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)

Escherichia coli DNA-directed RNA polymerase is shown to contain a novel phosphorolytic error correction activity which removes erroneous nucleotides, as rNDPs, from the 3'-end of the growing transcript. The activity we describe is biochemically similar to polynucleotide phosphorylase (PNP), yet in contrast to PNP is activated by Mn2+. We demonstrate that the activity, which is mediated by Pi, is dependent on the presence of an incorrectly incorporated nucleotide at the leading 3'-end of the transcript. The correction activity we describe exhibits a 4 x 10(4)-fold preference for the excision of incorrect nucleotides from the transcript. These findings suggest the possibility that RNA phosphorolysis may play a critical role in the process of transcriptional proofreading.
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PMID:Phosphorolytic error correction during transcription. 752 Jan 15

The ADP analogue in which the 5'-oxygen has been replaced by a methylene group can be prepared by condensing 5'-deoxy-5'-phosphonomethyladenosine with inorganic phosphate. This analogue readily polymerizes onto the primer A-A in the presence of the enzyme polynucleotide phosphorylase and either Mg2+ or Mn2+. The initial products are of the form A-A(-cA)n-cA (where "-" and "-c" stand for the normal phosphodiester linkage and the linkage in which the 5'-oxygen is replaced with the methylene group, respectively). Treatment of these with alkali yields adenosine 2'(3')-phosphate and the series (A(-cA)n-cA containing only phosphonomethylene linkages. The decamer A(-cA)8-cA interacts with two molecules of U(-U)8-U to form a triple-standard structure that has a stability similar to that exhibited by the analogous complex formed from A(-A)8-A and U(-U)8-U. This property, along with the resistance of these oligomer analogues toward nucleases that cleave phosphodiester linkages between the phosphorus and the 5'-oxygen, should provide a strong rationale for application of phosphonomethylene linkages in schemes for therapeutic drug design that use the antisense strategy.
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PMID:Synthesis and properties of adenosine oligonucleotide analogues containing methylene groups in place of phosphodiester 5'-oxygens. 839 23

The pnpA gene of Bacillus subtilis, which codes for polynucleotide phosphorylase (PNPase), has been cloned and employed in the construction of pnpA deletion mutants. Growth defects of both B. subtilis and Escherichia coli PNPase-deficient strains were complemented with the cloned pnpA gene. RNA decay characteristics of the B. subtilis pnpA mutant were studied, including the in vivo decay of bulk mRNA and the in vitro decay of either poly(A) or total cellular RNA. The results showed that mRNA decay in the pnpA mutant is accomplished despite the absence of the major, Pi-dependent RNA decay activity of PNPase. In vitro experiments suggested that a previously identified, Mn2+ -dependent hydrolytic activity was important for decay in the pnpA mutant. In addition to a cold-sensitive-growth phenotype, the pnpA deletion mutant was found to be sensitive to growth in the presence of tetracycline, and this was due to an increased intracellular accumulation of the drug. The pnpA deletion strain also exhibited multiseptate, filamentous growth. It is hypothesized that defective processing of specific RNAs in the pnpA mutant results in these phenotypes.
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PMID:Properties of a Bacillus subtilis polynucleotide phosphorylase deletion strain. 863 41

Campylobacter jejuni is a foodborne bacterial pathogen, which is now considered as a leading cause of human bacterial gastroenteritis. The information regarding ribonucleases in C. jejuni is very scarce but there are hints that they can be instrumental in virulence mechanisms. Namely, PNPase (polynucleotide phosphorylase) was shown to allow survival of C. jejuni in refrigerated conditions, to facilitate bacterial swimming, cell adhesion, colonization and invasion. In several microorganisms PNPase synthesis is auto-controlled in an RNase III (ribonuclease III)-dependent mechanism. Thereby, we have cloned, overexpressed, purified and characterized Cj-RNase III (C. jejuni RNase III). We have demonstrated that Cj-RNase III is able to complement an Escherichia coli rnc-deficient strain in 30S rRNA processing and PNPase regulation. Cj-RNase III was shown to be active in an unexpectedly large range of conditions, and Mn2+ seems to be its preferred co-factor, contrarily to what was described for other RNase III orthologues. The results lead us to speculate that Cj-RNase III may have an important role under a Mn2+-rich environment. Mutational analysis strengthened the function of some residues in the catalytic mechanism of action of RNase III, which was shown to be conserved.
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PMID:Characterization of the biochemical properties of Campylobacter jejuni RNase III. 2407 28

Posttranscriptional gene regulation often involves RNA-binding proteins that modulate mRNA translation and/or stability either directly through protein-RNA interactions or indirectly by facilitating the annealing of small regulatory RNAs (sRNAs). The human pathogen Streptococcus pneumoniae D39 (pneumococcus) does not encode homologs to RNA-binding proteins known to be involved in promoting sRNA stability and function, such as Hfq or ProQ, even though it contains genes for at least 112 sRNAs. However, the pneumococcal genome contains genes for other RNA-binding proteins, including at least six S1 domain proteins: ribosomal protein S1 (rpsA), polynucleotide phosphorylase (pnpA), RNase R (rnr), and three proteins with unknown functions. Here, we characterize the function of one of these conserved, yet uncharacterized, S1 domain proteins, SPD_1366, which we have renamed CvfD (conserved virulence factor D), since loss of the protein results in attenuation of virulence in a murine pneumonia model. We report that deletion of cvfD impacts the expression of 144 transcripts, including the pst1 operon, encoding phosphate transport system 1 in S. pneumoniae We further show that CvfD posttranscriptionally regulates the PhoU2 master regulator of the pneumococcal dual-phosphate transport system by binding phoU2 mRNA and impacting PhoU2 translation. CvfD not only controls expression of phosphate transporter genes but also functions as a pleiotropic regulator that impacts cold sensitivity and the expression of sRNAs and genes involved in diverse cellular functions, including manganese uptake and zinc efflux. Together, our data show that CvfD exerts a broad impact on pneumococcal physiology and virulence, partly by posttranscriptional gene regulation.IMPORTANCE Recent advances have led to the identification of numerous sRNAs in the major human respiratory pathogen S. pneumoniae However, little is known about the functions of most sRNAs or RNA-binding proteins involved in RNA biology in pneumococcus. In this paper, we characterize the phenotypes and one target of the S1 domain RNA-binding protein CvfD, a homolog of general stress protein 13 identified, but not extensively characterized, in other Firmicutes species. Pneumococcal CvfD is a broadly pleiotropic regulator, whose absence results in misregulation of divalent cation homeostasis, reduced translation of the PhoU2 master regulator of phosphate uptake, altered metabolism and sRNA amounts, cold sensitivity, and attenuation of virulence. These findings underscore the critical roles of RNA biology in pneumococcal physiology and virulence.
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PMID:S1 Domain RNA-Binding Protein CvfD Is a New Posttranscriptional Regulator That Mediates Cold Sensitivity, Phosphate Transport, and Virulence in Streptococcus pneumoniae D39. 3260 Oct 68


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