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)

Four urinary alkaline ribonucleases (RNase, EC 3.1.4.22) were purified from patients with nephrotic syndrome using phosphocellulose, DEAE-cellulose and Sephadex G-75 chromatographiy. These enzymes were designated as RNases 1--4, respectively, in order of elution on phosphocellulose chromatography. The respective purification of each fraction was 41-, 23-, 34- and 27-fold with a total recovery of 25%. The pH optima of these RNases were around 8.5 with Tris/HCl buffer and the reaction was activated by mono- and divalent cations, such as Na+, K+, Mg2+ and Ca2+, but inhibited by Fe2+, Cu2+ and Zn2+. EDTA had little effect on the velocity of reaction. The molecular weights of RNases 1--4 were estimated by gel filtration as 45 000, 32 000, 20 000, and 13 000, respectively. Each enzyme hydrolyzed pyrimidine nucleotides preferentially with higher affinity for poly(C) than poly (U) as determined with synthetic polymers and was free from other nucleolytic enzymes. The patients with renal disorders excreted one to four RNases in urine and the number of enzymes increased as the concentration of urinary protein increased. On the other hand, normal subjects excreted a single fraction essentially identical to RNase 1.
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PMID:Purification and properties of urinary alkaline ribonucleases from patients with nephrotic syndrome. 1 98

The DNAase in human urine was purified about 30-fold with a recovery of 28%. This involved DEAE-cellulose and phosphocellulose chromatography steps and gel filtration on Sephadex G-75. The enzyme required divalent cations such as Co2+, Mg2+, Mn2+ and Zn2+ for activity, but Ca2+, Cu2+ and Fe2+ were ineffective. EDTA and G-actin inhibited the reaction. The maximum activity was observed at pH 5.5 in acetate buffer plus Co2+ or Mg2+ and Ca2+. It had a molecular weight of approximately 38 000, estimated by gel filtration on Sephadex G-75 and isoelectric point of around pH 3.9. The enzyme is an endonuclease which hydrolyzes native, double-stranded DNA about 3 to 4 times faster than thermally denatured DNA to produce 5'-phosphoryl- and 3'-hydroxyl-terminated oligonucleotides. The final preparation was free of non-specific acid and alkaline phosphatases, phosphodiesterase and ribonuclease activities.
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PMID:Purification and properties of deoxyribonuclease from human urine. 2 31

Acid RNase was purified from normal human serum about 2400-fold by chromatography on phosphocellulose and Sephadex G-75 and rechromatography on Sephadex G-75. Assayed with yeast RNA as substrate, the enzyme showed the maximal activity at about pH 6.5 with sodium phosphate buffer. The reaction was activated by Na+, K+, and spermine, but it was not affected greatly by Mg2+, Co2+, and EDTA. Ca2+, Fe2+, Zn2+, and Cu2+ inhibited the reaction. Among the synthetic substrates examined, the enzyme preferentially hydrolyzed pyrimidine nucleotides, with a higher affinity for polycytidylate than for polyuridylate. The enzyme was thermolabile, but it stabilized with bovine plasma albumin. The molecular weight was approximately 15,000, estimated gel filtration on Sephadex G-75, and its isoelectric pH was above 11.0. From normal human leukocytes, acid RNase was purified about 400-fold by the same procedure described previously except that rechromatography on Sephadex G-75 was omitted. The properties of leukocytic RNase were found to be similar to those of serum acid RNase, but the latter enzyme differed in substrate specificity substantially from leukocytic RNase, preferring polyuridylate to polycytidylate. This evidence shows that serum RNase is not of leukocytic origin under normal physiological conditions.
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PMID:Purification and properties of acid ribonucleases in human serum and leukocytes. 2 64

By sequential acid treatment, gel filtration and KM-cellulose sorption a 18--20-fold purified preparation of ribonuclease with a yield of 50--60% was obtained from the culture liquid filtrate of Actinomyces rimosus 994. The preparation had a high specific activity of 450,000--600,000 units/mg protein, contained 85--98% protein, insignificant amounts of carbohydrates and hydroxytetracycline, and no quantities of DNase, phosphomonoesterases, phosphodiesterase or proteases. In RNA degradation (preparation of the total yeast RNA of the Sigma Co.) optimal results were obtained at 50 degrees C and pH 7.0--7.2 in phosphate buffer and 7.6--8.0 IN Tris-HCl buffer. The preparation was stable at high temperatures (80--100 degrees) in the wide pH range and during storage in the lyophilized form and in buffer solutions. RNase effect was inhibited by zinc, copper, iron and cobalt cations and activated by beta-mercaptoethanol, citrate and EDTA. Protamine sulphate and urea in low concentrations (0.01% and 1--4 M, respectively) accelerated and in high concentrations (1% and 8 M, respectively) terminated the enzyme reaction. With respect to many properties RNase from Act. rimosus 994 was similar to extracellular RNases, produced by other actinomycetes and fungi.
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PMID:[Preparation of extracellular ribonuclease form Actinomyces rimosus 994]. 3 16

HeLa (substrain Ho) grown in serum free medium showed an increase in the specific activity of alkaline phosphatase when fetal calf serum (10%) was added to the medium (9.7 nmoles/sec/mg protein to 86.8). Under the same conditions, eight intracellular enzymes showed no increase in activity. Similar results were obtained using a different serum or medium, and with a second strain of HeLa (substrain ATC). For a given set of growth conditions, the effect of serum was dependent on its concentration and required one or more culture generations to develop. The type of isozyme expressed did not change. Neither zinc nor a total serum lipid extract would substitute for serum. The enzyme expressed by HeLaHo was not induced by prednisolone, while that in HeLaATC was. However, for cells grown in excess prednisolone without serum, the specific activity was 25% of that found for cells grown with prednisolone and serum. Cortexolone, an antagonist of prednisolone, was without effect for HeLaHo grown in A3 medium with or without serum. The serum factor had the following characteristics. It was not lost on dialysis, treatment with DNase and RNase, or removal of lipoproteins. It was reduced after heating by 65% and after treatment with Pronase by 82%. The data are interpreted to indicate the presence of a factor (s) in serum, probably a protein, which is involved in stimulating alkaline phosphatase specific activity.
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PMID:Evidence for a high molecular weight factor(s) in serum which increases alkaline phosphatase specific activity in HeLa. 3 90

The reduction of nucleic acid by an endogenous polynucleotide phosphorylase and ribonuclease in cells of Brevibacterium JM98A (ATCC 29895) was studied. A simple process was developed for the activation of the endogenous RNA-degrading enzyme(s). RNA degradation was activated by the presence of Pi with 14.2 mumol of ribonucleoside 5'-monophosphate per g of cell mass accumulating extracellularly. The optimum pH for degradation of RNA was 10.5 and the optimum temperature was 55 to 60 degrees C. Enzymatic activity was inhibited by the presence of Ca2+, Zn2+, or Mg2+. Although some of the RNA-degrading enzymatic activity was associated with the ribosomal fraction, most was soluble. Both polynucleotide phosphorylase and ribonuclease activities were identified.
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PMID:Reduction of endogenous nucleic acid in a single-cell protein. 3 4

1. Autodegradation of yeast ribosomes is due to a 'latent' ribonuclease which is associated with the 40 S ribosomal subunit. 2. The ribonuclease was extracted in the presence of EDTA from ribosomes and purified 118-rold by protamine sulphate precipitation, (NH4)2SO4 fractionation and chromatography on DEAE-cellulose. 3. The optimum pH for this enzyme is 5 to 6.5 while the optimum temperature is 45 to 50 degrees C. Incubation for 10 min at 60 degrees C caused a reduction in enzyme activity of 70%. 4. The ribonuclease has an endonucleolytic activity against rRNA, tRNA, poly(A), poly(U) and poly(C) but does not degrade poly(G) or DNA. It hydrolyzes the homopolymers to nucleoside 3'-phosphates. 5. Zn2+, Mn2+, heparin, glutathione and p-chloromercuribenzoate inhibit the ribonuclease, while Na+, K+, EDTA and sermidine have only little or no effect. 6. It binds tightly to yeast ribosomes but only loosely to ribonuclease-free wheat germ ribosomes. 7. Polyribosomes possess less autodegradation activity than monoribosomes, isolated from the same homogenate.
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PMID:A ribonuclease from yeast associated with the 40 S ribosomal subunit. 4 79

Nucleoli isolated from Novikoff hepatoma cells of the rat were previously shown to carry out synthesis of predominantly ribosomal precursor RNA and methylation of this RNA in vitro. In order to develop in vitro systems for further detailed study of these processes and their interrelationships, isolated nucleoli were incubated in a complete RNA-synthesizing medium using (5-3H)cytidine 5'-triphosphate or S-adenoxyl(methyl-3H)methionine to measure the activities of RNA synthesis and methylation, respectively, under the same reaction conditions. Methylation of the ribose of the nascent ribosomal precursor RNA predominated. It occurred in close coordination with the transcriptional step by RNA polymerase as shown by the kinetic data, the analysis of labeled RNA in sucrose gradients, the inhibition by increased ionic strength or actinomycin D, and the release of labeled nucleotides by a 3'-exonuclease, venom phosphodiesterase. Methylation of the RNA bases occurred more slowly, continued longer after transcription ceased, and appeared to follow later in the processing of the RNA. Certain divalent cations (Mg2+, Mn2+, and Ca2+ at higher concentrations, and Zn2+ and Cu2+) inhibited both RNA synthesis and methylation to similar extents. RNase inhibitors (bentonite and dextran sulfate) at low concentration inhibited methylation while stimulating RNA synthesis, and pyrophosphate greatly decreased RNA synthesis with relatively little effect on methylation. These results indicated that RNA polymerase and ribosomal RNA methylases can function independently despite their close relationship. An exogenous substrate for the nucleolar rRNA methylases was found: nuclear RNA prepared from Novikoff hepatoma cells, cultured in the absence of methionine, served as a good substrate for methylation of both ribose and bases. Other exogenous RNAs, including cytoplasmic ribosomal RNA from these methionine-starved cells, nucleolar RNA from normal cells, and wheat germ ribosomal RNA were almost devoid of methyl-acceptor activity. A description of these parameters helps establish isolated nucleoli as a suitable system for further study of interaction of RNA polymerase, methylases, and nucleases in control of synthesis of ribosomal RNA.
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PMID:Interrelationships between synthesis and methylation of ribosomal RNA in isolated Novikoff Tumor nucleoli. 16 25

1. A ribonuclease (RNAase CL) (EC 3.1.4.23, ribonucleate 3'-oligonucleotide hydrolase) was extracted by EDTA/acetate buffer, pH 5.6 from acetonedried cells of Candida lipolytica and purified 1350-fold by acetone and (NH4)2SO4 fractionation, DEAE-cellulose and DEAE-Sephadex chromatography. 2. RNAase CL is an acidic protein having an isoelectric point of 4.2, and an approximate molecular weight of 32 000. 3. Optimal pH and temperature for the enzyme were 6.0 and 60 degrees C, respectively. It is stable at neutral pH up to 50 degrees C. At 64 degrees C for 30 min, 95, 49 and 64% inactivation of the enzyme occurred at pH values 4.2, 6.6 and 10.0, respectively. 4. RNAase CL inhibited by Zn2+ and Cu2+, sulfhydryl reactants and by high concentration of salts, but not by chelating agents. 5. RNAase CL degraded ribosomal RNA, transfer RNA, polyadenylic acid, polycytidylic acid and polyuridylic acid into acid-soluble nucleotides. Among the synthetic homopolymers, polycytidylic acid was most rapidly degraded. Polyguanylic acid and duplexes of synthetic homopolymers were less sensitive. DNA was not attacked. Specificity studies showed that RNAase CL preferentially cleaves pC-purine bonds. 6. Digestion of poly (C) by RNAase CL resulted in the liberation of cyclic 2',3'-CPM from the start of the reaction with no observable formation of intermediate oligonucleotides. This suggests that the enzyme depolymerizes by an exonucleolytic mechanism.
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PMID:Puridication and properties of an intracellular ribonuclease from Candida lipolytica. 23 21

Zn2+, Cd2+ and Hg2+ inhibit ribonuclease but Mn2+ does not except at very high concentrations. By high resolution NMR one can detect in the pH range 5-8 the C-2 protons of histidines 105, 12, and 119. The inhibiting ions produce large shifts of the resonance of His-12 but not of His-105. On the other hand Mn2+ broadens the C-2 proton of His-105 much more than it does those of His-12 and 119. The selective shifts suggest that the mechanism of inhibition is binding at or near the active site of which His-12 and 119 are a part. The selective broadening is a consequence of binding of the Mn2+ to a site very far from the active site but closer to His-105.
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PMID:Interaction of transition metal ions with ribonuclease A. II. The selective effects of Mn2+, Zn2+, Cd2+ and Hg2+ on the histidine magnetic resonance. 23 49


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