Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.27.3 (RNase T1)
 1,228 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The pseudouridylation of ribosomal RNA of Saccharomyces carlsbergensis was investigated with respect to its timing during the maturation of rRNA and its sequence specificity. Analysis of 37-S RNA, the common precursor to 17-S, 5.8-S and 26-S rRNA and most probably the primary ribosomal transcript, shows that this RNA molecule contains already most if not all of the 36-37 pseudouridine residues found in the mature rRNAs. Thus pseudouridylation is, like 2'-0-ribosemethylation, an early event in the maturation of rRNA, taking place immediately after, or even during, transcription. The data presented show that the non-conserved sequences of 37-S precursor rRNA contain very few pseudouridine residues if any. The pseudouridine residues within the rRNA sequences are apparently clustered to a certain degree as can inferred from the occurrence of a single oligonucleotide containing 3 pseudouridines, which was obtained by digestion of 26-S rRNA with ribonuclease T1.
Nucleic Acids Res 1979 Sep 11
PMID:Pseudouridylation of yeast ribosomal precursor RNA. 11 83

Infectious retroviruses have been isolated from gibbon apes and a woolly monkey. Previous studies have shown that these isolates share some antigenic determinants and that they exhibit partial nucleic acid homology. To further define the relationships in this group of viruses, we compared the RNAs of the viruses of the woolly monkey-gibbon ape class by two-dimensional polyacrylamide gel electrophoresis of the large RNase T1-resistant oligonucleotides. The degree of sequence identity between the RNAs was determined by the similarity of the fingerprint patterns and in some cases by partial sequence analysis of individual oligonucleotides. This technique permitted us to determine the degree of sequence identity in related RNA species. These studies showed that as much as 80% of the genomes of gibbon ape leukosis virus-Halls' Island and gibbon ape leukosis virus-brain could be identical. The other viruses, simian sarcoma-associated virus, gibbon ape leukosis virus-Thailand, and gibbon ape leukosis virus-San Francisco, showed an extensive but somewhat lower degree of sequence identity (between 40 to 60% of the genomes.
J Virol 1979 Sep
PMID:Structural analysis of the genomes of gibbon ape and woolly monkey leukosis viruses. 22 47

The reaction of Escherichia coli ribosomes with beta-ethoxy-alpha-ketobutyraldehyde (kethoxal) in a buffer containing 50--100 mM Tris.HCl at pH 7.4, 50 mM NH4Cl, and 5 mM Mg(OAc)2 readily released the 5S RNA from the ribosomes. When liberated, the 5S RNA is in a conformation such that position 44 is selectively reactive, in addition to the normally reactive quanines at positions 41 and 13. Positions 41 and 13 have been previously shown to react in the 5S RNA in situ. The resulting new RNase T1 resistant oligonucleotides 5'CCG 44K AAUCAG51(3') and 5'ACCCCAUG 41KCCG 44KAACUCAG51(3') have been isolated and identified. These oligonucleotides have not been found in RNase T1 digests of 5S RNA that is not released from the ribosome. The guanine at position 44 is part of the invariant sequence 5'CCG44AAC3' which includes that portion of the molecule thought to interact with the invariant 5'GT psi C3' of tRNAs in the ribosomal A site. This invariant sequence of the 5S RNA may also form part of the binding site for protein L5.
Proc Natl Acad Sci U S A 1979 Sep
PMID:Accessibility of guanine at position 44 in the invariant sequence 5'CCG44AAC3' of Escherichia coli 5S RNA to reaction with kethoxal. 38 42

A modification of the known method for obtaining radioactive fingerprints from non-radioactive nucleic acids by labelling a digest with 5'-hydroxyl polynucleotide kinase and [gamma-32P]-ATP has been applied to RNase T1 digests from various high molecular weight virus RNAs and to ovalbumin mRNA. Fractionation of the resultant [32P]-labelled T1 RNase digests by two-dimensional polyacrylamide electrophoresis demonstrates that in the case of virus RNAs, the fingerprints thus obtained are very similar to those derived from uniformly labelled RNAs. The value of this technique is that it requires only 1-5 microgram of purified virus RNA and at least three orders of magnitude less radioactivity than is routinely employed in preparing uniformly labelled RNA.
Nucleic Acids Res 1977 Sep
PMID:Oligonucleotide mapping of non-radioactive virus and messenger RNAs. 41 3

The fluorescence of the supposedly buried tryptophan in ribonuclease T1 has been found to be collisionally quenched by acrylamide with a rate constant of 3 X 10(8) M--1 sec--1. Only a slight decrease in the quenching rate is observed upon a 5-fold increase in the viscosity of the solution. For this to be the case, the diffusion of the quencher must be limited by the protein matrix. To explain the process of diffusion through this complex material, the formation of "holes" in the lattice of a protein due to nanosecond fluctuations must be invoked. Thus, the dynamic character of a protein molecule is revealed. The quenching rate constant has an activation energy of 9 kcal/mol which can be used to characterize the nature of the cohesive forces in the microenvironment about the indole ring. The mechanical properties of a portion of a protein matrix can, therefore, be described as one would for a fluid.
Proc Natl Acad Sci U S A 1975 Sep
PMID:Dynamics of a protein matrix revealed by fluorescence quenching. 81 Aug

The genomic RNA of the avian influenza A virus, fowl plague, was fractionated into eight species by electrophoresis in polyacrylamide-agarose gels containing 6 M urea. The separated 32P-labeled RNA species were characterized by digestion with RNase T1 and fractionation of the resulting oligonucleotides by two-dimensional gel electrophoresis; this demonstrated that each species has a distinct nucleotide sequence. A tentative correlation of each genome RNA species with the virus protein that it encodes was made.
Proc Natl Acad Sci U S A 1976 Sep
PMID:Influenza virus genome consists of eight distinct RNA species. 106

The frequency of oligonucleotides obtained from simian sarcoma virus RNA by digestion with ribonuclease T1 was compared with the frequency expected of an RNA molecule in which nucleotides are arranged in random distribution. Oligonucleotides containing C-residue attached to 3'-Gp were found significantly less in simian sarcoma virus 70S RNA than expected by random distribution.
J Virol 1975 Sep
PMID:Low frequency of (5'-3') -C-G- connection in 70S RNA from simian sarcoma virus. 117 95

The single-strand specific nuclease S1 from Aspergillus oryzae (EC 3.1.4.21) was purified 600-fold in 16% yield from dried mycelia. Determination of the isoelectric point of S1 nuclease as 4.3-4.4 allowed adjustment of chromatographic conditions such that the enzyme was isolated free of contaminating ribonucleases T1 and T2. S1 nuclease so purified was used for removal of single-stranded portions from the RNA of the Escherichia coli phage MS2, which has a helical content of about 65% in vitro. At 23 degrees, increasing amounts of enzyme converted the RNA to mononucleotides in about equimolar base ratios. No small intermediates of chain length 2-8 were found. At 0 degrees, MS2 RNA hydrolysis was slower and reached, in exhaustive digests, a plateau where 70% of the substrate RNA remained insoluble in 66% EtOH. With [32P]MS2 RNA, strip chart counting of 6% acrylamide-6 M urea electrophoresis patterns of such digests gave recoveries of 80-91% in the form of defined oligomer bands. On 2.5% acrylamide-0.5% agarose gels, the molecular weights of the major oligomers were found to range from 25,000 to 41,000. Similar to purified tRNAArg used as a control, these oligomers were not resistant to pancreatic RNase-RNase T1 hydrolysis at 37 degrees, and were not bound on hydroxylapatite at 50 degrees in 0.14 M sodium phosphate (pH 6.8). Melting of the oligomers gave complex profiles without a clear Tm and showed an increase in A260 of 35% at 93 degrees over that at 28 degrees. Upon formaldehyde denaturation of MS2 RNA prior to S1 nuclease hydrolysis, no resistant oligomers were found.
Biochemistry 1975 Sep 23
PMID:S1 nuclease hydrolysis of single-stranded nucleic acids with partial double-stranded configuration. 118 98

Kinetic intermediates in protein folding are short-lived and therefore difficult to detect and to characterize. In the folding of polypeptide chains with incorrect isomers of Xaa-Pro peptide bonds the final rate-limiting transition to the native state is slow, since it is coupled to prolyl isomerization. Incorrect prolyl isomers thus act as effective traps for folding intermediates and allow their properties to be studied more easily. We employed this strategy to investigate the mechanism of slow folding of ribonuclease T1. In our experiments we use a mutant form of this protein with a single cis peptide bond at proline 39. During refolding, protein chains with an incorrect trans proline 39 can rapidly form extensive secondary structure. The CD signal in the amide region is regained within the dead-time of stopped-flow mixing (15 ms), indicating a fast formation of the single alpha-helix of ribonuclease T1. This step is correlated with partial formation of a hydrophobic core, because the fluorescence emission maximum of tryptophan 59 is shifted from 349 nm to 325 nm within less than a second. After about 20 s of refolding an intermediate is present that shows about 40% enzymatic activity compared to the completely refolded protein. In addition, the solvent accessibility of tryptophan 59 is drastically reduced in this intermediate and comparable to that of the native state as determined by acrylamide quenching of the tryptophan fluorescence. Activity and quenching measurements have long dead-times and therefore we do not know whether enzymatic activity and solvent accessibility also change in the time range of milliseconds. At this stage of folding at least part of the beta-sheet structure is already present, since it hosts the active site of the enzyme. The trans to cis isomerization of the tyrosine 38-proline 39 peptide bond in the intermediate and consequently the formation of native protein is very slow (tau = 6,500 s at pH 5.0 and 10 degrees C). It is accompanied by an additional increase in tryptophan fluorescence, by the development of the fine structure of the tryptophan emission spectrum, and by the regain of the full enzymatic activity. This indicates that the packing of the hydrophobic core, which involves both tryptophan 59 and proline 39, is optimized in this step. Apparently, refolding polypeptide chains with an incorrect prolyl isomer can very rapidly form partially folded intermediates with native-like properties.
Protein Sci 1992 Sep
PMID:Structure of a rapidly formed intermediate in ribonuclease T1 folding. 130 94

From calculations of a model reaction scheme for base-catalyzed RNA hydrolysis, a pentacoodinate dianionic intermediate 2a (Storer, et al., J. Am. Chem. Soc., 1991, 113, 5216-5219) as well as two transition states, TS1 and TS2, to the intermediate have been located by ab initio calculations at the 3-21G* level. Although the intermediate, which has the well depth on the order of kBT, is unlikely to be kinetically significant, the overall rate-limiting transition state structure TS2 obtained at 3-21G* level is very close to the corresponding structure at the STO-3G level; it has an extended P-O(5') bond breaking character. These gas-phase calculation results are used to qualitatively interpret mutagenesis results of Barnase and RNase T1 where water molecules are absent from the active site.
Biochem Biophys Res Commun 1992 Sep 30
PMID:RNA hydrolysis via an oxyphosphorane intermediate. 138 73


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