Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.27.1 (RNase)
16,360 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Lysates containing folded chromosomes of competent Bacillus subtilis were prepared. The chromosomes were supercoiled, as indicated by the biphasic response of their sedimentation rates to increasing concentrations of ethidium bromide. Limited incubation of the lysates with increasing concentrations of ribonucleases resulted in a gradual decrease in the sedimentation velocity of the deoxyribonucleic acid (DNA) until finally a constant S value was reached. Incubation with sonicated, 4,5',8-trimethylpsoralen-monoadducted, denatured, homologous donor DNA molecules at 37 degrees C and concomitant irradiation with long-wave ultraviolet light of the nucleoid-containing lysates resulted in the formation of complexes of the donor DNA molecules and the recipient chromosomes. This complex formation was stimulated when nucleoids were previously (i) unfolded by ribonuclease incubation, (ii) (partially) relaxed by X irradiation, or (iii) subjected to both treatments. Monoadducts were not essential. On the other hand, the complex-forming capacity of recipient chromosomes previously cross-linked by 4,5',8-trimethylpsoralen diadducts was greatly reduced, suggesting that strand separation of the recipient molecule was involved in the formation of the complex. None of these effects has been observed when heterologous (Escherichia coli) donor DNA has been used. When the same kind of experiments were carried out at 70 degrees C, donor-recipient DNA complexes were also formed and required strand separation and homology similar to donor-recipient complex formation at 37 degrees C. However, in contrast to what was found at 37 degrees C, unfolding plus relaxation of the nucleoids, as well as the absence of monoadducts in the donor DNA fragments, resulted in a decrease in complex formation. On the basis of these results, we assume that superhelicity can promote the in vitro assimilation of single-stranded donor DNA fragments by nucleoids of competents B. subtilis cells at 70 degrees C, but that at 37 degrees C a different mechanism is involved.
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PMID:Assimilation of single-stranded donor deoxyribonucleic acid fragments by nucleoids of competent cultures of Bacillus subtilis. 678 88

When lymphocytes obtained from W/Fu rats primed with BCG are cultured in the presence of PPD, they elaborate a factor that is capable of potentiating the specific in vitro generation of cytotoxic lymphocytes to syngeneic (C58NT)D lymphoma cells and to BN alloantigen. Purification of the factor, achieved by gel filtration on Sephadex G-100, was facilitated by using a serum-free culture condition and the removal of the specific stimulating antigen, PPD, after an initial incubation period. The factor isolated contains DNA by its absorption spectrum, resistance to trypsin and RNase, but complete susceptibility to DNAase, and by the presence of ethidium bromide-positive material in the purified sample. It displays a 260/280 nm absorption ratio of 1.6 and a m.w. estimate of 10,000 to 30,000. Electrophoresis of the purified factor on agarose gel yielded three ethidium bromide reactive bands. Data obtained following the slicing and elution of these bands, and then testing for potentiating activity indicated that two of the three bands contained potentiating activity.
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PMID:Specific enhancement of immune responses by BCG: isolation of extracellular DNA from supernatants of specifically stimulated BCG-primed lymphoid cells. 698 4

The interaction of human platelets and Candida albicans was studied. Platelet-rich plasma was obtained from freshly drawn blood or outdated platelet concentrates. From the platelet-rich plasma, a platelet extract was derived which stimulated germ tube formation by C. albicans when incubated with yeast cells at 37 degrees C. The active component(s) was heat stable, trypsin sensitive, and ribonuclease and deoxyribonuclease insensitive, and possessed cationic properties since it readily attached to carboxymethyl-Sephadex. The active component(s) seemed to bind to heparin also, since germ tube-promoting activity was eluted from a heparin-cyanogen bromide-activated Sepharose 4B column. In addition, platelet-derived growth factor (Collaborative Research, Inc.) stimulated germination when incubated with low amounts (0.4% final concentration) of bovine calf serum. The aggregation of platelets, prepared as platelet-rich plasma by C. albicans cell wall or alkali-extracted cell wall fractions, was also studied. Aggregation of platelets was observed when cell wall or cell wall fractions were incubated with platelet-poor plasma at 37 degrees C for 20 min and then added to platelet-rich plasma. The component of platelet-poor plasma which promoted aggregation of platelets by C. albicans cell wall or alkali-extracted fractions was inactivated at 56 degrees C (30 min) and by cobra venom factor, indicating a role for the alternate complement pathway in the aggregation response.
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PMID:Platelet interactions with Candida albicans. 703 46

DNA synthesis in rat hepatocytes, from livers regenerating after 70% hepatectomy, was assessed by flow cytometric determination of nuclear DNA content and by incorporation of [3H]thymidine. Parenchymal liver cells were isolated by collagenase perfusion and low-speed centrifugation. Nuclei from the isolated cells were prepared for flow cytometry by a treatment with detergent, pepsin and RNase, and stained with ethidium bromide. Parallel samples of cells were incubated with [3H]thymidine and analysed for rate of incorporation of radioactivity into DNA and for labelling index determination. The flow cytometric measure of the replicative response, i.e. the presence of cells with S-phase DNA content within the diploid and tetraploid cell populations, was compared with the incorporation of [3H]thymidine. For each of fourteen animals, including two control rats and twelve partially hepatectomized animals killed either before (at 13 hr after hepatectomy), at the onset (16 and 18 hr) or at the peak (24 hr) of regenerating activity, a fairly good correlation was found between the different methods. Satisfactory resolution of the flow cytometric detection of S-phase cells was indicated by a sorting experiment using an Ortho (system 50-H) cell sorter which demonstrated that after [3H]thymidine injection in vivo 88% of the diploid and 84% of the tetraploid S-phase nuclei were labelled, while labelling in the G1-fractions was only 2 and 7%, respectively.
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PMID:The proliferation response of rat liver parenchymal cells after partial hepatectomy. A methodological study comparing flow cytometry of nuclear DNA content and in vivo and in vitro uptake of thymidine. 712 1

Plasma membrane from the brush border isolated from the tegument of Hymenolepis diminuta contains membrane-bound ribonuclease (RNase) and alkaline phosphatase activities. RNase (yeast RNA substrate), alkaline phosphatase (p-nitrophenyl phosphate substrate), and additional membrane proteins were solubilized by sonication or treatment with the detergents dodecyl trimethylammonium bromide, beta-octyl-D-glucopyranoside, sodium dodecyl sulfate (SDS), or ZwittergentTM 3-12 (N-dodecyl-N,N-dimethyl-3-ammonio-1-propanesulfonate). At optimal conditions, greater than 90% of both enzymes and total protein were solubilized by the latter two detergents, whereas beta-octyl-D-glucopyranoside, dodecyl trimethylammonium bromide, and sonication were only partially effective. Nonionic detergents did not solubilize the membrane effectively.
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PMID:Solubilization of membrane-bound ribonuclease (RNAse) and alkaline phosphatase from the isolated brush border of Hymenolepis diminuta (Cestoda). 739 87

Combined action of polyornithine and lecithin modified tobacco mosaic virus (TMV) virions making them sensitive to ribonuclease (RNase), pronase or Triton X-100. Sedimentational analysis and examination of the fluorescence spectrum revealed that the reaction product obtained after RNase treatments of modified TMV was a three-component complex made of coat protein, polyornithine and lecithin. The minimum requirement for the modification was completely fulfilled by cetyltrimethylammonium bromide, suggesting that a positively charged nitrogen group and an alkyl group of moderate size, C10--18, are necessary components. These components react with the surface region of TMV which is considered to have an important role in connecting coat protein subunits in TMV virions.
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PMID:Modification of tobacco mosaic virus by polyornithine and lecithin. 741 96

Five muscarinic receptor genes (m1-m5) that encode distinct muscarinic receptor subtypes have been cloned. Because of their structural homology and pharmacological similarity, ligand binding probes currently available do not clearly distinguish among the subtypes. To obtain a clear distribution within the CNS of molecularly defined muscarinic receptor subtypes, seven brain regions were examined for the expression of the respective mRNAs. The most sensitive method for detecting mRNA is through amplification of the respective cDNAs. Brain regions were obtained from male Wistar rats, and total RNA was isolated. The isolates were extensively treated with RNase-free DNase to remove any residual genomic DNA. Total RNA (1 microgram) was reverse-transcribed using random primers and reverse transcriptase. The resulting cDNA was amplified using a thermal cycler, and the polymerase chain reaction (PCR)-amplified products were analyzed by gel electrophoresis containing ethidium bromide and visualized with fluorescent illumination. PCR-amplified samples were also injected directly onto an HPLC anion exchange column and quantified by UV detection. Each of the five muscarinic subtypes was found in every brain region examined. The m1 subtype was most abundant in cortex and gradually declined in content caudally to the spinal cord. The m2 subtype was most abundant in thalamus-hypothalamus and ponsmedulla. The m4 subtype was found in greatest amount in the striatum, whereas m3 and m5 were expressed consistently throughout the CNS. The combination of RT-PCR and HPLC provides a rapid and sensitive method for quantifying the expression of mRNA coding for all five muscarinic receptor subtypes derived from the CNS.
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PMID:m1-m5 muscarinic receptor distribution in rat CNS by RT-PCR and HPLC. 751 60

A method for detecting the activity of ribonuclease inhibitors (RIs) after nondenaturing polyacrylamide gel electrophoresis and isoelectric focusing was developed. Both types of electrophoresis were performed using vertical slab polyacrylamide gels in the presence of dithiothreitol and in a sealed system. In each system, purified 50 kDa human RI was visualized as a single band by immunoblotting with a specific antibody. RI activity in the polyacrylamide gel slab was detected by sandwiching the gel slab between a cellulose acetate membrane moistened with a solution of bovine pancreatic ribonuclease A and a dried agarose film sheet containing substrate yeast RNA plus ethidium bromide and incubating at 37 degrees C. The ribonuclease penetrated the polyacrylamide gel and digested the substrate RNA in the agarose film. However, if an RI was present in the gel, the enzyme was inactivated by complex formation. Fluorescent bands corresponding to RIs were observed on a dark background under ultraviolet light. This activity staining had a high sensitivity allowing detection of less than 0.6 units of mammalian RIs (corresponding to 5 ng of purified human RI) and produced a sharp band which compared favorably with that obtained on immunoblotting. These electrophoretic techniques appear useful for the investigation of RIs in heterogeneous biological samples.
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PMID:Activity staining of mammalian ribonuclease inhibitors after electrophoresis in sealed vertical slab polyacrylamide gels. 754 54

Ribonucleases are widely found on the tissues of living organisms, but the functions of individual ribonucleases are not clear. To facilitate characterization of individual ribonucleases, I have developed a rapid method to separate and identify each ribonuclease from a crude sample by gel electrophoresis instead of by time-consuming purification steps. The ribonucleases in a crude sample are first separated by RNA-cast SDS-polyacrylamide gel electrophoresis and then eluted from the gel after ethidium bromide staining. To determine the base specificity of each ribonuclease, a 5' labelled oligonucleotide with known sequence is added to the enzyme eluate and the digested products are analyzed by denaturing gel electrophoresis. The base specificity of bovine pancreatic ribonuclease (RNase A), bullfrog oocyte-specific ribonuclease (RC-RNase), human serum ribonucleases and sweet potato leaf ribonucleases were determined by this method. Other properties of individual ribonucleases, e.g. substrate preference, may also be determined from crude samples by this method without further purification steps.
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PMID:Determination of base specificity of multiple ribonucleases from crude samples. 756 55

A novel S-alkylating reagent, N-(3-bromopropyl)-N,N,N',N',N'-pentamethyl-1,3-propanedi(ammonium bromide) (TAP2-Br) which carries two positive charges in the molecule, was prepared to increase the solubility or to decrease the hydrophobicity of cysteine-containing denatured proteins (or peptides). S-Alkylation with TAP2-Br introduces two positive charges per cysteine residue, which will effectively shift the net charge of a protein in the positive direction. Disulfide-containing proteins, such as hen egg-white lysozyme, RNase A, BSA, and soybean trypsin inhibitor (Kunitz type), were reduced and S-alkylated with TAP2-Br to evaluate the potential of this reagent compared with other S-alkylating reagents such as monoiodoacetic acid, bromosuccinic acid and (3-bromopropyl)trimethylammonium bromide. The solubilities of these denatured proteins in the pH range of 2-10 indicated that S-alkylation with TAP2-Br effectively solubilized not only basic proteins (lysozyme and RNase) but also an acidic protein containing a fairly large number of cysteine residues (BSA). Moreover, the retentions of cysteine-containing tryptic peptides derived from lysozyme on reversed-phase HPLC were greatly reduced by S-alkylation with TAP2-Br. These results indicate that TAP2-Br is very useful to increase the solubility of some cysteine-containing denatured proteins and to decrease the hydrophobicity of peptides containing cysteine residue(s).
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PMID:An S-alkylating reagent with positive charges as an efficient solubilizer of denatured disulfide-containing proteins. 788 61


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