Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.8 (polynucleotide phosphorylase)
 723 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Two procedures were investigated for the modification of tRNAs at the 3'-terminal nucleoside. The first involved the incubation of an enzymatically abreviated tRNA (tRNA-C-COH) with appropriate nucleoside triphosphates in the presence of CTP(ATP):tRNA nucleotidyltransferase from Escherichia coli and yeast. The E. coli enzyme did not utilize 2'- or 3'-deoxyadenosine 5'-triphosphate as substrates, but affected incorporation of the 2'- and 3'-O-methyladenosine triphosphates onto tRNA-C-Cou to the extent of 30 and 37%, respectively. Although incorporation of the deoxynucleotides could not be effected using the E. coli enzyme, yeast CTP(ATP:tRNA nucleotidyltransferase produced the desired tRNAs in yields of 45-65%. The second modification procedure involved incubation of tRNA-C-COH with (appropriately blocked) nucleoside diphosphates in the presence of polynucleotide phosphorylase. This procedure afforded the tRNAs terminating in 2'- and 3'-deoxyadenosine in yields of 4% (and the yield of the former was increased to 36% when the incubation was carried out in the presence of 20% methanol). The yields of tRNAs terminating in 2'- and 3'-O-methyladenosing produced by this procedure were 55 and 17%, respectively. Because only single isomers of most of the tRNAs terminating in 2'- and 3'-deoxy- and O-methyladenosine are aminoacylated, attempts were made to obtain the other isomericaminoacyl-tRNA by enzymatic introduction of chemically preaminoacylated nucleotides onto tRNA-C-COH. Although incubation of tRNA-C-COH with three aminoacylated nucleoside 5'-triphosphates and E. coli CTP(ATP):tRNA nucleotidyltransferase did not result in production of the desired tRNAs to a detectable extent, incubation with 2'-deoxy-3'-O-L-phenylalanyladenosine 5'-diphosphate and polynucleotide phosphorylase afforded E. coli tRNA terminating with the corresponding aminoacylated deoxynucleoside.
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PMID:Preparation of Escherichia coli tRNAs terminating of modified nucleosides by the use of CTP(ATP):tRNA nucleotidyltransferase and polynucleotide phosphorylase. 31 25

In the phosphorolytic degradation catalyzed by chicken liver PNPase (E.C. 2.4.2.1) inosine appears to behave as a better substrate than xanthosine. Hypoxanthine, xanthine, guanine and purine (1 X 10(-1)M) appear to be inhibitors of the pigeon liver PNPase, whereas allopurinol, ATP, ITP, CTP and UTP (1. X 10(-3) M) do not inhibit the enzyme. Both PNPase activities exhibit the same optimum temperature (37-40 degrees C). Chicken liver PNPase optimum pH is in the range 6.5-7, whereas that of pigeon liver is in the range 7-7.5. Lineweaver-Burk plots for the inosine phosphorolysis catalyzed by chicken liver PNPase yielded straight lines if substrate concentrations were lower than 1 X 10(-4) M but concave downward curves at higher concentrations. This activation increases when the homogenates are stored at 4 degrees C and pH = 7 during 24 h or more; pigeon liver PNPase does not show this activation phenomenon.
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PMID:[Purine metabolites in the activity of purine nucleoside phosphorylase (author's transl)]. 82 98

As a starting point for the study of the biosynthesis of polyadenylated RNA in bacteria, the characteristics of RNA synthesis by cells of Escherichia coli B made permeable to small molecules by treatment with toluene were examined. Such cells mediated the incorporation of radiolabeled ribonucleoside triphosphates into RNA in a reaction that was sensitive to inhibitors of RNA polymerase and required the simultaneous presence of the four ribonucleoside triphosphates. Between 10 to 15% of the RNA synthesized under these conditions was polyadenylated as shown by affinity chromatography on oligo(dT)-cellulose. The presence of orthophosphate or dADP, inhibitors of polynucleotide phosphorylase, had no effect on the reaction and the rate of RNA synthesis was indistinguishable in the polynucleotide phosphorylase-deficient strain PR-7 and in its otherwise isogenic parent strain PR-100. The poly(A) tracts associated with the newly synthesized RNA could be isolated after exhaustive digestion with pancreatic and T1 ribonucleases and accounted for 14% of the poly(A)-RNA. At least 74% of the poly(A) sequences were located at the 3' ends of RNA molecules and their weight-average length was 48 nucleotide residues. The size distribution of total RNA and poly(A)-RNA synthesized in the toluenized cell system was similar to that of the corresponding pulse-labeled fractions derived from growing cultures. The sequence complexity of poly(A)-RNA and unadenylated RNA synthesized in toluenized cells with [alpha-32P]CTP as the labeled substrate was analyzed by hybridization to fragments of Escherichia coli B DNA generated by digestion with EcoRI restriction endonuclease and immobilized on nitrocellulose sheets. Both RNA fractions hybridized with many DNA fractions, the hybridization patterns being similar with poly(A)-RNA and unadenylated RNA. This indicated that many different types of RNA transcripts synthesized in toluenized cells were subject to polyadenylation, but that polyadenylation was incomplete so that each transcript was present in both an adenylated and an unadenylated state.
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PMID:Synthesis of polyadenylate-containing RNA in vitro in permeable cells of Escherichia coli B. 619 64

1. By digitonin lysis of penicillin spheroplasts of Escherichia coli a particulate fraction P(1) was previously obtained that supported the sustained synthesis of alkaline phosphatase when supplied with amino acids, nucleotide triphosphates and other cofactors. This P(1) fraction, when subjected to mild ultrasonic treatment in the presence of sucrose and Mg(2+), yielded the P(1)(S) fraction, consisting of integrated particulate subcellular particles containing DNA and RNA. 2. The P(1)(S) fraction from E. coli K10 wild type (R(+) (1)R(+) (2)P(+)) grown under repressed conditions supported the immediate synthesis of alkaline phosphatase in vitro. The synthesis occurred in phases. The first was followed by a lag, and then there was a linear rapid phase that continued for at least 3hr. Actinomycin D inhibited the appearance of the second phase. It was concluded that the particles are programmed to synthesize enzyme even when prepared from repressed cells, and therefore that synthesis of the specific messenger RNA for alkaline phosphatase in vivo was not inhibited when the bacteria were grown in an excess of inorganic phosphate. 3. Phosphate inhibited synthesis of enzyme to the same extent with the P(1)(S) fractions of two constitutive strains as with the P(1)(S) fraction of the wild-type strain. 4. Inorganic phosphate inhibited amino acid incorporation with the P(1)(S) fraction and also inhibited enzyme synthesis in vitro. The effect on amino acid incorporation could be partially overcome by adding Mn(2+) to the incubation mixtures. However, Mn(2+) inhibited the synthesis of alkaline phosphatase. Also, inhibition of the incorporation of [(32)P]CTP into RNA was overcome by Mn(2+). The effect of phosphate on amino acid uptake was most probably due to a phosphorolysis of RNA by polynucleotide phosphorylase, also present in the P(1)(S) fraction. This phosphorolysis may be responsible for the instability of messenger RNA in vitro and in vivo. 5. Phosphate also specifically inhibited the formation of alkaline phosphatase, since it did not affect markedly the induced formation of beta-galactosidase by the same P(1)(S) fraction. The specific effect is attributed to the prevention of formation of the enzymically active dimer from precursors, a Zn(2+)-dependent reaction. It is suggested that the repression of the synthesis of alkaline phosphatase in vivo in the wild-type strain was the sum of these two effects.
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PMID:THE BIOSYNTHESIS OF ALKALINE PHOSPHATASE WITH A PARTICULATE FRACTION OF ESCHERICHIA COLI. 1433 60