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

2' and 3'-O-(N-acetyl-L-phenylalanyl)adenosine (Ac-Phe-Ado) were chemically synthesized. These two isomers were clearly separated from each other by high-performance liquid chromatography (HPLC). From the two isomers of [3H]Phe-tRNA in equilibrium, Ac-[3H]Phe-Ado was prepared, without any change in the 2'/3'-isomer ratio, by acetylation of the phenylalanyl residue with acetic anhydride followed by digestion with pancreatic RNase A. By HPLC analysis of this preparation of Ac-[3H]Phe-Ado, the abundance ratio of the 2'-isomer and the 3'-isomer of [3H]Phe-tRNA was found to be 0.20:0.80. Further, [3H]Phe-tRNA was bound to Escherichia coli polypeptide chain elongation factor Tu (EF-Tu) with the ligand of GTP or guanosine 5'-[beta, gamma-imido]triphosphate (GMP-P(NH)P). The ternary complex was treated with phenol and acetic anhydride, and then digested with pancreatic RNase A. By HPLC analysis of Ac-[3H]Phe-Ado, the abundance ratio of the 2'-isomer and the 3'-isomer of [3H]Phe-tRNA was determined to be 0.07:0.93 in the complex with EF-Tu.GTP and 0.04:0.96 in the complex with EF-Tu.GMP-P(NH)P. These results clearly indicate that the 3'-isomer, rather than the 2'-isomer, of aminoacyl-tRNA is exclusively involved in the ternary complex.
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PMID:Aminoacyl-tRNA exclusively in the 3'-isomeric form is bound to polypeptide chain elongation factor Tu. 385 54

The ribonuclease resistance assay has been used to probe the effect of trypsin modification of the Escherichia coli elongation factor Tu X GTP on the interaction with E. coli aminoacyl-tRNAs. First, the equilibrium dissociation constant of the trypsin-modified Tu X GTP X Thr-tRNA complex was determined to be 2.3 (0.1) X 10(-5)M at 4 degrees C, pH 7.4. Second, binding of 17 of 20 noninitiator aminoacyl-tRNAs and four sets of purified isoacceptor tRNAs to the modified protein was measured. At 4 degrees C, the complex stabilities vary 500-fold over the range of aminoacyl-tRNAs, with Gln-tRNA forming the strongest ternary complex and Val-tRNA, the weakest. The results are compared to a similar study of ternary complex formation using intact elongation factor Tu X GTP, and the major differences are discussed. An analysis of both data sets, particularly that for the leucine isoacceptor tRNAs, suggests that the trypsin modification of elongation factor Tu X GTP disrupts a region of protein that is involved with the aminoacyl side chain rather than that of the acceptor stem helix region of the aminoacyl-tRNA.
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PMID:Effect of trypsin modification of the Escherichia coli elongation factor Tu on the ternary complex with aminoacyl-tRNA. 389 46

On incubation of 50 S ribosomes, isolated from either tight couple (TC) or loose couple (LC) 70 S ribosomes, with elongation factor G (EG-G) and guanosine 5'-triphosphate, a mixture of TC and LC 50 S ribosomes is formed. There is almost complete conversion of LC 50 S ribosomes to TC 50 S ribosomes on treatment with EF-G, GTP, and fusidic acid. Similarly, TC 50 S ribosomes are converted to LC 50 S ribosomes, although partially, by treatment with EF-G and a GTP analogue like guanyl-5'-yl methylenediphosphate (GMP-P(CH2)P) or guanyl-5'-yl imidodiphosphate (GMP-P(NH)P) and including a polymer of 5'-uridylic acid (poly(U] in the incubation mixture. Furthermore, LC 23 S RNA isolated from LC 50 S ribosomes is converted to TC 23 S RNA on heat treatment, but similar treatment does not affect TC 23 S RNA. The interconversion was followed by several physical and biological characteristics of TC and LC 50 S ribosomes, like association capacities with 30 S ribosomes before and after kethoxal treatment, susceptibility to RNase I and polyphenylalanine-synthesizing capacity in association with 30 S ribosomes, as well as thermal denaturation profiles, circular dichroic spectra, and association capacity of isolated 23 S RNAs. These data strongly support the proposition that TC and LC 50 S ribosomes are the products of translocation during protein synthesis. The conformational change of 23 S RNA induced by EF-G and GTP is most probably responsible for the interconversion, and L7/L12 proteins play an important role in the process. A two-site model based on kethoxal data has also been proposed to explain the tightness and looseness of 70 S couples.
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PMID:Interconversion of tight and loose couple 50 S ribosomes and translocation in protein synthesis. 389 20

Cell ghosts have been prepared from mature chicken erythrocytes using 0.05% saponin. Such preparations are capable of incorporating label from [3H]UTP and provide a system, where the nucleus is permeable to nucleotides and macromolecules, for studying the low-level RNA synthesis characteristic of these cells. RNase A (50 micrograms/ml) eliminated all radioactivity binding to DE-81 filters, indicating that the product was RNA; and DNase (10 micrograms/ml) and actinomycin D (10 micrograms/ml) each inhibited UMP incorporation by 70%, suggesting that the synthesis was DNA-dependent. Polymerization was inhibited 90% by 0.1 microgram/ml alpha-amanitin, and maximum synthesis occurred in the presence of high salt (0.175 M KCl) and Mn2+ (0.5 mM). Polyacrylamide gel electrophoresis indicated that the newly synthesized RNA was heterogeneous in size, having a distribution from 5 to 60 S with a significant fraction migrating as 8-12 S. Approximately 15% of the total RNA was bound by an oligo(dT)-cellulose column, suggesting that some RNA processing was occurring, although attempts to detect the incorporation of label from [alpha-32P]GTP into a 5'-cap structure were unsuccessful. In comparison to RNA synthesis in reticulocyte nuclei, both the rate and extent of transcription in erythrocyte nuclei were much reduced. Moreover, about 25-30% of the reticulocyte nascent RNA was released from the nuclei during a 60-min incubation, while no release was observed for the erythrocyte nuclei. Hybridization of radiolabeled RNA to excess chicken DNA indicated that the majority (80%) of the in vitro transcripts were complementary to unique sequence DNA (C0t1/2 = 4.5 X 10(3)). When RNA synthesized by either erythrocyte or reticulocyte nuclei was hybridized to cDNA complementary to reticulocyte polysomal mRNA, about 8% of the reticulocyte nuclear RNA but less than 1% of the erythrocyte nuclear RNA were resistant to RNase A digestion. Taken together, these data suggest that nuclei prepared by saponin lysis of chicken erythrocytes synthesize messenger-like RNA via endogenous polymerase II activity. A fraction of this RNA is polyadenylated but contains few, if any, globin sequences or other transcripts found on reticulocyte polysomes.
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PMID:Synthesis of messenger-like RNA in avian erythrocyte nuclei. 390 27

The binding of ribosomes to mRNA is analyzed in a fractionated system from wheat germ with [3H]uridine-labeled poly(A)+ RNA prepared from germinating wheat embryos. The reaction requires factors eIF3, eIF4C, and eIF5; Met-tRNA and the Met-tRNA binding system; either GTP or GMP-PNP; ATP; and factors C1 and eIF4A. These requirements are identical to those previously found to be necessary for formation of ribosome X Met-tRNAMeti complexes, with the exception of ATP, and factors C1 and eIF4A. The function of factors C1 and eIF4A is therefore specifically related to the mRNA attachment reaction. The presence of GTP in the mRNA binding reaction results in the formation of 80 S ribosome complexes, while with GMP-PNP only 40 S ribosome complexes are formed. Ribosome binding to native reovirus RNA in the fractionated wheat germ system is similar to the reaction with poly(A)+ RNA, strongly requiring ATP and factors C1 and eIF4A. Binding to inosine-substituted reovirus RNA, however, is only partially dependent upon ATP, and both the ATP-dependent and the ATP-independent binding reactions strongly require factor C1 and are substantially stimulated by factor eIF4A. The ATP-independent reaction is inhibited by pm7GDP, has a strong requirement for Met-tRNAMeti, and the 40 S ribosome complex is stable to RNase. These results indicate that the ATP-independent binding of ribosomes to inosine-substituted reovirus RNA proceeds through the normal initiation process. They further suggest that neither factor C1 nor eIF4A function exclusively to unwind mRNA secondary structure. Since eIF4A is required for the ATP-independent binding to inosine mRNA, and at the same time interacts with ATP in the reaction with ATP-requiring mRNAs, this factor may have two roles in protein chain initiation, one related to the mRNA X ribosome interaction, and one related to the function of ATP.
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PMID:Initiation factors eIF4A and C1 from wheat germ and the formation of mRNA X ribosome complexes. 398 12

A cytosolic factor that stimulates transcription in isolated nuclei was purified approximately 4000-fold to near homogeneity from rat liver. The molecular weight of the factor was determined as 47 000 by SDS-polyacrylamide gel electrophoresis. The factor had no detectable deoxyribonuclease and protease activity but showed ribonuclease inhibitor activity. The factor could stimulate transcription in isolated nuclei by 50% at about 3.0 ng and the maximal stimulation was about 100%. When [gamma-S]ATP and [gamma-S]GTP were included in the reaction, the factor stimulated the synthesis of RNA which was able to bind to a mercury-Sepharose column and about 80% of the bound RNA was sensitive to a low concentration of alpha-amanitin. When heparin was added before initiation to preincubation mixture containing RNA polymerases II and DNA, a small but definite incorporation of [14C]UTP was observed. The factor alone had no stimulatory effect on the heparin-resistant incorporation of [14C]UTP but, in the presence of two rat liver nuclear fractions, phosphocellulose 0.5 and 1 M KCl step fractions, could stimulate the incorporation above the level with the combination of the two nuclear fractions. Antibody raised against the factor inhibited accurate transcription from the adenovirus 2 major late promoter in a nuclear lysate from Ehrlich ascites tumor cells, and the inhibition was neutralized by the factor.
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PMID:Purification of a cytosolic factor from rat liver that stimulates transcription in isolated nuclei and its action on purified RNA polymerase II-DNA system. 407 43

Treatment of insect polyribosomes with 1 M KCl released a messenger ribonucleoprotein with a pronounced 16S peak. Phenol extraction resulted in a defined peak of 10S RNA, which was judged as mRNA by the following criteria: it showed specificity for binding to ribosomes, and the formation of initiation complex was dependent on protein initiation factors, GTP, mRNA, and aminoacyl-tRNA. The complex directed protein synthesis upon the addition of elongation factors. mRNA was treated with phosphatase and phosphorylated at the 5'-end with [(32)P]cyanoethylphosphate. [(32)P]mRNA was digested by T1 ribonuclease to completion and chromatographed on DEAE-cellulose. The only fragment with (32)P was 15 nucleotides long; it was treated with pancreatic ribonuclease and fingerprinted. Fractions of AC, AAC, and AAAC were found. Initiation signal AUG or GUG in these mRNAs does not begin immediately at the 5'-end and may be at a distance greater than 15 nucleotides. Alkaline hydrolysis of mRNAs labeled in vivo with [(14)C]adenosine revealed Ap and pppAp. Alkaline hydrolysis of mRNA labeled with (32)P at the 5'-terminus resulted in pAp. Hence, these results suggest that in a heterogeneous population of mRNAs from insects, all start with A and have sequence homology at the 5'-termini. This sequence may reflect the signal for RNA polymerase on the gene or may promote the binding of mRNA to ribosomes.
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PMID:Sequence homology at the 5'-termini of insect messenger RNAs. 435 Nov 73

During replication of polyoma DNA in isolated nuclei, RNA was found attached to the 5' ends of growing progeny strands. This RNA starts with either ATP or GTP and can be labeled at its 5' end with (32)P from beta-labeled nucleotides. Digestion of progeny strands with pancreatic DNase released (32)P-labeled RNA that, on gel electrophoresis, gave a distinct peak in the position expected for a decanucleotide. We believe that this short RNA is involved in the initiation of the discontinuous synthesis of DNA and propose the name "initiator RNA" for it. The covalent linkage of initiator RNA to 5' ends of growing DNA chains was substantiated by the finding that (32)P was transferred to ribonucleotides by alkaline hydrolysis of purified initiator RNA obtained by DNase digestion of polyoma progeny strands synthesized from [alpha-(32)P]dTTP. While initiator RNA was quite homogeneous in size, it had no unique base sequence since digestion with pancreatic RNase of initiator RNA labeled at its 5' end with (32)P released a variety of different [(32)P]oligonucleotides. The switch from RNA to DNA synthesis during strand elongation may thus depend on the size of initiator RNA rather than on a specific base sequence.
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PMID:Initiator RNA in discontinuous polyoma DNA synthesis. 437 33

Rat liver mitochondria isolated in sucrose-N-tris(hydroxymethyl)methyl-2-aminoethane-sulphonic acid (TES) incorporated [(3)H]UTP into RNA for 1h. Incorporation was inhibited 50% by 1mug of actinomycin D/ml, 1mug of acriflavine/ml and 0.5mug of ethidium bromide/ml but was insensitive to rifampicin, rifamycin SV, streptovarcin and deoxyribonuclease. After the first 10min of incubation, the synthesis was insensitive to ribonuclease. RNA synthesis by mitochondria isolated in sucrose-EDTA was insensitive to actinomycin D and sensitive to ribonuclease during the first 10min of the incubation but thereafter the sensitivities were the same as for mitochondria isolated in sucrose-TES. In a hypo-osmotic medium the relative extent of incorporation of the four ribonucleoside triphosphates into RNA was CTP>UTP=ATP>>GTP. In an iso-osmotic medium the incorporation of CTP and GTP decreased. All four nucleotides were incorporated into RNA in a DNA-dependent process, as indicated by the inhibition by actinomycin D. In addition, CTP and ATP were incorporated into the CCA end of mitochondrial tRNA. ATP was also incorporated into an unidentified acid-insoluble compound, which hydrolysed in alkali to a product that was not ATP, ADP or 5'- or 2(3')-AMP. Atractyloside inhibited the incorporation of ATP into RNA with 50% inhibition at 2-3nmol/mg of protein. The [(3)H]UTP-labelled RNA had peaks of 16S and 13S characteristic of mitochondrial rRNA. In addition a peak at 20-21S was observed as well as heterogeneous RNA sedimenting throughout the gradient. The synthesis of all these species was inhibited by actinomycin D, indicating that rat liver mitochondrial DNA codes for mitochondrial rRNA as well as other as yet unidentified species.
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PMID:Synthesis of ribonucleic acid by isolated rat liver mitochondria. 440 94

Poly(A) polymerase activity is induced during vaccinia virus infection of HeLa cells. The enzyme is maximally induced at 3.5 h postinfection. Partial purification frees the preparation of RNase activity and RNA polymerase activity. ATP is the substrate for poly(A) synthesis. A small amount of poly(A) is produced from added adenosine diphosphate due to the production of ATP by an adenylate kinase present in the preparation. The incorporation of ATP into poly(A) is dependent on divalent cations (Mg(2+) or Mn(2+)) and is not inhibited by UTP, CTP, or GTP. Poly(U) stimulates ATP incorporation; poly(A) and poly(C) have little effect on ATP incorporation, and poly(dT) is extremely inhibitory. RNA prepared from HeLa cells and from the partially purified poly(A) polymerase (the enzyme preparation contains endogenous RNA [Brakel and Kates]) stimulates ATP incorporation by poly(A) polymerase which was subjected to DEAE-cellulose chromatography. RNase's, pancreatic and T(1), inhibit the production of poly(A). DNase has little effect. Poly(U) is able to stimulate poly(A) production in the presence of T(1) RNase.
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PMID:Poly(A) polymerase from vaccinia virus-infected cells. I. Partial purification and characterization. 441 6


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