Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “POLYNUCLEOTIDES”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 163 records · Page 9Linked to original sources
PubMed1996

Polynucleotide: adenosine glycosidase activity of saporin-L1: effect on DNA, RNA and poly(A).

The ribosome-inactivating proteins (RIPs) are a family of plant enzymes for which a unique activity has been determined: rRNA N-glycosidase, which removes adenine at a specific universally conserved position (A4324 in the case of rat ribosomes). Here we report that saporin-L1, a RIP from the leaves of Saponaria officinalis, recognizes other substrates, including RNAs from different sources, DNA and poly(A). Saporin-L1 depurinated DNA extensively and released adenine from all adenine-containing polynucleotides tested. Adenine was the only base released from DNA or artificial polynucleotides. The characteristics of the reactions catalysed by saporin-L1 have been determined: optimal pH and temperature, ionic requirements, and the kinetic parameters Km and kcat. The reaction proceeded without cofactors, at low ionic strength, in the absence of Mg2+ and K+. Saporin-L1 had no activity towards various adenine-containing non-polynucleotide compounds (cytokinins, cofactors, nucleotides). This plant protein may now be classified as a polynucleotide: adenosine glycosidase.

Animals↗
PubMed1981

Enzymatic synthesis of biotin-labeled polynucleotides: novel nucleic acid affinity probes.

Analogs of dUTP and UTP that contain a biotin molecule covalently bound to the C-5 position of the pyrimidine ring through an allylamine linker arm have been synthesized. These biotin-labeled nucleotides are efficient substrates for a variety of DNA and RNA polymerases in vitro. Polynucleotides containing low levels of biotin substitution (50 molecules or fewer per kilobase) have denaturation, reassociation, and hybridization characteristics similar to those of unsubstituted controls. Biotin-labeled polynucleotides, both single and double-stranded, are selectively and quantitatively retained on avidin-Sepharose, even after extensive washing with 8 M urea, 6 M guanidine hydrochloride, or 99% formamide. In addition, biotin-labeled polynucleotides can be selectively immunoprecipitated in the presence of antibiotin antibody and Staphylococcus aureus protein A. The unique features of biotin-labeled polynucleotides suggest that they will be useful affinity probes for the detection and isolation of specific DNA and RNA sequences.

Affinity Labels↗
PubMed1999

Activation of target-tissue immune-recognition molecules by double-stranded polynucleotides.

Abnormal expression of major histocompatibility complex (MHC) class I and class II in various tissues is associated with autoimmune disease. Autoimmune responses can be triggered by viral infections or tissue injuries. We show that the ability of a virus or a tissue injury to increase MHC gene expression is duplicated by any fragment of double-stranded (ds) DNA or dsRNA introduced into the cytoplasm of nonimmune cells. Activation is sequence-independent, is induced by ds polynucleotides as small as 25 bp in length, and is not duplicated by single-stranded polynucleotides. In addition to causing abnormal MHC expression, the ds nucleic acids increase the expression of genes necessary for antigen processing and presentation: proteasome proteins (e.g., LMP2), transporters of antigen peptides; invariant chain, HLA-DM, and the costimulatory molecule B7.1. The mechanism is different from and additive to that of gamma-interferon (gammaIFN), i.e., ds polynucleotides increase class I much more than class II, whereas gammaIFN increases class II more than class I. The ds nucleic acids also induce or activate Stat1, Stat3, mitogen-activated protein kinase, NF-kappaB, the class II transactivator, RFX5, and the IFN regulatory factor 1 differently from gammaIFN. CpG residues are not responsible for this effect, and the action of the ds polynucleotides could be shown in a variety of cell types in addition to thyrocytes. We suggest that this phenomenon is a plausible mechanism that might explain how viral infection of tissues or tissue injury triggers autoimmune disease; it is potentially relevant to host immune responses induced during gene therapy.

3T3 Cells↗
PubMed1987

Conformational peculiarities of polynucleotides with a nonrandom base sequence according to the 1H----3H exchange rate in C8H groups of purinic residues.

We have determined the 1H----3H exchange rate constants between water and C8H groups of purinic residues of alternating polynucleotides poly(dA-dT).poly(dA-dT), poly(dG-dC).poly(dG-dC) and poly(dA-dC).poly(dG-dT) as well as homopolynucleotides poly(dA).poly(dT) and poly(dG).poly(dC) in aqueous solutions with high-salt concentrations (3 M NaCl and 4-6 M CsF), in water-ethanol (60%) solution and in 0.15 M NaCl at 25 degrees C. The rate constants for adenine (kA) and guanine (kG) of polynucleotides were compared with corresponding constants for E. coli DNA. dGMP nd dAMP at the same conditions. The relation between exchange rates and conformations of polynucleotides permits the study of their conformational peculiarities in solution. Of three alternating polynucleotides examined in 0.15 M NaCl the exchange retardation was observed only for poly(dA-dT).poly(dA-dT) as compared with that in B-DNA, which is in good agreement with the B-alternating "wrinkled" DNA model. The conformations of poly(dG-dC).poly(dG-dC) and poly(dA-dC).poly(dG-dT), according to the exchange data obtained are within the B form. For homopolynucleotides in 0.15 M NaCl, the KA value for poly(dA).poly(dT) is nearly the same as kA for B-DNA, which indicates the similarity of their conformations, whereas the kG value for poly(dG).poly(dC) is 1.7-fold lower in comparison with the kG value in B-DNA. This seems to be connected with the existence of B = A conformation equilibrium for poly(dG).poly(dC) in solution. The increase of NaCl concentration to 3 M results in a B----Z transition in the case of poly(dG-dC).poly(dG-dC) and in the shift of B = A equilibrium towards the A-form in the case of poly(dG).poly(dC) as is evidenced by alterations of their KG values. Poly(dA-dT).poly(dA-dT) in 6 M CsF and poly(dA-dC).poly(dG-dT) in 4.3 M CsF maintain their inherent conformations in 0.15 M NaCl in spite of the fact that they are characterised by the "X-type" CD-spectrum at these conditions. According to the exchange data the conformation of poly(dA).poly(dT) in 6 M CsF corresponds to the "heteronomous" DNA model or some other structure with lower accessibility of C8H groups of adenylic residues.

Base Sequence↗
PubMed1995

Interaction specificity of benzimidazol group compounds with AT-containing polynucleotides.

The interaction between polynucleotides: poly(dA)-poly(dT), poly(dA-dT), poly(am2dA-dT), and the AT-specific compounds of benzimidazol group has been studied. It is been shown that these compounds bind to poly(dA)-poly(dT) and poly(dA-dT) at low and high salt concentration in solution. Poly(am2dA-dT) interacts with AT-specific compounds only at low salt, where this polynucleotide is in a B-form, but not at high salt, when the polynucleotide converts to another conformation. Thus, the interaction specificity of the groove-binding ligands is influenced not only by the minor groove substituents, but the peculiarities of the secondary structure of polynucleotides.

Benzimidazoles↗
PubMed1976

Thermostable polynucleotide phosphorylases from Bacillus stearothermophilus and Thermus aquaticus.

Polynucleotide phosphorylase from Bacillus stearothermophilus has been purified to homogeneity. Polyacrylamide gel electrophoresis run under denaturing conditions indicates that the enzyme is a tetramer with subunits of apparent molecular weight 51,000 daltons. A partial purification of polynucleotide phosphorylase from Thermus aquaticus has also been effected. The two enzymes show similar catalytic properties, which differ little from those of mesophilic polynucleotide phosphorylases. The use of thermostable polynucleotide phosphorylases for in vitro nucleic acid synthesis is discussed.

Geobacillus stearothermophilus↗
PubMed1975

The effects of thioketo substitution upon uracil-adenine interactions in polyribonucleotides. Synthesis and properties of the alternating polynucleotides poly(r(A-s2U)) and poly(r(A-s2s4U)).

The polynucleotides poly[r(A-s-2U)] and poly]r(A-s2s4U)] have been synthesized and characterized by nearest-neighbour analysis, sedimentation analysis as well as spectroscopic techniques. Absorption-temperature profile and absorption-pH profile of poly[r(A-s-2U)] did not reveal a structural transition between 10 and 95 degrees C even at low ionic strength, although a variety of properties indicated a helical structure of poly[r(A-s-2U)]: remarkable hyperchromicity of the absorption spectrum, circular dichroic spectrum displaying extrema of large amplitudes, resistance against hydrolysis by ribonuclease and interaction with ethidium bromide in a manner which is characteristic of helical polynucleotides. Our results show that interactions of the type A-s-2U and A-s-2s-4U do in fact exist in helical polynucleotides. The properties of poly]r(As-2U)] furthermore demonstrate the general stabilizing effect of 2-thioketopyrimidine bases in helical polynucleotides by virtue of vertical stacking interactions with neighbouring pyrimiding and purine bases.

Circular Dichroism↗
PubMed1977

Isolation and characterization of a polynucleotide phosphorylase from Bacillus amyloliquefaciens.

Bacillus amyloliquefaciens BaM-2 produces large amounts of extracellular enzymes, and the synthesis of these proteins appears to be dependent upon abnormal ribonucleic acid metabolism. A polynucleotide phosphorylase (nucleoside diphosphate:polynucleotide nucleotidyl transferase) was identified, purified, and characterized from this strain. The purification scheme involved cell disruption, phase partitioning, differential (NH4)2SO4 solubilities, agarose gel filtration, and diethylaminoethyl-Sephadex chromatography. The purified enzyme demonstrated the reactions characteristic of polynucleotide phosphorylase: polymerization, phosphorolysis, and inorganic phosphate exchange with the beta-phosphate of a nucleotide diphosphate. The enzyme was apparently primer independent and required a divalent cation. The reactions for the synthesis of the homopolyribonucleotides, (A)n and (G)n, were optimized with respect to pH and divalent cation concentration. The enzyme is sensitive to inhibition by phosphate ion and heparin and is partially inhibited by rifamycin SV and synthetic polynucleotides.

Adenosine Diphosphate↗
PubMed1989

Stabilization of the 3' one-third of Escherichia coli ribosomal protein S20 mRNA in mutants lacking polynucleotide phosphorylase.

Mutations which largely inactivate polynucleotide phosphorylase and which render RNase II thermolabile exert two effects on the metabolism of the two nested mRNAs which encode ribosomal protein S20. (i) The lifetime of both mRNA species is extended 2.5-fold at 38 degrees C in a strain harboring both mutations. (ii) A relatively stable truncated fragment of these mRNAs accumulates to significant levels in strains lacking polynucleotide phosphorylase. The truncated RNA (Po RNA) is 147 to 148 residues long and is coterminal with the 3' ends of intact S20 mRNAs. Its 5' end appears to be generated by endonucleolytic cleavage to the 5' side of a G residue in the sequence AACCGAUC. The data are consistent with the hypothesis that S20 mRNAs can be degraded by alternative pathways. The normal pathway depends on functional polynucleotide phosphorylase and is concerted, since S20 mRNAs disappear without accumulation of detectable intermediates in the decay process. The slower alternative pathway is followed when polynucleotide phosphorylase is inactivated by mutation. This pathway is distinguished by segmental rather than concerted degradation of S20 mRNAs and involves at least one endonucleolytic cleavage. The 5' two-thirds of S20 mRNAs decays significantly more quickly than the 3' third in this latter mode of mRNA turnover.

Alleles↗
PubMed1988

Cloned polynucleotide and synthetic oligonucleotide probes used in colony hybridization are equally efficient in the identification of enterotoxigenic Escherichia coli.

Restriction endonuclease-generated polynucleotide and synthetically produced oligonucleotide gene probes used in colony hybridization assays proved to be efficient for the detection and differentiation of enterotoxigenic Escherichia coli. To compare their relative efficiencies, these two sets of probes were radiolabeled with 32P and were applied to 74 strains of E. coli with known enterotoxin profiles and to 156 previously unexamined E. coli isolates. The enterotoxigenic bacteria Vibrio cholerae O1, Vibrio cholerae non-O1 (NAG), Yersinia enterocolitica, and E. coli harboring the plasmid vectors of the polynucleotide gene probes were examined for further evaluation of probe specificity. The two classes of probes showed a perfect concordance in their specific detection and differentiation of enterotoxigenic E. coli. In the analysis of six strains, the signal strength on autoradiography after hybridization with oligonucleotides was weaker than that obtained after hybridization with polynucleotide probes. The probes did not hybridize with DNA from V. cholerae O1, V. cholerae non-O1 (NAG), or Y. enterocolitica. The strains of E. coli harboring the plasmid vectors of the polynucleotide gene probes were, likewise, negative in the hybridization assays.

Base Sequence↗
PubMedDate not supplied

The formation of phosphate end groups in the radiolysis of polynucleotides in aqueous solution.

The polynucleotides poly(U), poly(C), poly(A) and poly(G) have been gamma-irradiated in N2O- and N2O/O2 (4:1)-saturated aqueous solutions. Hydroxyl radicals from the radiolysis of water react with the polynucleotides thereby producing among other lesions strand breaks. Strand breakage is connected with the formation of phosphomonoester end groups. Such end groups have been determined by measuring inorganic phosphate after a three hour incubation at 37 degrees C with acid or alkaline phosphatase. In the absence of oxygen G(phosphomonoester end groups) (in units of mumol J-1) are 0.47 (poly(U)), 0.17 (poly(C)) and less than or equal to 0.04 (poly(A) and poly(G)). In the case of poly(U) and poly(C) on heating the sample for one hour at 95 degrees C prior to incubation with phosphatases the above values increased by 0.14 and 0.07 mumol J-1, resp., whereas such treatment of the purine polynucleotides still did not produce a measurable yield of phosphomonoester end groups. Comparing these values with G values for strand breakage taken from the literature, about two phosphomonoester end groups are formed per strand break in poly(U) while for poly(C) this ratio is about unity. The purine polynucleotides show very low yields of strand breakage in agreement with the negligible phosphomonoester yields. In the presence of oxygen G(phosphomonoester end groups) are 0.46 (poly(U)), 0.21 (poly(C)), and less than or equal to 0.04 (poly(A) and poly(G)). On heating, these values increase, most markedly for poly(U) and poly(C). This is possibly linked to the decomposition of unstable hydroperoxides which are formed in high yields in poly(U) and poly(C) (G = 0.7 and 0.19 mumol J-1, resp.).(ABSTRACT TRUNCATED AT 250 WORDS)

Borohydrides↗
PubMed1987

Nucleotide sequence of the pnp gene of Escherichia coli encoding polynucleotide phosphorylase. Homology of the primary structure of the protein with the RNA-binding domain of ribosomal protein S1.

The pnp gene is located at 69 min on the Escherichia coli chromosome adjacent to the rpsO gene which encodes the ribosomal protein S15. In this paper, we present the sequence of a 3030-nucleotide DNA fragment containing the open reading frames coding for ribosomal protein S15 and polynucleotide phosphorylase. Translation of pnp is initiated by 5'-UUG-3' codon separated by 7 nucleotides from a good ribosome binding site. Codon usage in this gene is typical of highly expressed proteins of E. coli. Some of the transcripts of the pnp gene terminate just after the stem of the terminator t2 visible in the nucleotide sequence. However, a very strong read-through occurs at this site, thus permitting many of the pnp transcripts to extend beyond this transcription terminator. We also describe the primary structure homologies between a 69-amino-acid stretch of polynucleotide phosphorylase and the four homologous stretches of ribosomal protein S1 which form its RNA binding site. The possibility that this 69-amino-acid stretch constitutes the polynucleotide binding domain of polynucleotide phosphorylase is discussed.

Amino Acid Sequence↗
PubMedDate not supplied

[Interaction of gene 5 protein of phage f1 with single and double-stranded DNA and polynucleotides].

The fluorescence method was used to reveal some differences in the interaction of gene 5 protein of phage f1 with single- and double-stranded polynucleotides (DNA). The binding with the duplexes is non-cooperative and the Kapp is twice lower than that for the cooperative formation of the complex with single-stranded structures. In the complex with a double-stranded polynucleotide (DNA) the protein cover 3 nucleotide pairs. The complex dissociates with a lower concentration of salt and the contribution of the energy of nonelectrostatic interactions to the total energy of complex formation for it is lower than for the complex with single-stranded DNA. In the complex of protein with single-stranded structure the fluorescence of the tyrosine (Tyr) residues is quenched to a greater degree and their accessibility to the external quencher is lower than that of the complex with double-stranded polynucleotides (DNA). The suggestion is made that in destabilization of nucleic double helices by gene 5 protein of phage f1, a great role belongs to Tyr residues because of their high affinity to single-stranded structures and because of their different localization in the complexes with single- and double-stranded polynucleotides.

Coliphages↗
PubMedDate not supplied

[Glucocorticoid-receptor complexes of rat liver. II. Interaction with natural and synthetic polynucleotides].

The relative affinities of [3H]dexamethasone-labelled glucocorticoid-receptor complexes (GRC) from rat liver to various natural and synthetic polynucleotides differing in base composition and secondary structure have been determined. The modified competition assay procedure with use of DNA-cellulose was employed. The interaction of 150-2000 fold purified GRC with polyribo- and polydeoxyribonucleotides greatly depends on their base composition and nucleotide sequences. This interaction is hardly affected by the structure (ribo- or deoxyribo-) of the sugar-phosphate backbone or its size and is in fact independent of the secondary structure (single- or double-stranded forms) of polynucleotides. Mixtures of mononucleotides or apurinic DNA do not compete with DNA-cellulose for GRC binding. In the presence of cytosol, GRC bind double-stranded rather than single-stranded DNA, the fact possibly reflecting different effects of cytoplasmic proteins on the accessibility to GRC of polynucleotides of dissimilar secondary structure. The apparent equilibrium dissociation constants (Kd) of the ternary complexes of GRC with some synthetic polynucleotides were calculated. In case of double-stranded polydeoxyribonucleotide poly(dA) . poly(dT) Kd is equal to (1.2 +/- 0.6) x 10(-4) M; this value may characterize a nonspecific GRC--DNA interaction. It is supposed that GRC interact with high affinity (Kd approximately 10(-7)--10(-8) M) with short (5-10 nucleotides long) DNA sequences in which certain purine and pyrimidine residues are interspersed. Such DNA sequences may represent sites of primary action of GRC on the cellular genome in vivo.

Animals↗
PubMed1995

A carcinoembryonic antigen polynucleotide vaccine has in vivo antitumor activity.

We have constructed a plasmid DNA encoding the full-length cDNA for human carcinoembryonic antigen (CEA) driven by the cytomegalovirus early promoter/enhancer and demonstrated that this plasmid can function as a polynucleotide vaccine. The immune response elicited by the CEA polynucleotide vaccine is dose and schedule dependent. There appears to be a threshold dose of 50 micrograms capable of inducing CEA-specific lymphoblastic transformation, lymphokine release, and antibody response. Doses of 10 micrograms were significantly less effective. When 50-micrograms doses are employed, thrice weekly or weekly vaccination schedules more reliably elicit CEA-specific immune responses by day 43 than does an every-3-weeks schedule. Furthermore the CEA polynucleotide vaccine can immunoprotect against challenge with syngeneic CEA-transduced colon carcinoma cells as early as 3 weeks after the first vaccination. Studies are ongoing to demonstrate the ability of CEA polynucleotide vaccination to treat pre-existing syngeneic mouse colon and breast carcinomas expressing human CEA.

Animals↗
PubMed1987

Bacteriophage T4 anticodon nuclease, polynucleotide kinase and RNA ligase reprocess the host lysine tRNA.

Host tRNAs cleaved near the anticodon occur specifically in T4-infected Escherichia coli prr strains which restrict polynucleotide kinase (pnk) or RNA ligase (rli) phage mutants. The cleavage products are transient with wt but accumulate in pnk- or rli- infections, implicating the affected enzymes in repair of the damaged tRNAs. Their roles in the pathway were elucidated by comparing the mutant infection intermediates with intact tRNA counterparts before or late in wt infection. Thus, the T4-induced anticodon nuclease cleaves lysine tRNA 5' to the wobble position, yielding 2':3'-P greater than and 5'-OH termini. Polynucleotide kinase converts them into a 3'-OH and 5' P pair joined in turn by RNA ligase. Presumably, lysine tRNA depletion, in the absence of polynucleotide kinase and RNA ligase mediated repair, underlies prr restriction. However, the nuclease, kinase and ligase may benefit T4 directly, by adapting levels or decoding specificities of host tRNAs to T4 codon usage.

Base Sequence↗
PubMed1986

Purification of wheat germ RNA ligase. I. Characterization of a ligase-associated 5'-hydroxyl polynucleotide kinase activity.

An RNA ligase that catalyzes the formation of a 2'-phosphomonoester-3',5'-phosphodiester bond in the presence of ATP and Mg2+ was purified approximately 6000-fold from raw wheat germ. A 5'-hydroxyl polynucleotide kinase activity copurified with RNA ligase through all chromatographic steps. Both activities cosedimented upon glycerol gradient centrifugation even in the presence of high salt and urea. RNA ligase and kinase activities sedimented as a single peak on glycerol gradients with a sedimentation coefficient of 6.2 S. The purified polynucleotide kinase activity required dithiothreitol and a divalent cation for activity and was inhibited by pyrophosphate and by ADP. The kinase phosphorylated a variety of 5'-hydroxyl-terminated polynucleotide chains including some that were substrates for the RNA ligase (e.g. 2',3'-cyclic phosphate-terminated poly(A)) and others that were not ligase substrates (e.g. DNA or RNA containing 3'-hydroxyl termini). RNA molecules containing either 5'-hydroxyl or 5'-phosphate and 2',3'-cyclic or 2'-phosphate termini were substrates for the purified RNA ligase activity. The rate of ligation of 5'-hydroxyl-terminated RNA chains was greater than that of 5'-phosphate-terminated molecules, suggesting that an interaction between the wheat germ kinase and ligase activities occurs during the course of ligation.

Anions↗
PubMed2001

Three-way multivariate curve resolution applied to speciation of acid-base and thermal unfolding transitions of an alternating polynucleotide.

Analytical speciation of acid-base equilibria and thermal unfolding transitions of an alternating random polynucleotide containing cytosine and hypoxanthine, poly(C, I), is studied. The results are compared with those obtained previously for single-stranded polynucleotides, poly(I) and poly(C), and for the double-stranded poly(I). poly(C), to examine the influence of the secondary structure on the acid-base properties of bases. This study is based on monitoring acid-base titrations and thermal unfolding experiments by molecular absorption, CD, and molecular fluorescence spectroscopies. Experimental data were analyzed by a novel chemometric approach based on a recently developed three-way Multivariate Curve Resolution method, which allowed the simultaneous analysis of data from several spectroscopies. This procedure improves the resolution of the concentration profiles and pure spectra for the species and conformations present in folding-unfolding and acid-base equilibria. The results from acid-base studies showed the existence of only three species in the pH range 2-12 at 37 degrees C and 0.15M ionic strength. No cooperative effects were detected from the resolved concentration profiles, showing that equilibria concerning alternating polynucleotides like poly(C, I) are simpler than those involving poly(I). poly(C). Thermal unfolding experiments at neutral pH confirmed the existence of two transitions and one intermediate conformation. This intermediate conformation could only be detected and resolved without ambiguities when molecular absorption and CD spectral data were analyzed simultaneously.

Algorithms↗