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Query: EC:3.1.27.5 (RNase)
 17,967 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

RNAs synthesized in Escherichia coli infected with virulent phages T4, T5, T7 and BF23 were labelled with 32PO4 3- after phage infection. [32P]RNAs of low molecular weight were separated by two-dimensional polyacrylamide gel electrophoresis, in which electrophoresis was carried out in two dimensions at different concentrations of acrylamide. The fractionated RNAs were characterized by RNA-fingerprint patterns made after T1 ribonuclease digestion. The two-dimensional gel of 10% yields 20% acrylamide was suitable for RNA of less than 200 nucleotides, while that of 5% yields 10% was preferred for RNAs of about 150--400 nucleotides. With T4 phage, 16 RNA species were separable on a single slab gel. Among those, 11 were identified as the known RNA species, including eight T4 tRNAs, one tRNA precursor and two non-tRNA molecules. In the case of T5 and BF23, more than 20 RNA species were separated on a slab gel; 15 or more RNAs were found in the 4-S RNA region, and several in 5-S and 6-S region. The RNA-fingerprint patterns of many BF23 RNAs were very similar to those of corresponding RNAs of T5. Pseudouridine and ribosylthymidine, minor nucleosides generally present in tRNA, were found in several BF23 4-S RNAs tested. Possibility of those BF23 4-S RNAs as tRNAs is discussed. With phage T7, three RNAs were detected, two of which were much smaller than tRNAs.
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PMID:Two-dimensional polyacrylamide-gel electrophoresis for purification of small RNAs specified by virulent coliphages T4, T5, T7 and BF23. 109 84

Behaviour of modified nucleosides, tRNA components, and their analogues has been studied in the internucleotide bond formation catalysed by ribonucleases of various substrate specificity, polynucleotide phosphorylases, and T4 RNA ligase and the results are summarised in this paper. Pseudouridine, dihydrouridine, ribothymidine, 5-methylcytidine, inosine, and 6-methyladenosine can participate in the reaction of internucleotide bond formation the presence of most ribonucleases used, viz. Pb2, Pcl2, Pb1, Pch1, C2, T1, pancreatic RNase. 3-Methylcytidine and 4-acetylcytidine form internucleotide bond (as phosphate acceptors) usually by means of guanyl-specific ribonucleases, whereas 1-methylandenosine is incorporated with ribonuclease Pel2. 7-Methylguanosine and 1-methylguynosine 2',3'-cyclophosphates can be used as phosphate donors in the presence of ribonuclease Pb2; in the similar enzymatic reaction 6-isopentenyladenosine is an uneffective acceptor.
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PMID:[Enzymatic incorporation into oligonucleotides of modified nucleosides]. 367 47

Pseudouridine, an isomer of uridine, is probably the most common of many posttranscriptional RNA modifications found in nature. Although mass spectrometry has become widely used in the characterization of modified nucleic acids, its application to the recognition and sequence placement of pseudouridine has not been straightforward, particularly in the case of complex mixtures such as those resulting from selective enzymatic hydrolysis of RNA into oligonucleotides. We report results of a study of the characteristic dissociation reactions of pseudouridine-containing oligonucleotides following ionization by electrospray and use of those pathways in an LC/MS-based method applicable to direct analysis of RNase digests of RNA. As a consequence of the C-C (rather than C-N) glycosidic bond of pseudouridine, the otherwise common dissociation paths involving base loss do not occur, resulting in characteristic formation of a set of low-mass negative ions containing the intact glycosidic bond (m/z 225, 207, 189, 165, 164, 139), which permit recognition of pseudouridine-containing oligonucleotides. Those components can subsequently be subjected to sequence analysis by MS/MS, in which enhancement of selective sequence-determining ions (a-, w-, y-types), and absence of a - base ions, are observed at the site of pseudouridylation. Also, selected reaction pathways can be monitored in the LC/MS/MS analysis that are indicative of pseudouridine at the 5' terminus (m/z 225 --> 165), internal positions (m/z 207 --> 164), and in the RNase T1-derived product Psi pGp (m/z 668 --> 207) arising from the RNA sequence ...G Psi G... These procedures can be effectively integrated into an existing suite of LC/ESI-MS-based methods designed for the analysis of posttranscriptionally modified sites in RNA.
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PMID:Detection of the common RNA nucleoside pseudouridine in mixtures of oligonucleotides by mass spectrometry. 1605 77

Pseudouridine, the so-called fifth nucleoside due to its ubiquitous presence in ribonucleic acids (RNAs), remains among the most challenging modified nucleosides to characterize. As an isomer of the major nucleoside uridine, pseudouridine cannot be detected by standard reverse-transcriptase-based DNA sequencing or RNase mapping approaches. Thus, over the past 15 years, investigators have focused on the unique structural properties of pseudouridine to develop selective derivatization or fragmentation strategies for its determination. While the N-cyclohexyl-N'-beta-(4-methylmorpholinium)ethylcarbodiimide p-tosylate (CMCT)-reverse transcriptase assay remains both a popular and powerful approach to screen for pseudouridine in larger RNAs, mass-spectrometry-based approaches are poised to play an increasingly important role in either confirming the findings of the CMCT-reverse transcriptase assay or in characterizing pseudouridine sequence placement and abundance in smaller RNAs. This review includes a brief discussion of pseudouridine including a summary of its biosynthesis and known importance within various RNAs. The review then focuses on chemical derivatization approaches that can be used to selectively modify pseudouridine to improve its detection, and the development of mass-spectrometry-based assays for the identification and sequencing of pseudouridine in various RNAs.
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PMID:Mass spectrometry of the fifth nucleoside: a review of the identification of pseudouridine in nucleic acids. 1862 Sep 15