Escherichia coli polynucleotide phosphorylase (PNPase) is generally believed to be involved in mRNA degradation via its 3$\sp\prime$ to 5$\sp\prime$ exoribonuclease activity. However, there are many unexplained observations related to the physiological role of this enzyme. The projects conducted in this thesis are aimed at understanding more about the in vivo function of PNPase. Two approaches were employed to try to achieve this goal. First, a biochemical approach involving the purification of PNPase associated proteins was used. Specifically, the previously documented $\beta$ subunit present in the pentameric form of PNPase was examined as an initial step toward understanding the mechanism and possibly the regulation of PNPase catalysis. Second, a genetic approach was undertaken which focused on characterization of mutant strains lacking PNPase in combination with other known enzymes involved in RNA metabolism, in particular, tRNA nucleotidyltransferase, poly(A) polymerase and RNase PH.^ From the
TY - JOUR. T1 - Endogenous superoxide dismutase levels regulate iron-dependent hydroxyl radical formation in Escherichia coli exposed to hydrogen peroxide. AU - Mccormick, Michael L.. AU - Buettner, Garry R.. AU - Britigan, Bradley E.. PY - 1998/2/1. Y1 - 1998/2/1. N2 - Aerobic organisms contain antioxidant enzymes, such as superoxide dismutase (SOD) and catalase, to protect them from both direct and indirect effects of reactive oxygen species, such as O2- and H2O2. Previous work by others has shown that Escherichia coli mutants lacking SOD not only are more susceptible to DNA damage and killing by H2O2 but also contain larger pools of intracellular free iron. The present study investigated if SOD- deficient E. coli cells are exposed to increased levels of hydroxyl radical (OH) as a consequence of the reaction of H2O2 with this increased iron pool. When the parental E. coli strain AB1157 was exposed to H2O2 in the presence of an α-(4-pyridyl-1-oxide)-N-tert-butyl-nitrone (4-POBN)-ethanol ...
A polynucleotide molecule is a biopolymer composed of 13 or more nucleotide monomers covalently bonded in a chain. DNA (deoxyribonucleic acid) and RNA (ribonucleic acid) are examples of polynucleotides with distinct biological function. The prefix poly comes from the ancient Greek πολυς (polys, many). DNA consists of two chains of polynucleotides, with each chain in the form of a helical spiral. Although DNA and RNA do not generally occur in the same polynucleotide, the four species of nucleotides may occur in any order in the chain. The sequence of DNA or RNA species for a given polynucleotide is the main factor determining its function in a living organism or a scientific experiment. Polynucleotides occur naturally in all living organisms. The genome of an organism consists of complementary pairs of enormously long polynucleotides wound around each other in the form of a double helix. Polynucleotides have a variety of other roles in organisms. Polynucleotides are used in biochemical ...
SUMMARY: Polynucleotide phosphorylase has been purified from the cyanobacterium Nostoc sp. MAC. The enzyme requires a divalent cation such as Mg2+, has a pH optimum of 10·5 and catalyses the polymerization of ADP into polynucleotide in a primer-independent reaction at a rate of 2 μmol min-1 (mg protein)-1. It has an apparent native M r of 215000-240000. Non-denaturing polyacrylamide activity gels reveal a heterogeneous pattern of active bands similar to those previously observed with the corresponding enzyme from Micrococcus luteus, while SDS-denaturing activity gels reveal a single band of activity of M r 91000 in both crude extracts and the most highly purified fraction. Activity has also been demonstrated in a cetyltrimethyl-ammonium bromide non-denaturing activity gel. By analogy with other known polynucleotide phosphorylases, the Nostoc enzyme is probably a trimer of the 91000-M r subunit.
poly(2-chloro-2-deoxyadenylic acid): used as messenger RNAs in protein synthesizing systems in vitro; cpd not in Chemline 7/19/83
METHODS AND ARRAYS FOR DNA SEQUENCING - A method of sequencing a first polynucleotide strand having a first polynucleotide sequence, the first polynucleotide strand resembling a second polynucleotide strand having a known second polynucleotide sequence, the method employing a data set which, for one or more fragment(s) of the second polynucleotide sequence, contains: for each position along each said fragment: (i) first probe data describing the hybridization intensity of the first polynucleotide strand with a respective first probe designed to bind to a portion of the second polynucleotide strand centered at said position; and (ii) second probe data describing the respective hybridization intensities of the first polynucleotide strand with each of a set of second probes, each said second probe being designed to bind with a respective mutation of the corresponding portion of the second polynucleotide sequence which is formed by mutating the corresponding portion of the second polynucleotide ...
A numerical model has been developed to estimate the role of hydroxyl radicals in the competition of organic compounds during electrolysis using a boron-doped diamond (BDD) anode. It is well established that hydroxyl radicals are generated during water discharge. These radicals are free at the surface and act locally, their sphere of activity was estimated by the model to be 20 nm maximum from the anode. The implementation of the model taking account the presence of one and then two organic compounds in the solution can be used to predict the variation of the concentration of each organic with time during the electrolysis and their space profile in the electrochemical reactor too without using adjustable parameters. The role of hydroxyl radicals was highlighted in the competitive reaction with organics: the electrolysis of a solution containing an equimolar mixture of formic acid (FA) and maleic acid (MA) shows that FA oxidation only began when the MA had completely disappeared. The numerical ...
In the hydrothermal crystallization of zeolites from basic media, hydroxide ions (OH-) catalyze the depolymerization of the aluminosilicate gel by breaking the Si,Al-O-Si,Al bonds and catalyze the polymerization of the aluminosilicate anions around the hydrated cation species by remaking the Si,Al-O-Si,Al bonds. We report that hydroxyl free radicals (•OH) are involved in the zeolite crystallization under hydrothermal conditions. The crystallization processes of zeolites-such as Na-A, Na-X, NaZ-21, and silicalite-1-can be accelerated with hydroxyl free radicals generated by ultraviolet irradiation or Fentons reagent. ...
1. A new method has been developed for the preparation in good yield of highly purified Azotobacter vinelandii polynucleotide phosphorylase in its reduced form. 2. Aging or digestion with trypsin causes the enzyme to develop a primer requirement that is not eliminated by β-mercaptoethanol. 3. The development of a primer requirement is accompanied by marked changes of the electrophoretic mobility of the enzyme in polyacrylamide gels. 4. The enzyme is inactivated by aerial oxidation or thiol-specific reagents. The lost activity is restored by β-mercaptoethanol, but not by oligonucleotide primers.. ...
0111] In a preferred embodiment, the invention provides for a method of in vitro recombination comprising: [0112] (1) obtaining modified polynucleotide fragments according any one of the method described in any one of claims 1 to 5; [0113] (2) screening some or all of said modified polynucleotides to determine which polynucleotide or polynucleotides encode a protein or proteins of interest; [0114] (3) digesting said modified polynucleotides encoding a protein or proteins of interest with restriction enzymes to form fragments with single-stranded overhangs consisting of three nucleotide residues or of nucleotide residues in multiples of three; [0115] (4) modifying the obtained polynucleotide fragments of step (3) by removing and/or filling in the single-stranded overhangs to obtain new modified fragments by [0116] (i) removing, in multiples of three, all of the nucleotide residues of said overhanging end of one or more polynucleotide fragments; or [0117] (ii) extending the single strand of the ...
DNA ligase activity is strongly accelerated in the presence of high concentrations of non-specific polymers, with an accompanying change in product distribution characteristics. The rate of blunt-end ligation of DNA substrate by T4 DNA ligase is particularly affected, with a product shift from closed circular species to linear oligomers. The method provides a way to increase the rate of enzymatic DNA ligation and the size of the linear products. It may be useful when preparing large amounts of polymers by ligation of oligomers or when ligating amounts of DNA or deoxyribooligomers so low in concentration that a reduced yield would otherwise result.
DNA bases can be modified by endogenous agents (e.g. oxidized by products of respiration and photosynthesis or methylated by gene silencing processes) as well as by environmental agents (e.g. oxidized by UV light). In the process of removing modified bases, a 3-phosphate group is sometimes left in the resulting gap, and has to be removed since it blocks the incorporation of a new nucleotide by DNA polymerase. The aim of this thesis was the characterization of AtZDP, a plant enzyme with a DNA 3-phosphatase activity.. By homologous modeling, the existence of four domains was predicted in AtZDP, three independent zinc-finger and one DNA 3-phosphatase domains. AtZDP was found to be localized in the nucleus by bimolecular fluorescence complementation. Western blotting analysis showed that the enzyme was ubiquitously expressed in plant tissues.. AtZDP was found in a 600,000 molecular-weight protein complex by gel chromatography and glycerol gradient sedimentation centrifugation. The fractions ...
1. Remove the 10X T4 DNA Ligase Reaction Buffer* from the freezer to thaw. You can remove the T4 DNA Ligase enzyme from the freezer at this point but leave the ligase in a cold box to keep it close to -20○ C. Thawing is fast if the buffer tube is immersed in room temperature water. Once thawed, agitate the 10X T4 DNA Ligase Reaction Buffer until all precipitate goes into solution. 2. Add 11ul of H2O to a 200ul PCR tube. 3. Add 2ul from each of the digest to the tube.** 4. Add 2ul of 10X T4 DNA Ligase Reaction Buffer to the tube. 5. Add 1ul of the T4 DNA Ligase to the tube. 6. The total volume in each tube should now be 20ul. Ensue the ligation is well mixed by flicking the tube. You can spin the tube in a microcentrifuge for a few seconds to collect the liquid in the bottom of the tube again. *Repeated freeze-thaw cycles of the buffer can degrade the ATP in the buffer thereby making the ligation reaction less efficient. It is wise to aliquot the buffer into 10ul aliquots prior to freezing ...
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Last week, OCHIN, Inc. was awarded nearly \$7 million by the Patient-Centered Outcomes Research Institute to establish itself as part of PCORIs new Patient-Centered National Clinical Research Network. Jennifer DeVoe, M.D., DPhil, OCHINs research director and an associate professor in OHSUs Department of Family Medicine, is principal investigator on the project.. "By collaborating across the nation we hope to provide results that will inform policy and improve patient and population health outcomes," Dr. DeVoe said. "This opportunity with PCORI comes at a time when our nation is working to transform healthcare and needs the best evidence possible to ensure that care is more patient-centered. Im so proud to be serving on this team.". Read more.. ...
International Paint, Devoe High Performance Coatings launch integration project AkzoNobels International Paint Protective Coatings and Devoe High Performance Coatings have launched a two-year integration effort designed to expand its po
Definition of Oxygen-Derived Free Radical in the Legal Dictionary - by Free online English dictionary and encyclopedia. What is Oxygen-Derived Free Radical? Meaning of Oxygen-Derived Free Radical as a legal term. What does Oxygen-Derived Free Radical mean in law?
E. coli DNA Ligase catalyzes the formation of phosphodiester bonds between double-stranded DNA fragments containing juxtaposed 5-phosphate termini and 3-hydroxyl termini in the presence of the NAD cofactor. Both T4 DNA Ligase and E.coli DNA Ligase are used in various gene cloning experiments; however, unlike T4 DNA Ligase, E. coli DNA Ligase can only catalyze ligation of cohesive end-containing DNA fragments via standard reaction conditions. E. coli DNA Ligase is supplied in a buffer containing 10 mM potassium phosphate (pH 7.5), 50 mM KCl, 1 mM DTT, 1 mM EDTA and 50% glycerol.. ...
Abstract: Reactive oxygen species (ROS) such as hydrogen peroxide (H2O2) can produce hydroxyl radicals (OHo) that damage DNA. Oxidative damage affects the function or reproduction of cells leading to diseases such as cancer and neurodegenerative disorders. Living organisms have developed defense mechanisms such as antioxidants that can prevent or repair oxidation. Peroxiredoxins (prx), ubiquitous proteins that are found in organisms ranging from bacteria to humans, can act as antioxidants reducing H2O2 to H2O. In eukaryotic cells, prx are also regulators of H2O2-mediating signaling. In the presence of high levels of H2O2, peroxiredoxin becomes overoxidized, forming sulfinic acid. The overoxidized prx can no longer reduce H2O2 to H2O, allowing H2O2 to participate in signaling. The formation of sulfinic acid was believed to be irreversible; however, recent studies have shown that a yeast protein called sulfiredoxin (srx) can reduce the cysteine-sulfinic acid of yeast peroxiredoxin TsaI back to its ...
299057642 - EP 0785941 B1 2000-04-19 - PORPHYRIN LABELING OF POLYNUCLEOTIDES - [origin: WO9611937A1] The invention provides various porphyrin labeled compounds and reagents for labeling molecules with porphyrins. The porphyrin labeled compounds include porphyrin labeled nucleosides, nucleotides, polynucleotides and the like. The porphyrin labeled compounds of the invention are designed so that the porphyrin moiety on the labeled compound may be detected on the basis of a reaction product produced by the porphyrin catalyzed oxidation of a porphyrin detection reagent. The oxidation of the porphyrin detection reagent may result in the formation of light, i.e., a chemiluminescent reaction, or a colored compound, i.e., a colorimetric reaction. The compounds of the invention contain the general structure of: porphyrin-linker-base-sugar-(phos)n-OH. Another aspect of the invention is to provide methods of labeling and detecting polynucleotides by incorporating a porphyrin-labeled nucleotide into a
Involved in mRNA degradation. Catalyzes the phosphorolysis of single-stranded polyribonucleotides processively in the 3- to 5-direction.
In this study, we aim at evaluation the role of the Asp148Glu (rs1130409) variant at apurinic/apyrimidinic endonuclease (APE) gene in renal cell carcinoma (RCC) risk and the contribution of different genotypes to its transcriptional mRNA levels. In the case-control study, 92 RCC patients and 580 cancer-free patients matched by age and gender were recruited. The apurinic/APE genotyping work was con...
Mentors: Anderson, Hilt. Iron oxide nanoparticles have the ability to enhance the production of reactive oxygen species (ROS) within cells through catalyzing the Haber-Weiss reaction (Fenton chemistry) which produces the hydroxyl radical. Cancer cells are more susceptible to oxidative insults compared to normal cells due to fast cell proliferation and metabolism so additional ROS stress induced by exogenous agents can overwhelm the relatively low antioxidant capacity and disrupt the redox homeostasis inside cancer cells leading to selective tumor cell toxicity. Iron oxide nanoparticles have been previously studied due to their multitude of biological applications, inherent biocompatibility, magnetic properties, and lack of protein adsorption after proper coating. Therefore, iron oxide nanoparticles coated with dextran will be used to improve the efficacy of radiation by enhancing the intracellular ROS production.. Due to the half-life of the hydroxyl radical being on the order of a nanosecond, ...
Generally, free radicals react with other molecules, damaging them. However some processes such as the killing of bacteria by neutrophil granulocytes is beneficial. The most important oxygen-based free radicals are superoxide and the hydroxyl radical, produced from the reduction of molecular oxygen. These free radicals are extremely reactive and are often associated with cell damage, mutations, and even malignancies. Symptoms of aging such as atherosclerosis is thought to be due to oxidation by free radicals. The free radical theory of aging postulates that free radical damage is the primary mechanism of aging. Free radicals are also associated with liver damage due to alcohol and the development of emphysema from ciagrette smoking. The body has developed a number of mechanisms to minimize free radical damage and even repair damage. Enzymes such as superoxide dismutase, catalase, [[glutathione peroxidase]], and glutathione reductase, as well as antioxidants such as vitamin A, vitamin C, vitamin ...
0007] Various embodiments include one or more of the following features: [0008] a ligase reaction buffer contains a small molecule enhancer having a concentration of 1%-50% v/v that is capable of enhancing, by at least 25%, intramolecular or intermolecular ligation of a polynucleotide fragment or fragments than would otherwise be obtained in the absence of the enhancer; [0009] the one or more small molecule ligation enhancers are selected from an optionally substituted straight or branched chain diol or polyol containing 2 to 20 carbons, alcohols, zwitterionic compounds and polar aprotic molecules; [0010] the optionally substituted straight or branched diol or polyol is selected from the group consisting of 1,2-ethylene glycol, 1,2-propanediol (1,2-PrD), 1,3-propanediol (1,3-PrD), glycerol, pentaerythritol, sorbitol, diethylene glycol, dipropyleneglycol, neopentyl glycol, 2-methyl-1,3-propanediol, 2,2-dimethyl-1,3-propanediol, 1,3-butanediol, and 1,4-butanediol 1,5-pentanediol, 1,6-hexanediol, ...
TY - JOUR. T1 - An essential function for the phosphate-dependent exoribonucleases RNase PH and polynucleotide phosphorylase. AU - Zhou, Zhihua. AU - Deutscher, Murray P.. PY - 1997/7. Y1 - 1997/7. N2 - Escherichia coli cells lacking both polynucleotide phosphorylase (PNPase) and RNase PH, the only known P1-dependent exoribonucleases, were previously shown to grow slowly at 37°C and to display a dramatically reduced level of tRNA(Tryrsu3+) suppressor activity. Here we show that the RNase PH-negative, PNP-negative double-mutant strain actually displays a reversible cold-sensitive phenotype and that tRNA biosynthesis is normal. In contrast, ribosome structure and function are severely affected, particularly at lower temperatures. At 31°C, the amount of 50S subunit is dramatically reduced and 23S rRNA is degraded. Moreover, cells that had been incubated at 42°C immediately cease growing and synthesizing protein upon a shift to 31°C, suggesting that the ribosomes synthesized at the higher ...
TY - JOUR. T1 - Kinetic and stoichiometric assessment of the antioxidant activity of flavonoids by electron spin resonance spectroscopy. AU - McPhail, D B AU - Hartley, R C AU - Gardner, P T AU - Duthie, G G. PY - 2003/3/12. Y1 - 2003/3/12. N2 - dThere is current interest in the use of naturally occurring flavonoids as antioxidants for the preservation of foods and the prevention of diseases such as atherosclerosis and cancers. To establish the molecular characteristics required for maximum antioxidant activity, electron spin resonance (ESR) spectroscopy has been used to determine the stoichiometry and kinetics of the hydrogen-donating ability of 15 flavonoids and D-alpha-tocopherol to galvinoxyl, a resonance-stabilized, sterically protected aryloxyl radical. The second-order reaction rates, which will be governed by O-H bond dissociation energies, were myricetin , morin , quercetin , fisetin similar to catechin , kaempferol similar to luteolin , rutin , D-alpha-tocopherol , taxifolin , ...
Polynucleotide Adenylyltransferase: An enzyme that catalyzes the synthesis of polyadenylic acid from ATP. May be due to the action of RNA polymerase (EC 2.7.7.6) or polynucleotide adenylyltransferase (EC 2.7.7.19). EC 2.7.7.19.
Hydrogen peroxide and peroxynitrite induce relaxations via ATP-sensitive K+ channels, indicating that oxygen-derived free radicals may activate these channels. Levels of free radicals are increased throughout the arterial wall in animal models of atherosclerosis, and therefore, vasorelaxation via ATP-sensitive K+ channels may be augmented in chronic hypertension. The present study was designed to determine whether relaxations to an ATP-sensitive K+ channel opener, levcromakalim, are increased in the aorta from spontaneously hypertensive rats (SHR) and whether free radical scavengers reduce these relaxations. Rings of aortas without endothelium taken from age-matched Wistar-Kyoto rats (WKY) and SHR were suspended for isometric force recording. Relaxations to levcromakalim (10(-8) to 10(-5) M), which are abolished by glibenclamide (10(-5) M), were augmented in the aorta from SHR, compared to those in the aorta from WKY. In the aorta from SHR, catalase (1200 U/ml), but neither superoxide dismutase (150 U
Poly(ADP-ribose) polymerase (PARP) is a critical DNA repair enzyme involved in DNA single-strand break repair via the base excision repair pathway. PARP inhibitors have been shown to sensitize tumors to DNA-damaging agents and to also selectively kill homologous recombination repair-defective cancers, such as those arising in BRCA1 and BRCA2 mutation carriers. Recent proof-of-concept clinical studies have demonstrated the safety and substantial antitumor activity of the PARP inhibitor, olaparib in BRCA1/2 mutation carriers, highlighting the wide therapeutic window that can be achieved with this synthetic lethal strategy. Likewise, the PARP inhibitor, BSI-201, in combination with carboplatin and gemcitabine have produced promising results in "triple-negative" breast cancers. There are also currently numerous other PARP inhibitors in clinical development. The potential broader therapeutic application of these approaches to a wide range of sporadic tumors harboring specific defects in the ...
The cold acclimatization response in many bacterial species is a tightly regulated process, which ensures the correct folding of macromolecules. In enterobacteria, this response is in part dependent on polynucleotide phosphorylase, which is encoded by the gene pnp. Based on transcriptional analysis of the pnp locus of Salmonella enterica serovar Typhimurium, we show that pnp and the adjacent membrane lipoprotein nlpI gene form an operon with both genes contributing independently to the cold acclimatization response at 15 °C. Our findings thereby define a new role for NlpI in bacterial cold acclimatization.. ...
The accumulation of unfolded proteins in the endoplasmic reticulum (ER) triggers an intracellular signaling pathway, the unfolded protein response (UPR), that leads to increased transcription of genes encoding ER-resident proteins. Transcriptional activation is mediated by a dedicated transcription factor, Hac1p, whose activity is controlled by regulated splicing of its mRNA. We have identified a mutation in tRNA ligase that disrupts the UPR in the yeast Saccharomyces cerevisiae. In this mutant, splicing of HAC1 mRNA, but not tRNA, is blocked. In contrast, HAC1 mRNA splicing is not impaired in cells that are blocked in spliceosome-mediated mRNA splicing. Furthermore, the splice junctions of HAC1 mRNA do not conform to the consensus sequences of other yeast pre-mRNAs. Our results suggest that the regulated splicing of HAC1 mRNA occurs by a novel pathway, involving tRNA ligase and bypassing the spliceosome.. ...
Polyadenylated (poly(A)+) RNA molecules have been isolated from Methanococcus vannielii. Approximately 16% of the label in RNA isolated from cultures allowed to incorporate (3)Huridine for 3 min at 37C was poly(A)+ RNA. Electrophoretic separation of poly(A)+ RNA molecules showed a heterogeneous population with mobilities indicative of sizes ranging from 900 to 3,000 bases in length. Polyadenylate (poly(A)) tracts were isolated by digestion with RNase A and RNase T1 after 3 end labeling of the poly(A)+ RNA with RNA ligase. The radioactively labeled poly(A) oligonucleotides were shown by electrophoresis through DNA sequencing gels to average 10 bases in length, with major components of 5, 9, 10, 11, and 12 bases. Poly(A)+ RNA molecules from M. vannielii were labeled at their 5 termini with T4 polynucleotide kinase after dephosphorylation with calf intestine alkaline phosphatase. Pretreatment of the RNA molecules with tobacco acid pyrophosphatase did not increase the amount of phosphate ...
The present invention relates to isolated polypeptides having xylanase activity and polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.
Three IgM monoclonal anti-DNA antibodies were produced by hybridoma techniques from an MRL-lpr/lpr mouse using denatured DNA (dDNA) as the selection antigen. All three antibodies also bound poly(dT), poly(rA), and the single-stranded random copolymer poly(dI,dT), and each antibody displayed a unique preference for a limited array of other ribo- and deoxyribopolynucleotides based on direct binding as well as inhibition studies. Inability to identify a common primary structure in the polynucleotides reactive with each antibody suggested that higher ordered structures may be important. This notion was supported by the finding that oligomers of thymidine of 25-30 nucleotides or less were ineffective in blocking antibody binding to dDNA or poly(dT). However, deliberate destabilization of putative secondary structures by decreasing counterion concentration and increasing temperature had little effect on antibody binding to poly(dT). Since the antigenic polynucleotides in general contain little known ...
Dephosphorylates several organic phosphate monoesters including 3- and 5-nucleotides, 2-deoxy-5-nucleotides, pNPP, phenyl phosphate, glycerol 2-phosphate, ribose 5-phosphate, O-phospho-L-amino acids and phytic acid, showing the highest activity with aryl phosphoesters (pNPP, phenyl phosphate and O-phospho-L-tyrosine), and to a lesser extent with 3- and 5-nucleotides. No activity toward ATP, phosphodiesters, glycerol-1-phosphate, glucose 1-phosphate, glucose 6-phosphate, NADP, GTP or 3,5-cAMP, ADP or ATP. Also has a phosphotransferase activity catalyzing the transfer of low-energy phosphate groups from organic phosphate monoesters to free hydroxyl groups of various organic compounds. Capable of transferring phosphate from either pNPP or UMP to adenosine or uridine. Does not exhibit nucleotide phosphomutase activity.
DNA repair enzyme that can remove a variety of covalent adducts from DNA through hydrolysis of a 3-phosphodiester bond, giving rise to DNA with a free 3 phosphate. Catalyzes the hydrolysis of dead-end complexes between DNA and the topoisomerase I active site tyrosine residue. Hydrolyzes 3-phosphoglycolates on protruding 3 ends on DNA double-strand breaks due to DNA damage by radiation and free radicals. Acts on blunt-ended double-strand DNA breaks and on single-stranded DNA. Has low 3exonuclease activity and can remove a single nucleoside from the 3end of DNA and RNA molecules with 3hydroxyl groups. Has no exonuclease activity towards DNA or RNA with a 3phosphate ...
291459338 - EP 1254270 A1 2002-11-06 - METHODS AND MATERIALS RELATING TO CUB DOMAIN POLYPEPTIDES AND POLYNUCLEOTIDES - [origin: WO0157267A1] The invention provides novel polynucleotides and polypeptides encoded by such polynucleotides and mutants or variants thereof that correspond to a novel human secreted CUB domain polypeptide. These polynucleotides comprise nucleic acid sequences isolated from cDNA library from testis (Hyseq clone identification numbers 2924342 (SEQ ID NO: 1)). Other aspects of the invention include vectors containing processes for producing novel humain secreted CUB domain polypeptides, and antibodies specific for such polypeptides.[origin: WO0157267A1] The invention provides novel polynucleotides and polypeptides encoded by such polynucleotides and mutants or variants thereof that correspond to a novel human secreted CUB domain polypeptide. These polynucleotides comprise nucleic acid sequences isolated from cDNA library from testis (Hyseq clone identification numbers 2924342 (SEQ
We have characterized homologous recombination between linear DNA and the bacterial chromosome that depends on λ recombination functions, involves very short homologies, and is very efficient. We examined several parameters to establish a maximal efficiency for phage-mediated recombination with short homologies. Maximal recombination levels are achieved with induction times from 7.5-17.5 min at 42°C, and a homology segment of 40-50 bp. Recombination saturates at a linear DNA substrate concentration of about 300 molecules per cell.. The fact that 30- to 50-bp homologies are able to recombine in vivo opens a vast array of new possibilities for generating recombinant DNA. Several steps normally involved in generating recombinant DNA molecules are eliminated. Restriction enzyme digests are not required to generate DNA fragments, and DNA ligase reactions are not required to join different DNA fragments at novel junctions. PCR amplification followed by electroporation of the linear DNA into cells is ...
Prokayrotic FAB I polypeptides and DNA (RNA) encoding such FAB I and a procedure for producing such polypeptides by recombinant techniques is disclosed. Also disclosed are methods for utilizing such FAB I for the treatment of infection, such as bacterial infections. Antagonists against such FAB I and their use as a therapeutic to treat infections, such as staphylococcal infections are also disclosed. Also disclosed are diagnostic assays for detecting diseases related to the presence of FAB I nucleic acid sequences and the polypeptides in a host. Also disclosed are diagnostic assays for detecting polynucleotides encoding FAB I and for detecting the polypeptide in a host.
Biopolymers are easily denatured by heating, a change in pH or chemical substances when they are immobilized on a substrate. To prevent denaturation of biopolymers, we developed a method to trap a polynucleotide on a substrate by hydrogen bonding using silica particles with surfaces modified by aminoalkyl chains ([A-AM silane]/SiO2). [A-AM silane]/SiO2 was synthesized by silane coupling reaction of N-2-(aminoethyl)-3-aminopropyltrimethoxysilane (A-AM silane) with SiO2 particles with a diameter of 5 μm at 100 °C for 20 min. The surface chemical structure of [A-AM silane]/SiO2 was characterized by Fourier transform infrared spectroscopy and molecular orbital calculations. The surface of the silica particles was modified with A-AM silane and primary amine groups were formed. [A-AM silane]/SiO2 was trapped with single-stranded nucleic acids [(Poly-X; X = A (adenine), G (guanine) and C (cytosine)] in PBS solution at 37 °C for 1 h. The single-stranded nucleic acids were trapped on the surface of the [A-AM
Disclosed are devices for amplifying a preselected polynucleotide in a sample by conducting a polynucleotide polymerization reaction. The devices comprise a substrate microfabricated to define a sample inlet port and a mesoscale flow system, which extends from the inlet port. The mesoscale flow system includes a polynucleotide polymerization reaction chamber in fluid communication with the inlet port which is provided with reagents required for polymerization and amplification of a preselected polynucleotide. In one embodiment the devices may be utilized to implement a polymerase chain reaction (PCR) in the reaction chamber (PCR chamber). The PCR chamber is provided with the sample polynucleotide, polymerase, nucleoside triphosphates, primers and other reagents required for the polymerase chain reaction, and the device is provided with a device for thermally controlling the temperature of the contents of the reaction chamber at a temperature controlled to dehybridize double stranded polynucleotide, to
DNA ligase IV functions in DNA non-homologous end-joining, in V(D)J recombination, and during brain development. We previously reported a homozygous mutation (R278H) in DNA ligase IV in a developmentally normal leukemia patient who overresponded to radiotherapy. The impact of this hypomorphic mutation has been evaluated using cellular, biochemical, and structural approaches. Structural modeling using T7 DNA ligase predicts that the activity and conformational stability of the protein is likely to be impaired. We show that wild type DNA ligase IV-Xrcc4 is an efficient double-stranded ligase with distinct optimal requirements for adenylate complex formation versus rejoining. The mutation impairs the formation of an adenylate complex as well as reducing the rejoining activity. Additionally, it imparts temperature-sensitive activity to the protein consistent with the predictions of the structural modeling. At the cellular level, the mutation confers a unique V(D)J recombination phenotype affecting ...
Substitution Mutations: In substitution mutations, a nitrogenous base of a triplet codon of DNA is replaced by another nitrogen base or some derivative of the nitrogen base, changing the codon. The altered codon codes for a different amino acid substitution.The substitution mutations are of two types: 1.Transitions: It is the replacement of one purine in a polynucleotide chain by another purine(A by G or G by A) or one pyrimidine by another pyrimidine(T by C or C by T) 2.Transversions:A base pair substitution involving the substitution of a purine by pyrimidine or pyrimidine by a purine is called transversion. ...
The present invention contemplates chromophore-containing polynucleotides having at least two donor chromophores operatively linked to the polynucleotide by linker arms, such that the chromophores are positioned by linkage along the length of the polynucleotide at a donor-donor transfer distance, and at least one fluorescing acceptor chromophore operatively linked to the polynucleotide by a linker arm, such that the fluorescing acceptor chromophore is positioned by linkage at a donor-acceptor transfer distance from at least one of the donor chromophores, to form a photonic structure for collecting photonic energy and transferring the energy to an acceptor chromophore, and methods using the photonic structures.
The ligation reaction is made up of cDNAs and my vector. I am interested in obtaining more sequence information from only a single transcript and I have designed primers to do so once I have the transcript cloned. I am wondering if i can amplify directly from my ligation reaction. It would be so much simpler this way. I am worried that i wont have enough starting material though. any thoughts?. ...
The present invention provides novel polynucleotides encoding HGPRBMY23 polypeptides, fragments and homologues thereof. Also provided are vectors, host cells, antibodies, and recombinant and synthetic methods for producing said polypeptides. The invention further relates to diagnostic and therapeutic methods for applying these novel HGPRBMY23 polypeptides to the diagnosis, treatment, and/or prevention of various diseases and/or disorders related to these polypeptides, particularly renal diseases and/or disorders, colon cancer, breast cancer, and diseases and disorders related to aberrant NFKB modulation. The invention further relates to screening methods for identifying agonists and antagonists of the polynucleotides and polypeptides of the present invention.
The present invention relates to isolated polypeptides having cellulolytic enhancing activity and isolated polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.
Wnt-3 polypeptides and polynucleotides and methods for producing such polypeptides by recombinant techniques are disclosed. Also disclosed are methods for utilizing Wnt-3 polypeptides and polynucleotides in therapy, and diagnostic assays for such.
Dinucleoside phosphorodithioate phosphoramidite precursors useful in the synthesis of oligonucleotide phosphorodithioate which can be used as antisense inhibitors of translation.