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:3.1.26.9 (ribonuclease)
6,589 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A procedure for preparing polyribosome aminoacyl-tRNA free from contamination by supernatant aminoacyl-tRNA and free amino acids is described. Important features of the procedure are the use of acidic buffers to help protect the amino acid-tRNA linkage and the inclusion of sodium dodecyl sulphate, to inhibit ribonuclease activity. The specific radioactivity of polyribosome aminoacyl-tRNA is high within 30s and reaches a maximum in 2 1/2 min, well ahead of polyribosome peptides which, as described by Herrmann et al. (1971), attain maximum specific radioactivity in about 10 min.
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PMID:Preparation of polyribosome aminoacyl-transfer ribonucleic acid from the muscle of chick embryos. 24 28

The separation of rat T lymphocytes was investigated on anti-Ig--Ig columns. A simple and efficient method for the purification of rat Ig by precipitation of rat serum with sodium sulfate is presented. Protein binding characteristics of glass and plastic beads, as solid support of affinity columns, are described, as well as optimal parameters for coating beads with rat Ig (with BSA, ribonuclease and lysozyme, as comparison). Binding of Ig was primarily dependent on the concentration of the Ig solution. Maximal strong binding of Ig (6.2 X 10(3) molecules per micron2 of bead surface) was reached a 400 microng per ml concentration of purified Ig solution during 20 min of incubation. Higher concentrations increased only the amount of loosely bound Ig on the surface of beads whereas the amount of firmly bound Ig remained unchanged. Fractionation of lymphoid cell suspensions on anti-Ig--Ig affinity columns prepared at optimal conditions resulted in highly purified T-cell suspensions containing less than 1% of lymphocytes bearing surface Ig receptors.
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PMID:Optimal conditions for the separation of rat T lymphocytes on anti-immunoglobulin--immunoglobulin affinity columns. 30 Nov 58

M protein was extracted from type 24, group A streptococci with pepsin at pH 5.8 and was further purified by ammonium sulfate precipitation, ribonuclease digestion, ion-exchange chromatography, and isoelectric focusing. The purified pepsin extract of M (pep M) protein was shown to be free of nontype-specific immunoreactivity in (a) complement fixation tests with heterologous M antiserum, (b) skin tests in normal adult guinea pigs, and (c) passive hemagglutination tests for the presence of lipoteichoic acid sensitizing or antigenic activity. The pep M24 was highly immunogenic; two of three rabbits developed opsonic antibody titers of 1:256 and the third a titer of 1:32 6 wk after a single injection of 100-pg doses of pep M24 emulsified in complete Freund's adjuvant. The antisera lacked nontype-specific antibodies and produced single precipitin lines in agar gel diffusion tests against crude HC1 extracts of the homologous M protein. Thus, the type-specific antigenic determinant(s) of type 24 M protein appears to be separable from immunotoxic, cross-reactive antigens without loss of immunogenicity in rabbits. The mobility of pep M24 upon electrophoresis in 10 percent sodium dodecyl sulfate pelyacrylamide gel was consistent with an average mol wt of 33,500 daltons. Amino acid analysis demonstrated a predominance of alanine, followed by glutamic acid, lysine, leucine, and aspartic acid. Pep M24 contained an estimated six to seven methionine residues and approximately ten phenylalanine residues per molecule. No other aromatic amino acids were detected. Automatic Edman degradation of pep M24 yielded the sequence of the first 29 amino acids (the amino terminal amino acid being valine) of the amino terminal region of the molecule. The detection of only one new amino acid at each step of Edman degradation confirmed the homogeneity of the purified pep M24.
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PMID:Purification and properties of M protein extracted from group A streptococci with pepsin: covalent structure of the amino terminal region of type 24 M antigen. 32 68

Heat-stable enterotoxin (ST) produced by porcine strains of enterotoxigenic (ENT+) Escherichia coli has been purified to apparent homogeneity by sequential ultrafiltration, acetone fractionation, preparative gel electrophoresis, diethylaminoethyl Bio-Gel A ion-exchange chromatography, and Bio-Gel P-10 gel filtration. The enterotoxin, purified more than 1,500-fold, exhibited a molecular weight of 4,400, as determined by both sodium dodecyl sulfate-gel electrophoresis and gel filtration. A molecular weight of 5,100, representing 47 residues, was calculated from amino acid analysis data. The amino acid content was distinctive, with an unusually high proportion of cystines and few hydrophobic amino acids. A single amino-terminal residue, glycine, was observed. Purified ST was stable to heating (100 degrees C, 30 min) and did not lose biological activity after treatment with Pronase, trypsin, proteinase K, deoxyribonuclease, ribonuclease, and phospholipase C. Periodic acid oxidation and several organic solvents (acetone, phenol, chloroform, and methanol) had no effect on the biological activity of ST. Further, purified ST was stable to acid treatment at pH 1.0 but lost biological activity at pH values greater than 9.0. Neither lipopolysaccharide nor lipid contamination was evident in purified preparations. A characteristic absorption spectrum was observed during the course of the purification, which shifted from a maximum at 260 nm in crude preparations to 270 nm for the purified toxin. Antiserum obtained from rabbits immunized with ST or ST coupled to bovine serum albumin neutralized the action of the enterotoxin in suckling mice; however, passive hemagglutination and hemolysis titer assays suggested that ST is a poor antigen.
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PMID:Purification and chemical characterization of the heat-stable enterotoxin produced by porcine strains of enterotoxigenic Escherichia coli. 34 81

The 5. 8S ribosomal RNA of bakers yeast contains one particularly stable hairpin helix which is isolated by partial T1 ribonuclease digestion. Thermal hyperchromism analysis of the hairpin fragment showed that it dissociates cooperatively with 18% hyperchromism, with a Tm of 83 degree C at 2.7 mM sodium ion concentration, and with a hyperchromic difference spectrum indicative of over 90% G + C content. The probable secondary structure for the fragment was used to predict a helix free energy, delta G = -16.2 kcal/mole, which was the same as that determined from the melting equilibrium. The predicted enthalpy however, was 77% of the value, delta H = -114 kcal/mole, determined from the van't Hoff relationship. The effect on these data of a G.U base pair within the 9 base pair helix is discussed.
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PMID:Thermodynamics of a stable yeast 5.8S rRNA hairpin helix. 36 14

The biosynthesis of bacteriophage T4 tRNAPro, tRNASer, and tRNAIle requires enzymatic removal of extra nucleotides from the 3' terminus of the respective precursor RNAs. A ribonuclease activity capable of catalyzing such reactions has been partially purified from uninfected Escherichia coli using an artificial precursor RNA as substrate. A number of ribonuclease activities were resolved during purification. Use of E. coli strain BN, a mutant known to be deficient in the relevant ribonuclease activity, permitted us to identify it in wild-type cells. This activity was designated the BN ribonuclease. BN ribonuclease had an apparent molecular weight of 35,000 as measured by Sephadex gel filtration. Mg2+ was required for activity, which was optimal at [Mg2+] of 2mM. Activity did not require monovalent cations K+ or Na+. BN ribonuclease was less efficient at removing extra residues in the biosynthesis of tRNASer and tRNAIle than in the biosynthesis of tRNAPro.
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PMID:An Escherichia coli ribonuclease which removes an extra nucleotide from a biosynthetic intermediate of bacteriophage T4 proline transfer RNA. 36 22

Two dimensional PEI-cellulose thin layer chromatography can resolve sequentially degraded oligonucleotide fragments of tRNA. This technique entails the sequential degradation of the oligonucleotide with snake venom phosphodiesterase in the presence of bacterial alkaline phosphatase, and periodate oxidation followed by tritiated sodium borohydride reduction of the 3' terminal nucleoside. Subsequently the tritiated oligonucleotide fragments were resolved by two dimensional PEI-cellulose TLC. The results of these experiments indicate that, in some cases, the complete nucleotide sequence of a large oligonucleotide fragment may be determined by interpretation of the observed mobility shifts, thereby eliminiating the need for additional analysis of the oligonucleotide. In addition, the use of two-dimensional rather than one-dimensional resolution of the tritium labeled fragments allows for a complete separation of any interfering background spots from the sequentially degraded oligonucleotides. This procedure was applied to the complete nucleotide sequence analysis of several ribonuclease T1Val and ribonuclease A digestion products from human placenta tRNA.
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PMID:A two-dimensional thin layer chromatographic procedure for the sequential analysis of oligonucleotides employing tritium post-labeling. 41 70

The characteristics of an in vitro polyuridylic acid dependent amino acid incorporating system prepared from germinating macroconidia of Microsporum canis are described. The incorporation of 14C-phenylalanine into polyphenylalanine is dependent on S-30 extract, adenosine triphosphate, magnesium ions and polyuridylic acid. Incorporation is slightly enhanced by yeast transfer ribonucleic acid and pyruvate kinase. The system is highly sensitive to ribonuclease, puromycin and miconazole (an antifungal agent), moderately sensitive to sodium fluoride and much less sensitive to phenethylalcohol, cycloheximide, chloramphenicol and deoxyribonuclease. Cell-free extract from ungerminated conidia has less capacity to synthesize the protein and during germination a marked increase in the protein synthetic activity is observed. The results from experiments wherein ribosomes and S-100 fraction from germinated and ungerminated spores are unterchanged, revealed that the defect in the extract from the ungerminated spore is in the ribosomes.
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PMID:Studies on the macroconidia of Microsporum canis. Characteristics of in vitro amino acid incorporating system. 42

Chloroplasts, isolated from the leaves of 7-day-old pea seedlings, were incubated in the light with [35S]methionine or [3H]leucine. After extraction from the washed chloroplast membranes using a mixture of ethyl acetate, ethanol and ammonia, cytochrome f was precipitated with a monospecific antiserum and resolved by gradient polyacrylamide gel electrophoresis in sodium dodecylsulphate. The cytochrome f band was identified by its intrinsic fluorescence in ultraviolet light and was shown to be radioactive by autoradiography or fluorography of dried polyacrylamide gel. One-dimensional peptide mapping of the products of papain hydrolysis confirmed that the radioactivity was an integral part of cytochrome f. The incorporation of [35S]methionine into cytochrome f was inhibited by D(-)threo-chloramphenicol but not by cycloheximide and did not occur in the dark. The synthesis was resistant to ribonuclease. It is concluded that cytochrome f is synthesised in intact isolated pea chloroplasts.
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PMID:Synthesis of cytochrome f by isolated pea chloroplasts. 46 51

Though DNase does not contain any cysteine residues, incubation of the enzyme with 2-nitro-5-thiocyanobenzoic acid in the presence of Ca2+ at pH values above 7.5 results in an irreversible inactivation of the enzyme. The inactivation also occurs when Ca2+ is replaced by Mg2+, but not in their absence. Amino acid analyses after acid hydrolyses of the completely inactivated ant the native enzymes show no significant differences in composition, including tryptophan and half-cystine residues. However, sodium dodecyl sulfate gel electrophoresis indicates enzyme cleavage by the treatment with 2-nitro-5-thiocyanobenzoic acid. This reagent does not inactivate chymotrypsin and lysozyme, and under conditions where bovine DNase is inactivated, does not inactivate other nucleases such as ribonuclease, snake venom phosphodiesterase, and spleen acid DNase. However, it inactivates malt DNase and can, therefore, be considered a specific inhibitor of DNase I. The inactivation kinetics is pseudo-first order, resembling Michaelis-Menten, with an affinity constant of 16.7 mM. It is the cyano group, not the thionitrobenzoic acid of 2-nitro-5-thiocyanobenzoic acid that reacts to form cyano-DNase.
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PMID:Inactivation of bovine pancreatic DNase by 2-nitro-5-thiocyanobenzoic acid. I. A novel inhibitor for DNase I. 48 54


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