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)

We investigated the ribonucleolytic breakdown of poly(U), poly(A), RNA trascribed from calf thymus DNA with E. coli RNA polymerase, ribosomal RNA, tRNA and mengovirus RNA by an enzyme fraction obrained from a postribosomal supernatant of Ehrlich ascites tumor cells. The single-stranded homopolyribonucleotides are preferentially degraded by the enzyme fraction with the production of ribonucleoside 5'-monophosphates. The RNase activity is completely dependent on the presence of Mg2+ ions and is highest at Mg2+ and K+ concentrations optimal for cell-free protein synthesis. Ribonucleoside 5'-monophosphates, ribonucleoside 2'(3')-monophosphates, ribonucleoside 2'(3'),5'-bisphosphates and transition state analogs consisting of vanadyl sulfate and either ribonucleosides or ribonucleoside 5'-monophosphates in a molar ratio 1:1 inhibit the ribonucleolytic activity of the enzyme fraction. The ribonucleoside 2'(3'),5'-bisphosphates and the transition state analogs are the most effective inhibitors. However, only in the presence of ribonucleoside 2'(3'),5'-bisphosphates a concomitant stimulation by 50 to 60% of poly(U)-directed polyphenylalanine synthesis is observed; all the other RNase inhibitors tested also inhibit polypeptide synthesis. The results of preliminary experiments show that poly(U) and ribonucleoside 2'(3'),5'-bisphosphates are well suited as ligands for affinity chromatography of ribonucleases from Ehrlich ascites tumor cells.
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PMID:Inhibition of ribonucleases by ribonucleotides and transition state analogs in cell-free extracts from Ehrlich ascites tumor cells. 32 84

A nuclease (RNase D) that can recognize structurally altered transfer RNA molecules has been partially purified from Escherichia coli. The enzyme acts poorly on intact tRNA and is inactive with the synthetic polyribonucleotides, poly(A), poly(U), or double-stranded poly(A).poly(U). The enzyme requires Mg2+ for activity and is stimulated by the monovalent cations, K+ and NH4+. The products of the reaction are 5'-mononucleotides. The molecular weight of the protein is about 60,000 as judged by Sephadex G-100 chromatography. The enzyme does not correspond to any known E. coli ribonuclease and may represent an intracellular scavenging mechanism for denatured tRNAs and other inactive RNA molecules.
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PMID:Identification of an Escherichia coli nuclease acting on structurally altered transfer RNA molecules. 34 22

The synthetic tRNA precursors, tRNA-C-114C]U and tRNA-C-C-A-[14C]C-C, as well as poly (a) and diesterase-treated tRNA, have been used to identify and purify potential 3'processing nucleases. Four activities have been separated by this analysis; and three of them have been characterized. Two of the enzymes, which are well-separated on hydroxylapatite columns, act on poly(A), require K+ and Mg2+ for activity, and have molecular weights of about 90,000. These activities have properties previously ascribed to RNase II. The third enzyme does not act on poly(A), requires Mg2+ for activity, and has a molecular weight of about 60,000. It is identical to RNase D, previously characterized as an exonuclease acting on tRNAs with altered structure. Each of the enzymes can remove nucleotides from the tRNA precursor containing extra nucleotides beyond the 3'terminus, whereas they are relatively inactive with intact tRNA or tRNA-C-U. The greatest specificity was displayed by RNase D. The possibility that RNase D is a 3'processing nuclease is discussed.
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PMID:Purification of potential 3' processing nucleases using synthetic tRNA precursors. 36 19

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

A second major species of leucine tRNA, tRNA Leu UAG (formerly designated tRNA Leu CUA) was purified from baker's yeast in a three-step procedure entailing BD-cellulose chromatography in the presence and absence of Mg2+ and Sephadex G-100 gel filtration. Results of aminoacylation and partial RNase T1 digestion experiments showed that this tRNA retains a native conformation under conditions that denature yeast tRNA Leu m5CAA (tRNA3 Leu). The primary structure of baker's yeast tRNA Leu UAG was elucidated by application of sensitive radioactive isotope derivative ("postlabeling") methods. Complete RNase T1 and A and partial RNase U2 fragments, prepared from non-radioactive tRNA and 5'-half and 3'-half molecules, were separated by two-dimensional polyethyleneimine-cellulose anion-exchange thin-layer chromatography and isolated by a novel micropreparative procedure affording high yields of these compounds in sufficient purity for subsequent tritium derivative analysis. Base composition and sequence of oligonucleotides were analyzed by tritium derivative methods. Molar ratios of the fragments were determined from the radioactivity of 3H-labeled nucleoside trialcohols in combination with base analysis. 2'-O-Methylated guanosine was characterized using the [gamma-32P]ATP/polynucleotide kinase reaction. The analysis of classical complete and partial RNase digests by the tritium derivative methods yielded the complete nucleotide sequence of the tRNA. A total of about 20 A260 units of the RNA was used for analysis, i.e. considerably less material than required for conventional spectrophotometric analysis. A different sequencing approach, consisting of a combination of "readout sequencing" with tritium sequencing of complete RNase T1 and A fragments, was applied to the 3'-half molecule. The 3'-half molecule was labeled with 32P at its 5' terminus, partially degraded with RNase T1, U2, and Phy1 and with alkali, and subjected to polyacrylamide gel electrophoresis. The sequence was read off the gel on the basis of cleavage patterns and size of the fragments. While the readout procedure provided only the positions of A, U, C, and G residues in the chain, additional information from tritium derivative analysis was utilized to define the positions of the modified nucleosides. The readout sequencing procedure was found to require less than 0.01 A260 unit of RNA and the analysis of the complete fragments about 6 A260 units. Interesting structural features of tRNA Leu UAG are (a) the location of unique, leucine tRNA iso-acceptor-specific sequences next to U-8, a constant nucleotide participating in synthetase recognition, (b) the occurrence of 1-methyladenosine in the T loop, a modification not present in the structurally related tRNA Leu m5CAA, and (c) the unusual presence of an unmodified uridine in the first position of the anticodon, which may be related to the unusual coding properties reported for this tRNA.
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PMID:Yeast tRNA Leu UAG. Purification, properties and determination of the nucleotide sequence by radioactive derivative methods. 37 75

The early stages of thermal unfolding of the tertiary structure of yeast tRNAPhe have been followed, in the presence and absence of Mg2+, by measuring changes in the chemical accessibility of the bases uracil and guanine. The reagent used in these studies is 1-cyclohexyl 3-[2-morpholino(4)-ethyl]carbodiimide methotosylate. 32P-labelled tRNA was used so that the points of modification could be examined with ribonuclease digestion and established fingerprinting techniques. Two regions of protection of Mg2+ have been found. One is within the oligonucleotide U8-A-m2G10 and the other is in the vicinity of residue U-59. The tertiary interactions and the D stem are the most readily melted parts of the teritary structure. In the absence of Mg2+ the region of U-59 is the first part of the tertiary structure to become accessible to the reagent. This is closely followed by the opening up of the 'wobble' G-U base pair in the aminoacyl stem. Most of the triple interactions in the augmented D helix are also disrupted early in the melting. The region of intricate interactions between the invariant G-G part of the D loop and the T-psi-C-G loop contains the most stable set of tertitary structure interactions.
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PMID:Initial stages of the thermal unfolding of yeast phenylalanine transfer RNA as studied by chemical modification: the effect of magnesium. 41 74

Studies using isogenic transductant strains mlpA+ and mlpA as well as reversion analysis suggested that the physiological consequences of a structural gene mutation in murein lipoprotein include (i) increased sensitivity toward chelating agents ethylenediaminetetraacetic acid and ethyleneglycol-bis (beta-aminoethyl ether)-N,N-tetraacetic acid, (ii) leakage of periplasmic enzyme ribonuclease, (iii) weakened association between the outer membrane and the rigid layer accentuated by Mg2+ starvation, resulting in the formation of outer membrane blebs, and (iv) decreased growth rate in media of low ionic strength or low osmolarity. It is suggested that the bound form of lipoprotein plays an important role in the maintenance of the structural integrity of the outer membrane of the Escherichia coli cell envelope. Other outer membrane components may also contribute to the anchorage of outer membrane to the rigid layer, probably through ionic interactions with divalent cations. Using the phenotype of ribonuclease leakage as an unselected marker in a three-factor cross with P1 transduction, we were able to establish the gene order of man mlpA aroD pps on the E. coli chromosome.
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PMID:Physiological characterization of an Escherichia coli mutant altered in the structure of murein lipoprotein. 41 67

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

The Mg2+-precipitable polysomes in experimental granulation tissue increase up to three weeks and decrease slowly thereafter. Polysomes from young granulation tissue are, on a weight basis, more active in cell-free protein synthesis than polysomes from old granulation tissue. RNase activity is highest in polysomes from 1-week tissue. The polysomes contain both acid and alkaline RNase, but the alkaline RNase is the latent form, manifested by the addition of p-chloromercuribenzoate (pCMB). The high RNase activity of granulation tissue renders the analysis of polysomal patterns and functions of the polysomes difficult. In the post-polysomal supernatant of granulation tissue the RNase activity is highest in 2-4 week old tissue; it is maximal at pH 7-8 and only slightly influenced by pCMB.
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PMID:Ribonuclease activities in developing experimental granulation tissue with reference to polysomes. 50 66

Myosin purified from Dictyostelium amoebae has approximately 10% by weight of RNA associated with it, unless specific steps (DEAE cellulose chromatography or RNase digestion) are taken to remove it. This RNA has significant effects on the structural states formed by the myosin at low ionic strength in the presence of Mg2+. Rapid precipitation of RNA-free myosin by dilution generates bipolar thick filaments (540 nm long, 33 nm thick), often with a bare zone and a 15-nm transverse repeat. Rapid precipitation of myosin with copurified RNA yields linear aggregates of bipolar filaments, showing some lateral association. Slow precipitation of RNA-free myosin by dialysis yields very long filaments or ribbons (greater than 5 micrometer, 30--60 nm wide) in which the myosin may be packed diagonally across the filament, similar to the "side-polar" aggregates formed by other nonmuscle myosins and by smooth muscle myosin (Craig R, Megerman J: J Cell Biol 75:990, 1977; Hinssen H, D'Haese J, Small JV, Sobieszek A: J Ultrastruct Res 64:282, 1978). Slow precipitation of myosin with copurified RNA generates linear filaments with repeat intervals of 290 and 650 nm. Other polyanions were tested for their effects on myosin aggregation. Total RNA and ribosomal RNA from Dictyostelium, when added to RNA-free myosin, also induced the extensive linear aggregation seen with the copurified RNA/myosin complex, although higher concentrations of RNA were required to obtain quantitatively the same effect. DNA and heparin were also effective inducers of linear aggregation, whereas homopolymers of nucleotides and of acidic or basic amino acids were poorly effective.
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PMID:Structural states of dictyostelium myosin. 54 31


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