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https://www.readbyqxmd.com/read/28448625/structural-and-functional-studies-of-ferredoxin-and-oxygenase-components-of-3-nitrotoluene-dioxygenase-from-diaphorobacter-sp-strain-ds2
#1
Archana Kumari, Deepak Singh, S Ramaswamy, Gurunath Ramanathan
3-nitrotoluene dioxygenase (3NTDO) from Diaphorobacter sp. strain DS2 catalyses the conversion of 3-nitrotoluene (3NT) into a mixture of 3- and 4-methylcatechols with release of nitrite. We report here, X-ray crystal structures of oxygenase and ferredoxin components of 3NTDO at 2.9 Å and 2.4 Å, respectively. The residues responsible for nitrite release in 3NTDO were further probed by four single and two double mutations in the catalytic site of α-subunit of the dioxygenase. Modification of Val 350 to Phe, Ile 204 to Ala, and Asn258 to Val by site directed mutagenesis resulted in inactive enzymes revealing the importance of these residues in catalysis...
2017: PloS One
https://www.readbyqxmd.com/read/28446668/serine-235-is-the-primary-ns5a-hyper-phosphorylation-site-responsible-for-hcv-replication
#2
Shih-Chin Hsu, Chieh-Wen Lo, Ting-Chun Pan, Kuan-Ying Lee, Ming-Jiun Yu
The non-structural protein 5A (NS5A) of the hepatitis C virus (HCV) is a phosphoprotein with two phosphorylation states: hypo- and hyper-phosphorylation. Genetic mutation studies have demonstrated a cluster of serine residues responsible for NS5A hyper-phosphorylation and functions in viral replication and assembly; however, the phosphorylation levels and potential interactions among the serine residues are unclear. We used three specific antibodies to measure NS5A phosphorylation at S222, S235, and S238 that were identified in our previous proteomics study...
April 26, 2017: Journal of Virology
https://www.readbyqxmd.com/read/28446513/frequent-hypomorphic-alleles-account-for-a-significant-fraction-of-abca4-disease-and-distinguish-it-from-age-related-macular-degeneration
#3
Jana Zernant, Winston Lee, Frederick T Collison, Gerald A Fishman, Yuri V Sergeev, Kaspar Schuerch, Janet R Sparrow, Stephen H Tsang, Rando Allikmets
BACKGROUND: Variation in the ABCA4 gene is causal for, or associated with, a wide range of phenotypes from early onset Mendelian retinal dystrophies to late-onset complex disorders such as age-related macular degeneration (AMD). Despite substantial progress in determining the causal genetic variation, even complete sequencing of the entire open reading frame and splice sites of ABCA4 identifies biallelic mutations in only 60%-70% of cases; 20%-25% remain with one mutation and no mutations are found in 10%-15% of cases with clinically confirmed ABCA4 disease...
April 26, 2017: Journal of Medical Genetics
https://www.readbyqxmd.com/read/28445755/mapping-ryanodine-binding-sites-in-the-pore-cavity-of-ryanodine-receptors
#4
Van A Ngo, Laura L Perissinotti, Williams Miranda, S R Wayne Chen, Sergei Y Noskov
Ryanodine (Ryd) irreversibly targets ryanodine receptors (RyRs), a family of intracellular calcium release channels essential for many cellular processes ranging from muscle contraction to learning and memory. Little is known of the atomistic details about how Ryd binds to RyRs. In this study, we used all-atom molecular dynamics simulations with both enhanced and bidirectional sampling to gain direct insights into how Ryd interacts with major residues in RyRs that were experimentally determined to be critical for its binding...
April 25, 2017: Biophysical Journal
https://www.readbyqxmd.com/read/28443131/a-bioinformatics-based-alternative-mrna-splicing-code-that-may-explain-some-disease-mutations-is-conserved-in-animals
#5
Wen Qu, Pablo Cingolani, Barry R Zeeberg, Douglas M Ruden
Deep sequencing of cDNAs made from spliced mRNAs indicates that most coding genes in many animals and plants have pre-mRNA transcripts that are alternatively spliced. In pre-mRNAs, in addition to invariant exons that are present in almost all mature mRNA products, there are at least 6 additional types of exons, such as exons from alternative promoters or with alternative polyA sites, mutually exclusive exons, skipped exons, or exons with alternative 5' or 3' splice sites. Our bioinformatics-based hypothesis is that, in analogy to the genetic code, there is an "alternative-splicing code" in introns and flanking exon sequences, analogous to the genetic code, that directs alternative splicing of many of the 36 types of introns...
2017: Frontiers in Genetics
https://www.readbyqxmd.com/read/28441772/structural-and-biochemical-properties-of-novel-self-cleaving-ribozymes
#6
REVIEW
Ki-Young Lee, Bong-Jin Lee
Fourteen well-defined ribozyme classes have been identified to date, among which nine are site-specific self-cleaving ribozymes. Very recently, small self-cleaving ribozymes have attracted renewed interest in their structure, biochemistry, and biological function since the discovery, during the last three years, of four novel ribozymes, termed twister, twister sister, pistol, and hatchet. In this review, we mainly address the structure, biochemistry, and catalytic mechanism of the novel ribozymes. They are characterized by distinct active site architectures and divergent, but similar, biochemical properties...
April 24, 2017: Molecules: a Journal of Synthetic Chemistry and Natural Product Chemistry
https://www.readbyqxmd.com/read/28441004/insights-into-the-mechanisms-by-which-clostridial-neurotoxins-discriminate-between-gangliosides
#7
Joshua R Burns, Gregory S Lambert, Michael R Baldwin
Botulinum neurotoxins (BoNTs) and Tetanus neurotoxin (TeNT) are the causative agents of the paralytic diseases botulism and tetanus, respectively. Entry of toxins into neurons is mediated through initial interactions with gangliosides, followed by binding to a protein co-receptor. Herein we aimed to understand the mechanism through which individual neurotoxins recognize the carbohydrate motif of gangliosides. Using cell-based and in vitro binding assays, in conjunction with structure-driven site directed mutagenesis, a conserved hydrophobic residue within the BoNTs that contributes to both affinity and specificity towards Sia5-containing gangliosides was identified...
April 25, 2017: Biochemistry
https://www.readbyqxmd.com/read/28440507/a-novel-splice-site-mutation-of-wrn-c-ivs28-2t-c-identified-in-a-consanguineous-family-with-werner-syndrome
#8
Pan-Feng Wu, Jie-Yuan Jin, Jing-Jing Li, Ji-Qiang He, Liang-Liang Fan, Min Jin, Hao Huang, Kun Xia, Ju-Yu Tang, Rong Xiang
Werner Syndrome (WS) is a rare, adult‑onset progeroid syndrome that is associated with multiple age‑associated complications and relatively short life expectancy. The characteristics of WS include a 'bird‑like' appearance, canities, cataracts and ulcerations around the ankles. In addition, certain patients develop hypogonadism with atrophic genitalia and infertility. The average life span of affected individuals is 54 years. Previous studies have demonstrated that mutations in the Werner syndrome RecQ like helicase gene (WRN) may contribute to WS...
April 12, 2017: Molecular Medicine Reports
https://www.readbyqxmd.com/read/28438176/the-pink1-p-i368n-mutation-affects-protein-stability-and-ubiquitin-kinase-activity
#9
Maya Ando, Fabienne C Fiesel, Roman Hudec, Thomas R Caulfield, Kotaro Ogaki, Paulina Górka-Skoczylas, Dariusz Koziorowski, Andrzej Friedman, Li Chen, Valina L Dawson, Ted M Dawson, Guojun Bu, Owen A Ross, Zbigniew K Wszolek, Wolfdieter Springer
BACKGROUND: Mutations in PINK1 and PARKIN are the most common causes of recessive early-onset Parkinson's disease (EOPD). Together, the mitochondrial ubiquitin (Ub) kinase PINK1 and the cytosolic E3 Ub ligase PARKIN direct a complex regulated, sequential mitochondrial quality control. Thereby, damaged mitochondria are identified and targeted to degradation in order to prevent their accumulation and eventually cell death. Homozygous or compound heterozygous loss of either gene function disrupts this protective pathway, though at different steps and by distinct mechanisms...
April 24, 2017: Molecular Neurodegeneration
https://www.readbyqxmd.com/read/28437635/three-cysteine-residues-of-slc52a1-a-receptor-for-the-porcine-endogenous-retrovirus-a-perv-a-play-a-critical-role-in-cell-surface-expression-and-infectivity
#10
Winston Colon-Moran, Takele Argaw, Carolyn A Wilson
Porcine endogenous retrovirus-A (PERV-A), a gammaretrovirus, infects human cells in vitro, thus raising the potential risk of cross-species transmission in xenotransplantation. Two members of the solute carrier family 52 (SLC52A1 and SLC52A2) are PERV-A receptors. Site-directed mutagenesis of the cDNA encoding SLC52A1 identified that only one of two putative glycosylation signals is occupied by glycans. In addition, we showed that glycosylation of SLC52A1 is not necessary for PERV-A receptor function. We also identified that at a minimum, three cysteine residues are sufficient for SLC52A1 cell surface expression...
April 20, 2017: Virology
https://www.readbyqxmd.com/read/28436954/optimized-expression-based-microdissection-of-formalin-fixed-lung-cancer-tissue
#11
Markus Grafen, Thurid R Hofmann, Andreas H Scheel, Julia Beck, Alexander Emmert, Stefan Küffer, Bernhard C Danner, Ekkehard Schütz, Reinhardt Büttner, Andreas Ostendorf, Philipp Ströbel, Hanibal Bohnenberger
Analysis of specific DNA alterations in precision medicine of tumors is crucially important for molecular targeted treatments. Lung cancer is a prototypic example and one of the leading causes of cancer-related deaths worldwide. One major technical problem of detecting DNA alterations in tissue samples is cellular heterogeneity, that is, mixture of tumor and normal cells. Microdissection is an important tool to enrich tumor cells from heterogeneous tissue samples. However, conventional laser capture microdissection has several disadvantages like user-dependent selection of regions of interest (ROI), high costs for dissection systems and long processing times...
April 24, 2017: Laboratory Investigation; a Journal of Technical Methods and Pathology
https://www.readbyqxmd.com/read/28436537/error-prone-replication-bypass-of-the-imidazole-ring-opened-formamidopyrimidine-deoxyguanosine-adduct
#12
Yan Sha, Irina G Minko, Chanchal K Malik, Carmelo J Rizzo, R Stephen Lloyd
Addition of hydroxyl radicals to the C8 position of 2'-deoxyguanosine generates an 8-hydroxyguanyl radical that can be converted into either 8-oxo-7,8-dihydro-2'-deoxyguanosine or N-(2-deoxy-d-pentofuranosyl)-N-(2,6-diamino-4-hydroxy-5-formamidopyrimidine) (Fapy-dG). The Fapy-dG adduct can adopt different conformations and in particular, can exist in an unnatural α anomeric configuration in addition to canonical β configuration. Previous studies reported that in 5'-TGN-3' sequences, Fapy-dG predominantly induced G → T transversions in both mammalian cells and Escherichia coli, suggesting that mutations could be formed either via insertion of a dA opposite the 5' dT due to primer/template misalignment or as result of direct miscoding...
April 24, 2017: Environmental and Molecular Mutagenesis
https://www.readbyqxmd.com/read/28433102/the-genetics-of-wilson-disease
#13
Irene J Chang, Si Houn Hahn
Wilson disease (WD) is an autosomal-recessive disorder of hepatocellular copper deposition caused by pathogenic variants in the copper-transporting gene, ATP7B. Early detection and treatment are critical to prevent lifelong neuropsychiatric, hepatic, and systemic disabilities. Due to the marked heterogeneity in age of onset and clinical presentation, the diagnosis of Wilson disease remains challenging to physicians today. Direct sequencing of the ATP7B gene is the most sensitive and widely used confirmatory testing method, and concurrent biochemical testing improves diagnostic accuracy...
2017: Handbook of Clinical Neurology
https://www.readbyqxmd.com/read/28432359/role-of-the-ion-channel-extracellular-collar-in-ampa-receptor-gating
#14
Maria V Yelshanskaya, Samaneh Mesbahi-Vasey, Maria G Kurnikova, Alexander I Sobolevsky
AMPA subtype ionotropic glutamate receptors mediate fast excitatory neurotransmission and are implicated in numerous neurological diseases. Ionic currents through AMPA receptor channels can be allosterically regulated via different sites on the receptor protein. We used site-directed mutagenesis and patch-clamp recordings to probe the ion channel extracellular collar, the binding region for noncompetitive allosteric inhibitors. We found position and substitution-dependent effects for introduced mutations at this region on AMPA receptor gating...
April 21, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28432356/a-role-of-human-rnase-p-subunits-rpp29-and-rpp21-in-homology-directed-repair-of-double-strand-breaks
#15
Enas R Abu-Zhayia, Hanan Khoury-Haddad, Noga Guttmann-Raviv, Raphael Serruya, Nayef Jarrous, Nabieh Ayoub
DNA damage response (DDR) is needed to repair damaged DNA for genomic integrity preservation. Defective DDR causes accumulation of deleterious mutations and DNA lesions that can lead to genomic instabilities and carcinogenesis. Identifying new players in the DDR, therefore, is essential to advance the understanding of the molecular mechanisms by which cells keep their genetic material intact. Here, we show that the core protein subunits Rpp29 and Rpp21 of human RNase P complex are implicated in DDR. We demonstrate that Rpp29 and Rpp21 depletion impairs double-strand break (DSB) repair by homology-directed repair (HDR), but has no deleterious effect on the integrity of non-homologous end joining...
April 21, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28432336/structure-function-analysis-of-the-dna-binding-domain-of-a-transmembrane-transcriptional-activator
#16
Andreas Schlundt, Sophie Buchner, Robert Janowski, Thomas Heydenreich, Ralf Heermann, Jürgen Lassak, Arie Geerlof, Ralf Stehle, Dierk Niessing, Kirsten Jung, Michael Sattler
The transmembrane DNA-binding protein CadC of E. coli, a representative of the ToxR-like receptor family, combines input and effector domains for signal sensing and transcriptional activation, respectively, in a single protein, thus representing one of the simplest signalling systems. At acidic pH in a lysine-rich environment, CadC activates the transcription of the cadBA operon through recruitment of the RNA polymerase (RNAP) to the two cadBA promoter sites, Cad1 and Cad2, which are directly bound by CadC...
April 21, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28432123/an-amphipathic-%C3%AE-helix-directs-palmitoylation-of-the-large-intracellular-loop-of-the-sodium-calcium-exchanger
#17
Fiona Plain, Samitha Dilini Congreve, Rachel Sue Zhen Yee, Jennifer Kennedy, Jacqueline Howie, Chien-Wen Kuo, Niall Fraser, William Fuller
The electrogenic sodium/calcium exchanger (NCX) mediates bidirectional calcium transport controlled by the transmembrane sodium gradient. NCX inactivation occurs in the absence of phosphatidylinositol 4,5-bisphosphate (PIP2), and is facilitated by palmitoylation of a single cysteine at position 739 within NCX's large intracellular loop. The aim of this investigation was to identify the structural determinants of NCX1 palmitoylation. Full-length NCX1 (FL-NCX1) and a YFP fusion protein of the NCX1 large intracellular loop (YFP-NCX1) were expressed in HEK cells...
April 21, 2017: Journal of Biological Chemistry
https://www.readbyqxmd.com/read/28431042/cis-regulatory-divergence-in-gene-expression-between-two-thermally-divergent-yeast-species
#18
Xueying C Li, Justin C Fay
Gene regulation is a ubiquitous mechanism by which organisms respond to their environment. While organisms are often found to be adapted to the environments they experience, the role of gene regulation in environmental adaptation is not often known. In this study, we examine divergence in cis-regulatory effects between two Saccharomyces species, S. cerevisiae and S. uvarum, that have substantially diverged in their thermal growth profile. We measured allele specific expression (ASE) in the species' hybrid at three temperatures, the highest of which is lethal to S...
April 14, 2017: Genome Biology and Evolution
https://www.readbyqxmd.com/read/28424158/generation-of-a-multipurpose-prdm16-allele-by-targeted-trapping
#19
A Strassman, F Schnütgen, Q Dai, J C Jones, A C Gomez, L Pitstick, N E Holton, R Moskal, E R Leslie, H von Melchner, D R Beier, B C Bjork
Gene trap mutagenesis is a powerful tool to create loss-of-function mutations in mice and other model organisms. Modifications of traditional gene trap cassettes, including addition of conditional features in the form of Flip-excision (FlEx) arrays enabling directional gene trap cassette inversions by Cre and Flpe site-specific recombinases, greatly enhanced their experimental potential. By taking advantage of these conditional gene trap cassettes, we developed a generic strategy for generating conditional mutations and validated this strategy in mice carrying a multipurpose allele of the Prdm16 transcription factor gene...
April 19, 2017: Disease Models & Mechanisms
https://www.readbyqxmd.com/read/28423625/inhibition-activity-of-a-disulfide-stabilized-diabody-against-basic-fibroblast-growth-factor-in-lung-cancer
#20
Yaxiong Cai, Shuange Yao, Jiangchuan Zhong, Jinxia Zhang, Haowu Jiang, Yanrui Deng, Ning Deng
The over-expression of basic fibroblast growth factor (bFGF) plays a crucial role in the development, invasion and metastasis of lung cancer. Therefore, neutralizing antibodies against bFGF may inhibit the growth of lung cancer. In this study, a Disulfide-stabilized diabody (ds-Diabody) against bFGF was constructed by site-directed mutation and overlap extension PCR (SOE-PCR) at the position of VH44 and VL100 in the scFv. The ds-Diabody was constructed and expressed in Pichia pastoris. We found that the ds-Diabody against bFGF could efficiently suppress the proliferation, migration and invasion of human lung cancer A549 cells in vitro...
March 21, 2017: Oncotarget
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