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)

Because of evidence of an immunologic role for ribonuclease II (E.C. 3.1.27.5) in mammals, its presence in milk was further characterized to provide a basis for study of possible contributions of its activity to the health of infants. Isoenzymes of ribonuclease II were quantitatively resolved from milk samples as small as 1 ml or less by chromatography on phosphocellulose. Three isoenzymes detected in bovine milk were the previously reported ribonucleases A and B and a form termed ribonuclease II-1. These isoenzymes were in the ratio of 70:30:1. Form II-1 was unique in its inability to hydrolyze polycytidylate. Bovine colostrum contained 10 to 15 times more ribonuclease II-1 than does milk and three times more total ribonuclease II per unit volume. Human milk contains about 1% the concentration of ribonuclease II found in cows' milk. Ribonuclease II activity in milk was quite stable in the acidic conditions of whey production and during low heat treatments. However, most of its enzymatic activity was lost with high heat treatments. No commercially manufactured milk-based or soybean-based infant formula assayed contained nearly as much ribonuclease activity as either human or bovine milk.
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PMID:Ribonuclease activity and isoenzymes in raw and processed cows' milk and infant formulas. 366 41

1. Ribonuclease II of Escherichia coli degrades pulse-labelled RNA associated with ribosomes and polyuridylic acid on ribosomes and in solution to mononucleotides. 2. Ribosomal and pulse-labelled RNA in solution and ribosomal RNA in chloramphenicol particles (protein-deficient ribosomes) are degraded to oligonucleotides. 3. Ribosomal RNA in mature ribosomes is not attacked by the enzyme. 4. From the mode of action of ribonuclease II, which is specific for single-stranded polyribonucleotides and does not attack helical forms, it is inferred that pulse-labelled RNA associated with ribosomes of E. coli exists as a single-stranded structure and that ribosomal RNA in chloramphenicol particles has a pronounced helical character. 5. The different behaviour of ribonuclease II towards newly synthesized RNA, ribosomal RNA and chloramphenicol-particle RNA in E. coli ribosomes is discussed.
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PMID:The conformation of ribonucleic acids in Escherichia coli ribosomes. Inferences from the mode of action of ribonuclease II. 486 Jun 40

Ribonuclease II (RNase II), encoded by the rnb gene, is one of the two major Escherichia coli exonucleases involved in mRNA degradation. Some of the ribonucleases implicated in this process have recently been shown to be inter-regulated. In this paper we studied the effects of the endonucleases RNase E and RNase III in rnb expression. We have shown that RNase E cleaves the rnb message internally: when this ribonuclease is inactivated rnb mRNA accumulates with a concomitant increase in RNase II activity. RNase III also affects RNase II expression but in an indirect way. We discuss these implications for the regulation of mRNA degradation.
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PMID:The role of endonucleases in the expression of ribonuclease II in Escherichia coli. 764 46

Ribonuclease II is a processive 3' exoribonuclease in Escherichia coli. It degraded substrates with 3'-OH or 2',3'-cyclicP ends slightly faster than those with 3'-P or 2'-P groups with a turnover number of approximately 70 nt/s at 37 degrees C. RNase II does not degrade DNA but the specificity for ribose was not for the cleavage bond but rather for ribo-bonds three to four nucleotides (nt) upstream, which could explain why the limit digest is a dimer. Oligonucleotides (oligos) of deoxy(C) were reversible competitive inhibitors of the enzyme and indicated a strong upstream binding site (approximately 15 to 27 nt from the 3' end). These oligos could protect RNase II from inactivation by heat or from diethylpyrocarbonate, an agent that preferentially reacts with His residues. Compared to oligo(dC), oligos of (dA) were at least 500 times less effective inhibitors of RNase II. Using mixed oligo(dAdC) inhibitors, an obligatory 3' to 5' direction of binding into the catalytic site was shown. From the reaction kinetics of RNase II under different conditions it was concluded that the enzyme recognition differs for poly(A), poly(C) and poly(U). Poly(C) was degraded more slowly than poly(A) or poly(U) with a 3.5 times slower Vmax, while rate differences between small oligos were extreme; oligo(A)7 was degraded > 100 times faster than oligo(C)7. Ethanol, which weakens hydrophobic interactions, increased the reaction velocity of poly(C) to that of poly(A) and poly(U). It had no effect on the reaction velocities of poly(A) or poly(U), but decreased the binding of poly(A) markedly. Oligo(A) was bound more strongly to a hydrophobic column than was oligo(C). Salt, which affects charge interactions, decreased the binding affinity and/or association rate of poly(C) to RNase II, had a lesser effect on poly(U), but the reactions of poly(A) were unaffected even in much higher concentrations of salt. A clue to the slower reaction velocity of poly(C) was shown when the reaction intermediates were viewed by PAGE. At lower temperatures of reaction (< 25 degrees C), there were more intense bands separated by discrete distances of approximately 12 nt during the degradation of poly(C) by RNase II. Chase experiments showed that these stops were accounted for by dissociation of poly(C) from the enzyme. They were not seen when poly(C) was degraded at 37 degrees C or degraded in the presence of 20% ethanol at any temperatures, nor were they seen when poly(A) or poly(U) was degraded even at low temperatures.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:The processive reaction mechanism of ribonuclease II. 796 9

Ribonuclease II (RNase II) is a major exonuclease in Escherichia coli that hydrolyzes single-stranded polyribonucleotides processively in the 3' to 5' direction. To understand the role of RNase II in the decay of messenger RNA, a strain overexpressing the rnb gene was constructed. Induction resulted in a 300-fold increase in RNase II activity in crude extracts prepared from the overexpressing strain compared to that of a non-overexpressing strain. The recombinant polypeptide (Rnb) was purified to apparent homogeneity in a rapid, simple procedure using conventional chromatographic techniques and/or fast protein liquid chromatography to a final specific activity of 4,100 units/mg. Additionally, a truncated Rnb polypeptide was purified, solubilized, and successfully renatured from inclusion bodies. The recombinant Rnb polypeptide was active against both [3H]poly(A) as well as a novel (synthetic partial duplex) RNA substrate. The data show that the Rnb polypeptide can disengage from its substrate upon stalling at a region of secondary structure and reassociate with a new free 3'-end. The stalled substrate formed by the dissociation event cannot compete for the Rnb polypeptide, demonstrating that duplexed RNAs lacking 10 protruding unpaired nucleotides are not substrates for RNase II. In addition, RNA that has been previously trimmed back to a region of secondary structure with purified Rnb polypeptide is not a substrate for polynucleotide phosphorylase-like activity in crude extracts. The implications for mRNA degradation and the proposed role for RNase II as a repressor of degradation are discussed.
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PMID:Overexpression, purification, and properties of Escherichia coli ribonuclease II. 855 29

Ribonuclease II is a processive 3'- to 5'-exoribonuclease in Escherichia coli with two binding sites: a catalytic site associated with the first few 3'-nucleotides and an anchor site binding nucleotides approximately 15 to 25 from the 3'-end. When RNase II degrades single-stranded helical poly(C), the enzyme-substrate complex dissociates at discrete intervals of 12 nucleotides. RNase II stalled at the last rC of single-stranded 3'-(rC)(n)(dC)(m) oligonucleotides. The more residues released, the faster the stalled complex dissociated and the less it inhibited RNase II activity, i.e. the enzyme-substrate association weakened progressively. Using phosphodiesterase I (PDE I) as a probe, a method was developed to identify cytidine residues in (32)P-oligonucleotides interacting with a protein. PAGE bands corresponding to nucleotides 1-6 from the 3'-end were consistent with interaction at the catalytic site, and following a gap, bands approximately 15 to 25 from the 3'-end, with anchor site association. Both 3' and 5' binding were necessary to maintain the complex. Of most significance, the original anchor site nucleotides remained fixed at the anchor site while the 3'-end was pulled, or threaded, through the catalytic site, i.e. the substrate did not 'slide' through the enzyme. DNA oligonucleotides with double-stranded stem-loops were good competitive inhibitors of RNase II. A 3'-single-stranded arm was essential, while optimal binding required both 5'- and 3'-arms. PDE I probing indicated that the nucleotides at the anchor site were specified by the spatial distance from the catalytic site, and on only one of the duplex strands. When degradation of a structured RNA paused or stopped, the RNase II-product commenced cycles of dissociation-reassociation. Duplex strand binding by RNase II made complex DNA or RNA structures accessible to degradation by other nucleases and further verified the PDE I footprinting method.
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PMID:The reaction mechanism of ribonuclease II and its interaction with nucleic acid secondary structures. 1044 70

The human ISG20/HEM45 gene was identified independently on the basis of its increased level of expression in response to either interferon or estrogen hormone. Notably, the encoded protein is homologous with members of the 3' to 5' exonuclease superfamily that includes RNases T and D, and the proofreading domain of Escherichia coli DNA polymerase I. We provide here direct biochemical evidence that Isg20 acts as a 3' to 5' exonuclease in vitro. This protein displays a pH optimum of approximately 7.0, prefers Mn2+ as a metal cofactor, and degrades RNA at a rate that is approximately 35-fold higher than its rate for single-stranded DNA. Along with RNase L, Isg20 is the second known RNase regulated by interferon. Previous data showed that Isg20 is located in promyelocytic leukemia (PML) nuclear bodies, known sites of hormone-dependent RNA polymerase II transcription and oncogenic DNA viral transcription and replication. The combined data suggest a potential role for Isg20 in degrading viral RNAs as part of the interferon-regulated antiviral response and/or cellular mRNAs as a regulatory component of interferon and estrogen signaling.
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PMID:The human interferon- and estrogen-regulated ISG20/HEM45 gene product degrades single-stranded RNA and DNA in vitro. 1140 64

Exoribonuclease II (RNase II), encoded by the rnb gene, is a ubiquitous enzyme that is responsible for 90% of the hydrolytic activity in Escherichia coli crude extracts. The E. coli strain SK4803, carrying the mutant allele rnb296, has been widely used in the study of the role of RNase II. We determined the DNA sequence of rnb296 and cloned this mutant gene in an expression vector. Only a point mutation in the coding sequence of the gene was detected, which results in the single substitution of aspartate 209 for asparagine. The mutant and the wild-type RNase II enzymes were purified, and their 3' to 5' exoribonucleolytic activity, as well as their RNA binding capability, were characterized. We also studied the metal dependency of the exoribonuclease activity of RNase II. The results obtained demonstrated that aspartate 209 is absolutely essential for RNA hydrolysis, but is not required for substrate binding. This is the first evidence of an acidic residue that is essential for the activity of RNase II-like enzymes. The possible involvement of this residue in metal binding at the active site of the enzyme is discussed. These results are particularly relevant at this time given that no structural or mutational analysis has been performed for any protein of the RNR family of exoribonucleases.
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PMID:A single mutation in Escherichia coli ribonuclease II inactivates the enzyme without affecting RNA binding. 1565 75

Three enzymes with ribonuclease activity, one of which also had deoxyribonuclease activity, have been isolated and partially purified from corn seeds and seedlings. The purification of Ribonuclease I from mature seed was previously reported. This enzyme has a pH optimum near 5.0, is loosely adsorbed to carboxymethyl-cellulose, and has a molecular weight of 23,000, determined by gel filtration.Ribonuclease II was isolated from the microsomes of corn roots, and was partially purified by gel filtration. It has a pH optimum plateau from 5.4 to 7.0, and molecular weight of 17,000.Nuclease I hydrolyzes both RNA and DNA. It was isolated from the large particles of a corn root homogenate and was partially purified on a carboxymethyl-cellulose column. It has a pH optimum at 6.2 and a molecular weight of 31,000.The relative activities of the 3 enzymes for deoxyribonuclease and at pH 5 and pH 6.2 for ribonuclease may be used to characterize them during purification operations. Assays on homogenates of corn roots, and especially of the root tips, suggested that a fourth enzyme, which possesses deoxyribonuclease activity, is also present.
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PMID:Plant nucleases. I. Separation and purification of two ribonucleases and one nuclease from corn. 1665 16

Besides linear RNAs, pre-mRNA splicing generates three forms of RNAs: lariat introns, Y-structure introns from trans-splicing, and circular exons through exon skipping. To study the persistence of excised introns in total cellular RNA, we used three Escherichia coli 3' to 5' exoribonucleases. Ribonuclease R (RNase R) thoroughly degrades the abundant linear RNAs and the Y-structure RNA, while preserving the loop portion of a lariat RNA. Ribonuclease II (RNase II) and polynucleotide phosphorylase (PNPase) also preserve the lariat loop, but are less efficient in degrading linear RNAs. RNase R digestion of the total RNA from human skeletal muscle generates an RNA pool consisting of lariat and circular RNAs. RT-PCR across the branch sites confirmed lariat RNAs and circular RNAs in the pool generated by constitutive and alternative splicing of the dystrophin pre-mRNA. Our results indicate that RNase R treatment can be used to construct an intronic cDNA library, in which majority of the intron lariats are represented. The highly specific activity of RNase R implies its ability to screen for rare intragenic trans-splicing in any target gene with a large background of cis-splicing. Further analysis of the intronic RNA pool from a specific tissue or cell will provide insights into the global profile of alternative splicing.
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PMID:Characterization of RNase R-digested cellular RNA source that consists of lariat and circular RNAs from pre-mRNA splicing. 1668 42


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