-gfh-factors-from-deinococcus-radiodurans
#17
Aleksei Agapov, Daria Esyunina, Danil Pupov, Andrey Kulbachinskiy
The Gre-family transcription factors bind within the secondary channel of bacterial RNA polymerase (RNAP) directly modulating its catalytic activities. Universally conserved Gre factors activate RNA cleavage by RNAP, by chelating catalytic metal ions in the RNAP active site, and facilitate both promoter escape and transcription elongation. Gfh factors are Deinococcus/Thermus -specific homologues of Gre factors whose transcription functions remain poorly understood. Recently, we found that Gfh1 and Gfh2 proteins from Deinococcus radiodurans dramatically stimulate RNAP pausing during transcription elongation in the presence of Mn(2+) but not Mg(2+) ions...
October 17, 2016: Biochemical Journal
https://www.readbyqxmd.com/read/27744351/rna-editing-tutase-1-structural-foundation-of-substrate-recognition-complex-interactions-and-drug-targeting
#18
Lional Rajappa-Titu, Takuma Suematsu, Paola Munoz-Tello, Marius Long, Özlem Demir, Kevin J Cheng, Jason R Stagno, Hartmut Luecke, Rommie E Amaro, Inna Aphasizheva, Ruslan Aphasizhev, Stéphane Thore
Terminal uridyltransferases (TUTases) execute 3' RNA uridylation across protists, fungi, metazoan and plant species. Uridylation plays a particularly prominent role in RNA processing pathways of kinetoplastid protists typified by the causative agent of African sleeping sickness, Trypanosoma brucei In mitochondria of this pathogen, most mRNAs are internally modified by U-insertion/deletion editing while guide RNAs and rRNAs are U-tailed. The founding member of TUTase family, RNA editing TUTase 1 (RET1), functions as a subunit of the 3' processome in uridylation of gRNA precursors and mature guide RNAs...
December 15, 2016: Nucleic Acids Research
https://www.readbyqxmd.com/read/27742790/matrix-mechanics-controls-fhl2-movement-to-the-nucleus-to-activate-p21-expression
#19
Naotaka Nakazawa, Aneesh R Sathe, G V Shivashankar, Michael P Sheetz
Substrate rigidity affects many physiological processes through mechanochemical signals from focal adhesion (FA) complexes that subsequently modulate gene expression. We find that shuttling of the LIM domain (domain discovered in the proteins, Lin11, Isl-1, and Mec-3) protein four-and-a-half LIM domains 2 (FHL2) between FAs and the nucleus depends on matrix mechanics. In particular, on soft surfaces or after the loss of force, FHL2 moves from FAs into the nucleus and concentrates at RNA polymerase (Pol) II sites, where it acts as a transcriptional cofactor, causing an increase in p21 gene expression that will inhibit growth on soft surfaces...
November 1, 2016: Proceedings of the National Academy of Sciences of the United States of America
https://www.readbyqxmd.com/read/27739412/induction-of-apoptosis-in-tnf-treated-l929-cells-in-the-presence-of-necrostatin-1
#20
Hirofumi Sawai
It has been shown that necroptosis-caspase-independent programmed necrotic cell death-can be induced by treatment with tumor necrosis factor (TNF) in the L929 murine fibrosarcoma cell line, even in the absence of a caspase inhibitor. Although it was reported that necrostatin-1-a specific inhibitor of necroptosis-inhibited TNF-induced necroptosis in L929 cells, it has not been elucidated whether the cells eventually die by apoptosis in the presence of necrostatin-1. In this paper, induction of apoptosis was demonstrated in TNF-treated L929 cells in the presence of necrostatin-1...
October 7, 2016: International Journal of Molecular Sciences
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