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

The substrate specificity of pancreatic ribonuclease A is discussed in light of observations based on accurate X-ray structure analysis of several enzyme-nucleotide complexes. A hypothesis for protein-nucleic acid recognition is presented which proposes that: (a) pyrimidine bases in RNA are recognised by ribonuclease due to the charge complementarity of two groups (the amide nitrogen and the side chain oxygen (OG) of threonine 45) of the protein and relevant atoms in the heterocyclic base (O2 and N3 in pyrimidine nucleotides); (b) interaction of the protein with the ribose moiety of the nucleotides is non-specific; and (c) conformational flexibility in the region of the scissile P-O bond is provided by different locations of the phosphoryl oxygens, rather than by an overall translation of the phosphate moiety.
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PMID:Enzyme specificity: base recognition and hydrolysis of RNA by ribonuclease A. 619 18

The modified purine nucleotide 8-oxo-guanosine-2'-phosphate binds at the pyrimidine binding site of ribonuclease-A. The O8-2'GMP inhibitor is in a syn conformation, with an intramolecular hydrogen bond between the N-3 atom of the base and the O-5' atom of the ribose. The essential groups of the protein involved in base recognition are O gamma 45 and N-45, which form hydrogen bonds to the five-membered ring of the heterocyclic base. Mobility of enzyme side-chains (viz. Lys41, Lys66, His119) close to the catalytic cleft of the protein allows conformational flexibility in the substrate binding region of ribonuclease-A. Inhibitor binding alters the solvent structure of the protein but the overall shape of the enzyme is not effected.
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PMID:Specificity of pancreatic ribonuclease-A. An X-ray study of a protein-nucleotide complex. 631 34

The pH, ionic strength, and solvent deuterium isotope dependence of the steady-state kinetics of the ribonuclease A catalyzed hydrolysis of cytidine cyclic 2',3'-phosphate has been investigated by using, primarily, the technique of flow microcalorimetry to monitor the kinetics. The pH dependence of the Michaelis-Menten parameters has been analyzed by assuming the participation of His-12 and -119 of the enzyme and a third ionizing group, postulated to be on the pyrimidine ring of the substrate, to determine the pH-independent rate constant kc, and Michaelis constant Km. The reported pH analysis, together with existing NMR data and chemical modification studies, allows an assignment of the functional roles of His-12 and -119 as being those of general acid and general base catalytic residues, respectively. At high pH, the apparent Km value is found to increase to unity. This drop in affinity between the enzyme and the substrate at high pH indicates that the substrate binds to the enzyme primarily through an electrostatic interaction with the active-site histidine residues, particularly His-12. The apparent absence of an interaction with the riboside portion of the substrate is suggested to be due to the fact that the substrate exists in a syn conformation about its glycosidic bond and thus cannot interact optimally with the enzyme's binding pocket. This will result in a relative destabilization of the enzyme-substrate complex, which can then be relieved upon the formation of the transition state. The ionic strength dependence of ribonuclease activity is shown to be primarily a result of its effect on the pKa of the histidine residues and a concomitant change in the value of Km.
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PMID:Energetics of ribonuclease A catalysis. 1. pH, ionic strength, and solvent isotope dependence of the hydrolysis of cytidine cyclic 2',3'-phosphate. 631 13

The results of X-ray analyses of three ribonuclease-A-nucleotide complexes, at 2.3 A, are reported. A modified purine mononucleotide, 8-oxo-guanosine 2'-phosphate in a syn conformation, binds at the pyrimidine-binding site of the catalytic cleft. Solvent molecules are expelled from the active site due to inhibitor binding. The positions of the side-chains of the active-site residues Gln-11, His-12 and Thr-45 are well defined and do not alter on inhibitor binding. The mobility of Lys-41 is greatly reduced in the protein-nucleotide complexes and the terminal amino group interacts directly with the 2'-phosphate group of the nucleotides. In the complex of the enzyme with a modified pyrimidine, cytidine-N(3)-oxide 2'-phosphate, His-119 is stabilised in the minor site of the native protein [Borkakoti, N., Moss, D.S. and Palmer, R.A. (1982) Acta Crystallogr. B38, 2210-2217], while in the protein-purine derivative the imidazole group is located in the major site. Inhibitor binding induces movements in the side-chains of Lys-7 and Lys-66 which also modify the conformation of the active-site cleft of ribonuclease A.
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PMID:The active site of ribonuclease A from the crystallographic studies of ribonuclease-A-inhibitor complexes. 640 32

A new monovalent and divalent requiring endoribonuclease has been identified in extracts of calf heart tissue using a natural substrate molecule. This ribonuclease creates 5' hydroxyl and 3' phosphate groups in its cleavage products. It cleaves predominantly between pyrimidine and A residues though not all such possible sites are attacked in the substrates tested.
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PMID:Properties of a monovalent and divalent requiring endoribonuclease with novel specificity from calf heart. 640 82

The synthesis of RNA catalysed by RNA polymerase from Escherichia coli is terminated at specific sites on DNA templates through the action of a multimeric basic protein known as rho (refs 1, 2). Three lines of evidence suggest that an interactions of rho with the nascent RNA is important for this termination. First, rho binds strongly to RNA; second, rho expresses an RNA-dependent ATPase activity which is essential for termination; third, RNA polymerase does not terminate RNA synthesis at rho-dependent sites when the nascent RNA is digested by ribonuclease during transcription. From the fact that certain RNAs, particularly single-stranded, pyrimidine-rich polymers containing at least 10% cytidylate residues, are more effective than other RNAs at promoting rho-ATPase, it has been proposed that rho recognises specific sites oion on a mRNA transcribed from bacteriophage lambda DNA.
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PMID:A rho-recognition site on phage lambda cro-gene mRNA. 644 43

A ribonuclease (RNase) has been isolated from normal human pancreas obtained upon autopsy. About 5 mg of RNase is normally recovered per kilogram of pancreas, equivalent to ca. 70% of the total activity and a 700-fold purification from the initial acidified extract. The specific activity of the purified enzyme is identical with that of bovine pancreatic ribonuclease, and a single component is found in sodium dodecyl sulfate-polyacrylamide gel electrophoresis, gel filtration, and reversed-phase high-pressure liquid chromatography. Aggregation of the protein is found upon ultracentrifugation under native and denaturing conditions, and several bands of equal specific activity are seen in polyacrylamide gel electrophoresis of the native protein. At least two components are glycoproteins. A molecular weight of 15 000 is estimated from sodium dodecyl sulfate gel electrophoresis, gel filtration, and amino acid and peptide analyses. The enzyme is related to bovine pancreatic RNase, but distinguishable by amino acid analysis, tryptic peptide maps, and low cross-reactivity of antibodies with the heterologous enzymes. The human enzyme is also inactivated by treatment with iodoacetic acid at pH 5.5 and is essentially identical with bovine RNase in its far-ultraviolet circular dichroism spectrum. The human RNase is like bovine pancreatic RNase catalytically; RNA is cleaved at pyrimidine residues, and activity against poly(cytidylic acid) is high.
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PMID:Purification and characterization of human pancreatic ribonuclease. 678 51

Partially fragmented 12-21S rat liver messenger ribonucleoprotein (mRNP), labelled either with [3H]-orotic acid or [3H]-adenine was treated with 5 (micrograms/ml or 0.1 microgram/ml pancreatic ribonuclease (EC 3.1.27.5) and the resistant fragments were separated by sucrose gradient centrifugation. Two types of fragments were obtained. Digestion of mRNP with ribonuclease at a concentration of 5 micrograms/ml resulted in 9S poly(A)-protein particles of mRNP as evidenced by their characteristic sedimentation, electrophoretic mobility of the RNA moiety and protein composition. In contrast, ribonuclease at a concentration of 0.1 microgram/ml produced 2-5S pyrimidine labelled fragments carrying polynucleotide sequences consisting of approximately 16-50 residues. Two polypeptides of rat liver mRNP with molecular weights of 38 000 (P38) and 44 000 (P44) were found to be attached to these sequences. The data demonstrate that RNA sequences other than poly(A) interact with protein in mRNP. Electron microscopic studies revealed that the liberation of mRNP from the polysomes by EDTA changes its surface properties since EDTA releases the mRNP mainly in the form of globular particles. However, a small proportion of the mRNP remains in fibril-like form. Among the fibrils several blobs could be seen which were probably the proteins attached to the mRNA. From the available data a model for the structure of an "average" mRNP molecule is proposed.
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PMID:Studies on the structure of rat liver messenger ribonucleoprotein. II. Non-poly(A) RNase resistant fragments and electron microscopic appearance of mRNP. 733 82

We have purified a high molecular weight ribonuclease (hmRNase) from human milk by a two-step column chromatographic procedure and characterized the enzyme. The molecular mass of hmRNase is 80 kDa as determined from SDS-polyacrylamide gel electrophoresis. The pH optimum of the enzyme is in the range of 7.5-8.0, similar to other secretory RNases. hmRNase is pyrimidine-specific and cleaves the phosphodiester bond 3' to a pyrimidine residue. It selectively degrades the pyrimidine strand in poly(rA):poly(rU) and poly(dA):poly(rU) double stranded substrates. The extent of degradation for naturally occurring RNAs vary in the order tRNA < rRNA < mRNA at low enzyme concentrations. hmRNase shows allosteric behavior with positive cooperativity in its reaction on polynucleotide substrates. The activity of the enzyme is enhanced in the presence of monoribonucleotides. Antiserum obtained against purified hmRNase did not cross-react with low molecular weight RNase which is also present in milk. In addition, an immunologically cross-reacting species could not be detected in the serum, suggesting the origin of hmRNase in the mammary gland but not blood.
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PMID:Purification and characterization of a high molecular weight ribonuclease from human milk. 768 36

To determine the localization of the pyrimidine-guanine sequence-specific ribonuclease in Rana catesbeiana (bullfrog) oocytes, the RNase was first isolated and used to prepare a specific rabbit antiserum. Only one protein of similar molecular size to the RNase was immunoprecipitated from ovary homogenate by the antiserum, but two bands were observed by Western blotting analysis. These two proteins were shown by further purification of antibody and Western blotting analysis to have similar antigenicity. Immunoprecipitation and Western blotting of tissue homogenates showed that the RNase was found predominantly in the ovary, but not in other tissues. The specific localization of the RNase was determined by immuno-electron microscopy of oocyte sections incubated with the specific antiserum; the yolk granules, but not other organelles, were found to contain the RNase. Most of the RNase was evenly distributed in the lateral amorphous area of the yolk granule but not in the central yolk crystal area which contains stored vitellogenin proteins. Our results indicate that the RNase is compartmentalized in the yolk granules of oocytes, which might prevent damage to cellular RNAs.
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PMID:Immunocytochemical localization of ribonuclease in yolk granules of adult Rana catesbeiana oocytes. 778 Oct 23


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