Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Formamides”

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 541 records · Page 30Linked to original sources
PubMed2001

The enhancement of PCR amplification by low molecular weight amides.

Amplification of a DNA target by the polymerase chain reaction (PCR) often requires laborious optimization efforts. In this regard, the use of certain organic chemicals such as dimethyl sulfoxide, polyethylene glycol, betaine and formamide as cosolvents has been found to be very helpful. Unfortunately, very little is known about the precise structural features that make these additives effective and, accordingly, the number of such chemicals currently known to enhance PCR is limited. In order to address these issues, we decided to focus on formamide and undertook an extensive study of low molecular weight amides as a class to see how changing the substituents in the amide structure influences its effect on PCR. We describe here the results of this study, which involved 11 different amides, and present observations that provide a cohesive picture of structure-activity relations in this group of additives. We found several of these amides to be exceptionally effective and introduce them as novel PCR enhancers.

Acetamides↗
PubMed2001

Temperature sensitive nop2 alleles defective in synthesis of 25S rRNA and large ribosomal subunits in Saccharomyces cerevisiae.

Using molecular genetic techniques, we have generated and characterized six temperature sensitive (ts) alleles of nop2. All failed to support growth at 37 degrees C and one was also formamide sensitive (fs) and failed to grow on media containing 3% formamide. Conditional lethality is not due to rapid turnover of mutant Nop2p proteins at 37 degrees C. Each allele contains between seven and 14 amino acid substitutions and one possesses a nonsense mutation near the C-terminus. Mapping experiments with one allele, nop2-4, revealed that a subset of the amino acid substitutions conferred the ts phenotype and that these mutations have an additive effect. All six mutants exhibited dramatic reductions in levels of 60S ribosome subunits under non-permissive conditions as well as some reduction at permissive temperature. Processing of 27S pre-rRNA to mature 25S rRNA was defective in all six mutants grown under non-permissive conditions. Levels of the 40S ribosomal subunit and 18S rRNA were not significantly affected. Amino acid substitutions in nop2 conditional alleles are discussed in the context of the hypothesis that Nop2p functions both as an RNA methyltransferase and a trans-acting factor in rRNA processing and large ribosomal subunit biogenesis.

Alleles↗
PubMed1994

Alternative routes of enzymic cyanide metabolism in Pseudomonas fluorescens NCIMB 11764.

Cell-free extracts from Pseudomonas fluorescens NCIMB 11764 catalysed the degradation of cyanide into products that included CO2, formic acid, formamide and ammonia. Cyanide-degrading activity (CDA) was localized to cytosolic cell fractions and was observed at substrate concentrations as high as 100 mM (2600 mg CN-I-1). At least two different CDAs could be distinguished by: (i) the determination of reaction product stoichiometries, (ii) requirements for NADH and oxygen, and (iii) kinetic analysis. The first activity produced CO2 and NH3 as reaction products, was dependent on oxygen and NADH for activity, and displayed an apparent Km for cyanide of 1.2 mM. The second activity generated formic acid (and NH3) plus formamide as reaction products, was oxygen independent, and had an apparent Km of 12 mM for cyanide. The first enzymic activity was identified as cyanide oxygenase as previously described [Harris, R. E. & Knowles, C. J. (1983) FEMS Microbiol Lett 20, 337-341] whereas the second activity is believed to consist of two enzymes, a cyanide nitrilase (dihydratase) and hydratase (EC 4.2.1.66). In addition to these enzymes, cyanide-grown cells were also induced for formate dehydrogenase (EC 1.2.1.2) thereby providing a means of recycling NADH utilized by cyanide oxygenase. A mutant strain having lost the ability to grow on cyanide as a nitrogen source was isolated and shown to be defective in cyanide oxygenase, but not the cyanide nitrilase/hydratase enzymes. This finding together with results showing that the substrate affinity of cyanide oxygenase was tenfold greater than for the nitrilase/hydratase enzymes, indicates that it is this enzyme that is most important in cyanide assimilation.(ABSTRACT TRUNCATED AT 250 WORDS)

Ammonia↗
PubMed1990

Fructose-1,6-bisphosphate-activated pyruvate kinase from Escherichia coli. Nature of bonds involved in the allosteric mechanism.

The allosteric properties of the fructose-1,6-bis-phosphate-activated pyruvate kinase from Escherichia coli were examined in the presence of a number of fructose bisphosphate analogues, as well as of increased ionic strength (NaCl) and of the hydrogen-bond-breaking agent, formamide. Fructose 2,6-bisphosphate, ribulose 1,5-bisphosphate and 5-phosphorylribose 1-pyrophosphate gave allosteric activation (additive to that of fructose 1,6-bisphosphate). Formamide always decreased Vmax, but left unchanged the Km for phosphoenolpyruvate, while it decreased the concentration of fructose bisphosphate required to give half-maximal activity (K0.5). NaCl increased the K0.5 for both phosphoenolpyruvate and fructose bisphosphate, leaving Vmax unchanged. These results are consistent with ionic binding of fructose bisphosphate through phosphates and with a critical role of hydrogen bonds in stabilizing both the inactive and the active enzyme conformers.

Allosteric Regulation↗
PubMed1990

Ciguatoxin enhances quantal transmitter release from frog motor nerve terminals.

1. Ciguatoxin (CTX), a marine toxin produced by the benthic dinoflagellate Gambierdiscus toxicus, is responsible for a complex endemic disease in man known as ciguatera fish poisoning. In the present study we have investigated the effects of purified CTX extracted for Gymnothorax javanicus moray-eel liver on frog isolated neuromuscular preparations with conventional electrophysiological techniques. 2. CTX (1-2.5 nM) applied to cutaneous pectoris nerve-muscle preparations induced, after a short delay, spontaneous fibrillations of the muscle fibres that could be suppressed with 1 microM tetrodotoxin (TTX) or by formamide to uncouple excitation-contraction. 3. In preparations treated with formamide, CTX (1-2.5 nM) caused either spontaneous or repetitive muscle action potentials (up to frequencies of 60-100 Hz) in response to a single nerve stimulus. Recordings performed at extrajunctional regions of the muscle membrane revealed that during the repetitive firing a prolongation of the repolarizing phase of the action potential occurred. At junctional sites the repetitive action potentials were triggered by repetitive endplate potentials (e.p.ps). 4. CTX (2.5 nM) caused a TTX-sensitive depolarization of the muscle membrane. 5. In junctions equilibrated in solutions containing high Mg2+ + low Ca2+, addition of CTX (1.5 nM) first induced an average increase of 239 +/- 36% in the mean quantal content of e.p.ps. Subsequently CTX reduced and finally blocked nerve-evoked transmitter release irreversibly. 6. CTX (1.5-2.5 nM) increased the frequency of miniature endplate potentials (m.e.p.ps) in junctions bathed either in normal Ringer, low Ca2(+)-high Mg2+ medium or in a nominally Ca2(+)-free solution containing EGTA.2+ Extensive washing with toxin-free solutions did not reverse the effect. Furthermore, Cd2 + (0.1 mM), a potent calcium channel blocker, neither antagonized nor abolished the increase in transmitter release caused by CTX. 7. TTX (1 microM) completely prevented the effect of CTX (2.5nM) on m.e.p.p. frequency. This effect was independent of the presence of extracellular Ca2 +. TTX, when added after CTX (2.5 nM) exposure, antagonized the increase in m.e.p.p. frequency. The antagonism was complete in Ca2 +-free medium. These results strongly suggest that increased permeability of the nerve terminal to Na+ is responsible for the increase in m.e.p.p. frequency caused by CTX. It is likely that CTX may trigger calcium release from internal stores due to an increase of intraterminal Na+ concentration. 8. It is concluded that CTX exerts, in the nanomolar concentration range, a selective action on sodium channels of the neuromuscular junction causing both pre- and postsynaptic effects.

Animals↗
PubMed1973

Inhibition of macrophage migration by nucleotide-containing streptococcal preparations.

Certain extracts of streptococcal cell walls are known to inhibit macrophage migration in vitro. In this study, we attempted to identify the streptococcal components responsible for this phenomenon. Trypsinized cell walls and cytoplasm from groups A and B streptococci were extracted with hot formamide followed by acetone precipitation. Subsequent gel filtration in aqueous solutions yielded a fraction devoid of C-carbohydrate and containing mostly oligonucleotides, apparently derived from streptococcal cytoplasm. This fraction significantly inhibited the migration of peritoneal exudate cells from rats sensitized to groups A and B streptococci. It was noteworthy that no inhibition of migration was observed with cells from nonsensitized animals or control rats injected with BCG or complete Freund adjuvant. Similarly, no inhibition was obtained with formamide extracts of calf thymus RNA. Although the inhibition does not show specificity for streptococcal groups, it seems to have immunological specificity since prior sensitization with streptococci is required for migration inhibition.

Animals↗
PubMed1967

Ultrastructure of the cell wall of Bacillus polymyxa.

The macromolecular arrangement on the surface of Bacillus polymyxa was revealed by metal shadowing of whole cells and wall fragments; it consisted of a rectangular array of 70-A globules with a repeating interval of 100 A. The substructure was studied in plan with phosphotungstic acid (pH 6) or uranyl acetate as negative stains of fragments and was studied also in profile with sections of embedded material. Staining of sections of cells fixed with glutaraldehyde showed that layering (approx. 80-A dense, 40-A light, and 120-A dense layers, outermost layer first) could be demonstrated in the cell wall with lead or uranyl acetate, used together or separately. The outer "dense" layer corresponded to the regularly arrayed structure (RS); it was removed by guanidine hydrochloride, sodium lauryl sulfate, cold formamide, and by trypsin. The RS layer (isolated by a hydrogen bond breaking reagent, guanidine hydrochloride) was disrupted by agents such as sodium lauryl sulfate or damaged by 3 m sodium chloride. Qualitative chemical tests, ultraviolet absorption, and removal by trypsin indicated that the structured layer consisted mainly of protein, but exact characterization was not attempted. The globular units making up the layer consisted of a small number of subunits, imperfectly resolved by negative staining. The underlying polysaccharide appeared to be covalently bound to the deepest (probably mucopeptide) layer since it required "hot" formamide for its removal. A survey of species was not made.

Bacillus↗
PubMed1972

Isolation and characterization of simian virus 40 ribonucleic acid.

Deoxyribonucleic acid-ribonucleic acid (RNA) hybridization in formamide was used to isolate simian virus 40-specific RNA. Early in the lytic cycle, a 19S viral RNA species was observed. Late in the lytic cycle, 16S and 19S viral species were found. The 16S and 19S species of viral RNA were localized in the cytoplasm. High-molecular-weight heterogeneous RNA, containing viral sequences, was isolated from the nuclear fraction of infected cells late in the lytic cycle. This RNA may contain non-viral sequences linked to viral sequences. The formamide hybridization technique can be used to isolate intact late lytic viral RNA which is at least 99% pure.

Animals↗
PubMed1975

RNA subunit structure of Mason-Pfizer monkey virus.

Mason-Pfizer monkey virus 60-70S RNA has a molecular weight of 8 times 10-6 when analyzed on polyacrylamide gels. Dissociation of 60-70S RNA of Mason-Pfizer monkey virus and murine leukemia virus by heat or formamide (40%) resulted in conversion to identical subunit structures of 2.8 times 10-6 daltons; treatment with lower amounts of formamide revealed a partial dissociation of Mason-Pfizer monkey virus 60-70S RNA released three low-molecular-weight RNA species of 10-5, 3,5 times 10-4, and 2.5 times 10-4.

Animals↗
PubMed1976

Comparative electrophoresis of the 18-22S RNAs of Newcastle disease virus.

Between 80 and 90% of the 18-22S Newcastle disease virus intracellular RNA molecules contain poly(A) sequences. Electrophoresis of the 18S RNA in formamide-polyacrylamide gels resolves five species resolved by electrophoresis in aqueous gels. Thus, these five RNA species are probably unique size classes of RNA and not different conformations of the same RNAs. They are of sufficient size to code for the five smaller Newcastle disease virus proteins, and their combined molecular weights represent 60% of the viral genome-a value identical to that obtained by annealing 18-22S RNA with genome RNA. Formamide or heat treatment of the 22S RNA converts most of it into species with migration rates similar to those of the 18S species. Thus, the 22S RNA may not contain unique RNA species.

Chromatography, Gel↗
PubMed1989

Form-determining functions in Sindbis virus nucleocapsids: nucleosomelike organization of the nucleocapsid.

Purified intact Sindbis virus nucleocapsids were treated at different pH values or with various concentrations of divalent cations, cation chelators, salt, or formamide. The resulting structures were examined by velocity sedimentation, electron microscopy, and protein-protein cross-linking. Changes in each of the test conditions led to alterations in the sedimentation profile of treated nucleocapsids. Appropriate concentrations of formamide or divalent cations generated beaded strandlike structures similar in morphology to those generated from adenovirus cores and nucleosomes. The capsid protein and RNA remained associated with each other at NaCl concentrations less than or equal to 1 M or after treatment of the structures with alkaline pH up to and including pH 10.7. Protein and RNA were dissociated by salt concentrations of greater than 1 M, suggesting that the arginine-rich, amino-terminal portion of the capsid protein is responsible for binding the RNA. Protein-protein cross-linking also indicated that the capsid proteins remained associated in small aggregates under some of the conditions that caused dissociation of the nucleocapsid and suggested the presence of more than one type of protein-protein interaction in the nucleocapsids. Collectively, these data suggest that, like histones and adenovirus core proteins, the Sindbis virus capsid protein serves to package segments of the genome into nucleoprotein beads which are capable of interacting with each other to form the nucleocapsid structure.

Capsid↗
PubMed1996

Isoelectric focusing of crystallins in microsections of calf and adult bovine lens. Identification of water-insoluble crystallins complexing under nondenaturing conditions: demonstration of chaperone activity of alpha-crystallin.

Topographic studies of crystallin fractions from the young adult bovine lens revealed that lenses do not have a homogeneous distribution of crystallins. There are, however, gradual differences between the cortices and the nucleus. The isolated lenses were separated mechanically into lens equator and inner cylinder. The latter was then sectioned in a special sectioning machine into 11-12 morphological layers (from anterior cortex through nucleus to posterior cortex). Matters of the lens sections were separated into water-soluble (WS) and water-insoluble (WI) crystallins. The WI fractions were solubilized with 100% formamide, or dissolved into 7 M urea. Crystallin profiles were obtained for each lens layer, using thin-layer isoelectric focusing in polyacrylamide gel. WS crystallins from the lens equator revealed a separation into HM-, alpha L-, beta H-, beta L-, beta S- and gamma-crystallins. The WI fractions of the layers dissolved in urea gave a separation into the individual HM- (3 components), alpha L- (4 components), beta- (6 component groups), beta S- (2 components) and gamma- (11 components) crystallins in the different morphological layers. The results confirm that a significant age-related increase in several beta- and gamma-crystallins incorporated into alpha-crystallins exists in the patterns of WI fractions of the different layers from lenses of 2.2 and 5.9 years. The WI crystallins solubilized in formamide showed only the presence of HM weight and alpha-crystallin moieties, due to the action of chaperone activity of alpha-crystallin. The nature of the WI protein fraction in the separated lens layers reflected to the aggregated state of: alpha L-, beta L-, beta S- and gamma-crystallins in the different regions of the lens, concealed in the central cavity of the alpha-crystallin chaperone model.

Aging↗
PubMed2004

Immunoassay for single-stranded DNA in apoptotic cells.

The apoptosis assay described in this chapter is based on the selective denaturation of DNA in condensed chromatin of apoptotic cells and the detection of denatured DNA with a monoclonal antibody highly specific to single-stranded DNA. Optimal results are obtained by the heating at a relatively low temperature in the presence of formamide. The assay detects apoptotic cells but not necrotic cells or cells with DNA breaks in the absence of apoptosis. The sensitivity of the assay reflects the detection of early and late apoptosis. Apoptotic cells are detected in the sections of frozen or formalin-fixed paraffin-embedded tissues by immunohistochemistry and in the cell suspensions by flow cytometry or fluorescence microscopy. Apoptosis enzyme-linked immunoassay based on DNA denaturation by formamide in microtiter plates and one-step immunostaining is applied for high-throughput screening of drugs. The enzyme-linked immunoassay has the ability to distinguish anticancer drugs from toxic chemicals, to predict selective toxicity to cancer cells, and to detect drug synergism.

Animals↗
PubMed2003

Kinetics of denaturation of human and chicken hemoglobins in the presence of co-solvents.

The stability of four hemoglobins (Hb) in dimer forms (low concentration) were investigated by the kinetics of denaturation. The rate constants of denaturation were obtained by variation of 280 nm absorption versus time in 10 mM Tris-HCl, 10 mM EDTA, pH 8.0 at 45 degrees C in the absence and presence of 0.5 M ethanol, dimethyl sulfoxide (DMSO), formamide, and glycerol. The results show the trend of rate constants in different co-solvents in the following order: chicken hemolysate < human hemolysate and chicken Hb D < chicken Hb A. The buried surface area was calculated for Hb samples in the absence of co-solvents. Accordingly, the trend points out that: chicken Hb D > chicken Hb A > human Hb A. These results suggest that both chicken hemolysate and chicken Hb D are relatively more stable than human and chicken Hb A, respectively. However, the denaturation rate constants of Hb in different co-solvents have designated the following order: ethanol > DMSO > formamide > glycerol. As a matter of fact, this phenomenon is an indication of an increase in the denaturation capacity (DC) and hydrophobicity, and a decrease in the surface tension of the solution in the preceding co-solvents.

Animals↗
PubMed1975

[Comparison of the activities of free and carrier fixed horseradish peroxidase].

Commerical horseradish-peroxidase was covalently bound to BrCN-activated sepharose. The activity parameters Vmax and Km were determined by the leukomalachite green reaction. Compared with the soluble enzyme, the immobilized POD has a relative residual activity of 22%. Dimethylsulfoxide and formamide were found to diminish the enzymatic activity in a concentration-dependent manner. The activity of the free enzyme in aqueous formamide solution (10%) is reduced by 43%, that of the insolubilized enzyme by 68%. Dimethylsulfoxide (15%) does not alter the LMG conversion rate of free POD, whilst a rate loss by 60% was observed for the immobilized enzyme.

Binding Sites↗
PubMed1992

Evaluation of DNA probe removal from nylon membrane.

Genetic fingerprinting is one of the most challenging applications of any hybridization membranes. Forensic DNA fingerprinting typically uses samples in the range of 100-400 ng of genomic DNA. To ensure the ability to successfully reprobe the samples, it is imperative that repeated stripping of sample DNA be minimized while stripping of the probe DNA be maximized. By using standard dilutions of K562 cell line, we compared the following three stripping techniques: NaOH at 25 degrees C, formamide (HCONH2) at 65 degrees C and 0.1 x standard saline citrate and 0.1% sodium dodecyl sulfate at 95 degrees C (high-temperature stripping). The largest amount of genomic DNA was stripped from the membrane with NaOH, with the other two techniques removing less. Formamide and high-temperature procedures resulted in a loss of approximately 5-10 ng of DNA per strip. In contrast, the NaOH resulted in a loss of approximately 10-20 ng per strip.

Autoradiography↗
PubMed1976

Stepwise transition of aggregate structure of high-molecular-weight avian myeloblastosis virus RNA. Mode of releasing of associated 4S RNA.

Mode of releasing of associated 4S RNA species was studied during a controlled transition of aggregate structure of high-molecular-weight AMV-RNA. It has been found that associated 4S RNA constitutes 2.5% of 60S AMV-RNA complex. Approximately 60% of associated 4S RNA is successively released during treatment of viral RNA with increasing formamide concentration, concomitantly with the transition of 60S RNA aggregate through 50--55S RNA intermediate into the final 30--40S RNA subunits. 40% of 4S RNA remains associated with 30--40S RNA subunits prepared by formamide treatment and can be released from them by heating. A procedure is thus provided both for the isolation of oncornaviral RNA subunit structures deprived of various partions of associated 4S RNA and for the fractionation of 4S RNA species according to their binding affinity to the genome oncornaviral RNA.

Avian Leukosis Virus↗
PubMedDate not supplied

Formation and reversibility of S-linked conjugates of N-(1-methyl-3,3-diphenylpropyl)isocyanate, an in vivo metabolite of N-(1-methyl-3,3-diphenylpropyl)formamaide, in rats.

The metabolic disposition of N-(1-methyl-3,3-diphenylpropyl) formamide was studied in rats. The water-soluble metabolites, N-acetyl-S-[N-(1-methyl-3,3-diphenylpropylcarbamoyl)]cysteine and S-[N-(1-methyl-3,3-diphenylpropylcarbamoyl)]glutathione, were identified in urine and bile, respectively, of rats doses with the secondary formamide. The structures of these metabolites were confirmed by comparison with synthetic standards and by using liquid chromatography mass spectrometry and fast atom bombardment mass spectrometry. Synthetic standards of these metabolites were obtained by reacting the N-(1-methyl-3,3-diphenylpropyl)isocyanate with glutathione or N-acetylcysteine in methanolic solutions. The isocyanate was obtained in high yield by reacting 1-methyl-3,3-diphenylpropylamine with trichloromethyl chloroformate. The S-linked conjugates released the isocyanate in mild alkali, but were stable under acidic conditions. The released isocyanate was characterized by comparison with the synthetic standard using GC/MS and HPLC. A mechanism is proposed for the base-catalyzed elimination of the isocyanate from the thiol conjugates.

Acetylcysteine↗