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Query: EC:3.1.27.4 (ribonuclease)
 6,621 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The stochastic boundary molecular dynamics method is used to study the structure, dynamics, and energetics of the solvated active site of bovine pancreatic ribonuclease A. Simulations of the native enzyme and of the enzyme complexed with the dinucleotide substrate CpA and the transition-state analog uridine vanadate are compared. Structural features and dynamical couplings for ribonuclease residues found in the simulation are consistent with experimental data. Water molecules, most of which are not observed in crystallographic studies, are shown to play an important role in the active site. Hydrogen bonding of residues with water molecules in the free enzyme is found to mimic the substrate-enzyme interactions of residues involved in binding. Networks of water stabilize the cluster of positively charged active site residues. Correlated fluctuations between the uridine vanadate complex and the distant lysine residues are mediated through water and may indicate a possible role for these residues in stabilizing the transition state.
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PMID:Active site dynamics of ribonuclease. 386 34

The terminal fragment obtained by hydrolysis with ribonuclease T(1) of the ribonucleic acid from the bacteriophage Qbeta has been isolated and purified. The results of chemical and enzymatic hydrolysis of this fragment and of the intact RNA itself indicate that the fragment has the composition (10 Cp, 4 Up), and that the RNA has the terminal sequence -GP(9 Cp, 4 Up) CpA. These conclusions are supported by the results of an application of the sedimentation-equilibrium method in which the molecular weight of the Qbeta fragment was compared with that of the corresponding fragment from f2 phage RNA for which the terminal sequence, -GpUpUpApCpCpApCpCpCpA had previously been determined.
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PMID:Comparison of RNA terminal sequences of phages f2 and Q-beta: chemical and sedimentation equilibrium studies. 565 59

A nucleolar ribonuclease specific for single-stranded ribonucleic acid (RNA) has been isolated and extensively purified from Ehrlich ascites carcinoma cells. The enzyme is optimally active at neutral pH and degrades RNA via a 2',3'-cyclic intermediate leaving 3'- or 2',3'-cyclic terminated oligonucleotides. The ribonuclease has an apparent molecular weight of 38 500 as judged by sedimentation equilibrium and is a basic protein having an isoelectric point greater than 9.0. The enzyme preferentially cleaves poly(C) over poly (U), poly(A), or poly(C).poly(I). Limit digestion products of poly(C) degratation are on the average tri-, tetra-, and pentanucleotides. In the partial digestion of yeast 5.8S rRNA, the nucleolar ribonuclease cleaves only CpA phosphodiester bonds. Spermidine, spermine, and histone I inhibit the activity of nucleolar ribonuclease. Antibodies directed toward pancreatic RNase do not cross-react with the Ehrlich nucleolar ribonuclease.
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PMID:Properties of a purified nucleolar ribonuclease from Ehrlich ascites carcinoma cells. 739 14

With the use of a high yield prokaryotic expression system, large amounts of human eosinophil cationic protein (ECP) have been obtained. This has allowed a thorough kinetic study of the ribonuclease activity of this protein. The catalytic efficiencies for oligouridylic acids of the type (Up)nU>p, mononucleotides U>p and C>p, and dinucleoside monophosphates CpA, UpA, and UpG have been interpreted by the specific subsites distribution in ECP. The distribution of products derived from digestion of high molecular mass substrates, such as poly(U) and poly(C), by ECP was compared with that of RNase A. The characteristic cleavage pattern of polynucleotides by ECP suggests that an exonuclease-like mechanism is predominantly favored in comparison to the endonuclease catalytic mechanism of RNase A. Comparative molecular modeling with bovine pancreatic RNase A-substrate analog crystal complexes revealed important differences in the subsite structure, whereas the secondary phosphate-binding site (p2) is lacking, the secondary base subsite (B2) is severely impaired, and there are new interactions at the po, Bo, and p-1 sites, located upstream of the P-O-5' cleavable phosphodiester bond, that are not found in RNase A. The differences in the multisubsites structure could explain the reduced catalytic efficiency of ECP and the shift from an endonuclease to an exonuclease-type mechanism.
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PMID:Kinetic and product distribution analysis of human eosinophil cationic protein indicates a subsite arrangement that favors exonuclease-type activity. 1033 57

Kinetic parameters of cleavage of CpA and UpA sequences in an oligoribonucleotide under the action of artificial ribonuclease ABL3C1 were measured. The compounds were built of RNA-binding domain B, catalytic fragment C, linker L3 comprising 3 methylene groups, and aliphatic fragment A. The rate of cleavage of phosphodiester bonds in CpA sequence within decaribonucleotide UUCAUGUAAA was shown to be 3.4 +/- 0.2 times higher than in UpA sequence. The rate of cleavage of phosphodiester bonds were found to depend on substrate length: a thousandfold increase in cleavage rate constant was observed for CpA sequence in decaribonucleotide as compared with diribonucleotide monophosphate CpA. A slight decrease in the cleavage rates was observed for the reactions proceeding in different buffers at pH 7.0: imidazole > HEPES > phosphate > cacodylate. At the same time, the ratio of cleavage rates for CpA and UpA sequences remained constant.
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PMID:[Kinetic parameters of hydrolysis of CpA and UpA sequences in an oligoribonucleotide by compounds functionally mimicking ribonuclease A]. 1250 May 46

Cleavage of phosphodiester bonds by small ribonuclease mimics within different bulge-loops of RNA was investigated. Bulge-loops of different size (1-7 nt) and sequence composition were formed in a 3' terminal fragment of influenza virus M2 RNA (96 nt) by hybridization of complementary oligodeoxynucleotides. Small bulges (up to 4 nt) were readily formed upon oligonucleotide hybridization, whereas hybridization of the RNA to the oligonucleotides designed to produce larger bulges resulted in formation of several alternative structures. A synthetic ribonuclease mimic displaying Pyr-Pu cleavage specificity cleaved CpA motifs located within bulges faster than similar motifs within the rest of the RNA. In the presence of 10 mM MgCl2, 75% of the cleavage products resulted from the attack of this motif. Thus, selective RNA cleavage at a single target phosphodiester bond was achieved by using bulge forming oligonucleotides and a small ribonuclease A mimic.
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PMID:Enhanced RNA cleavage within bulge-loops by an artificial ribonuclease. 1573 40

A recombinant ribonuclease, cusativin, was characterized for its cytidine-specific cleavage ability of RNA to map chemical modifications. Following purification of native cusativin protein as described before (Rojo et al. Planta 194:328, 17), partial amino acid sequencing was carried out to identify the corresponding protein coding gene in cucumber genome. Cloning and heterologous expression of the identified gene in Escherichia coli resulted in successful production of active protein as a C-terminal His-tag fusion protein. The ribonuclease activity and cleavage specificity of the fusion protein were confirmed with a variety of tRNA isoacceptors and total tRNA. Characterization of cusativin digestion products by ion-pairing reverse-phase liquid chromatography coupled with mass spectrometry (IP-RP-LC-MS) analysis revealed cleavage of CpA, CpG, and CpU phosphodiester bonds at the 3'-terminus of cytidine under optimal digestion conditions. Ribose methylation or acetylation of cytosine inhibited RNA cleavage. The CpC phosphodiester bond was also resistant to cusativin-mediated RNA cleavage; a feature to our knowledge has not been reported for other nucleobase-specific ribonucleases. Here, we demonstrate the analytical utility of such a novel feature for obtaining high-sequence coverage and accurate mapping of modified residues in substrate RNAs. Graphical abstract Cytidine-specific novel ribonuclease activity of cusativin.
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PMID:Novel ribonuclease activity of cusativin from Cucumis sativus for mapping nucleoside modifications in RNA. 2873 Mar 4