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:2.7.7.8 (polynucleotide phosphorylase)
723 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Photorhabdus sp. strain K122 was found to produce higher levels of the protein CAP87K when cultured at 9 degrees C than when cultured at 28 degrees C. NH2-terminal sequencing of this protein revealed homology with the NH2 terminus of Escherichia coli polynucleotide phosphorylase. A 4.5-kb DNA fragment from strain K122 was cloned and sequenced and found to have 75% identity to the E. coli rpsO-pnp operon coding for ribosomal protein S15 and polynucleotide phosphorylase, respectively. Predicted proteins encoded by this sequence were found to have 86% identity with ribosomal protein S15 and polynucleotide phosphorylase from E. coli, and the genes were called rpsO and pnp, respectively. Quantitation of rpsO and pnp mRNA transcripts from K122 revealed that there was a 2.4-fold increase in the level of pnp mRNA and a 1.9-fold decrease in the level of rpsO mRNA at 9 degrees C relative to 28 degrees C. Primer extension analysis revealed the positions of possible promoters controlling the expression of rpsO and pnp in K122, suggesting that the genes are expressed independently. The increase in the level of pnp mRNA at 9 degrees C was not due to any relative increase in its stability compared with that of the rpsO transcript. However, there was evidence to suggest that it may be a result of a cold-inducible promoter, P2, in the intergenic region between rpsO and pnp. Several features of P2 support the suggestion that it may be cold inducible.
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PMID:The gene coding for polynucleotide phosphorylase in Photorhabdus sp. strain K122 is induced at low temperatures. 820 56

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

The S1 domain, originally identified in ribosomal protein S1, is found in a large number of RNA-associated proteins. The structure of the S1 RNA-binding domain from the E. coli polynucleotide phosphorylase has been determined using NMR methods and consists of a five-stranded antiparallel beta barrel. Conserved residues on one face of the barrel and adjacent loops form the putative RNA-binding site. The structure of the S1 domain is very similar to that of cold shock protein, suggesting that they are both derived from an ancient nucleic acid-binding protein. Enhanced sequence searches reveal hitherto unidentified S1 domains in RNase E, RNase II, NusA, EMB-5, and other proteins.
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PMID:The solution structure of the S1 RNA binding domain: a member of an ancient nucleic acid-binding fold. 900 64

Escherichia coli cells lacking both polynucleotide phosphorylase (PNPase) and RNase PH, the only known P(i)-dependent exoribonucleases, were previously shown to grow slowly at 37 degrees C and to display a dramatically reduced level of tRNA(Tyr)su3+ suppressor activity. Here we show that the RNase PH-negative, PNP-negative double-mutant strain actually displays a reversible cold-sensitive phenotype and that tRNA biosynthesis is normal. In contrast, ribosome structure and function are severely affected, particularly at lower temperatures. At 31 degrees C, the amount of 50S subunit is dramatically reduced and 23S rRNA is degraded. Moreover, cells that had been incubated at 42 degrees C immediately cease growing and synthesizing protein upon a shift to 31 degrees C, suggesting that the ribosomes synthesized at the higher temperature are defective and unable to function at the lower temperature. These data indicate that RNase PH and PNPase play an essential role that affects ribosome metabolism and that this function cannot be taken over by any of the hydrolytic exoribonucleases present in the cell.
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PMID:An essential function for the phosphate-dependent exoribonucleases RNase PH and polynucleotide phosphorylase. 920 58

The psychrotrophic bacterium Yersinia enterocolitica is characterized by temperature-dependent adaptations. To investigate Y. enterocolitica genes involved in cold adaptation, a mutant restricted in its ability to grow at 5 degrees C was isolated from a transposon mutant library. The transposon insertion site in this psychrotrophy-defective (PD) mutant mapped 16 bp upstream of an open reading frame whose predicted amino acid sequence showed 93% similarity with the Escherichia coli exoribonuclease polynucleotide phosphorylase (PNPase), encoded by pnp. Expression of this gene was blocked in the PD mutant. However, the introduction of a second copy of pnp, including 0.33 kbp sequences upstream of its coding region, into the chromosome of the PD mutant restored pnp expression as well as the ability to grow at 5 degrees C. Furthermore, the expression of pnp appeared to be temperature dependent: in the parental Y. enterocolitica strain, the levels of both pnp mRNA and PNPase were 1.6-fold higher at 5 degrees C compared with 30 degrees C. A similarly enhanced level of PNPase at 5 degrees C was observed in the merodiploid recombinant strain, which indicates that the 0.33 kbp region upstream of pnp harboured a cold-inducible promoter. A putative cold shock promoter motif (ATTGG) was observed in this region.
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PMID:The psychrotrophic bacterium Yersinia enterocolitica requires expression of pnp, the gene for polynucleotide phosphorylase, for growth at low temperature (5 degrees C). 963 58

The cellular content of major cold shock protein (MCSP) mRNA transcribed from the tandem gene duplication cspA1/A2 and growth of Yersinia enterocolitica were compared when exponentially growing cultures of this bacterium were cold shocked from 30 to 20, 15, 10, 5, or 0 degrees C, respectively. A clear correlation between the time point when exponential growth resumes after cold shock and the degradation of cspA1/A2 mRNA was found. A polynucleotide phosphorylase-deficient mutant was unable to degrade cspA1/A2 mRNA properly and showed a delay, as well as a lower rate, of growth after cold shock. For this mutant, a correlation between decreasing cspA1/A2 mRNA and restart of growth after cold shock was also observed. For both wild-type and mutant cells, no correlation of restart of growth with the cellular content of MCSPs was found. We suggest that, after synthesis of cold shock proteins and cold adaptation of the cells, MCSP mRNAs must be degraded; otherwise, they trap ribosomes, prevent translation of bulk mRNA, and thus inhibit growth of this bacterium at low temperatures.
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PMID:Restart of exponential growth of cold-shocked Yersinia enterocolitica occurs after down-regulation of cspA1/A2 mRNA. 1080 13

Polynucleotide phosphorylase (PNPase, polyribonucleotide nucleotidyltransferase, EC 2.7.7.8) is one of the cold shock-induced proteins in Escherichia coli and pnp, the gene encoding it, is essential for growth at low temperatures. We have analysed the expression of pnp upon cold shock and found a dramatic transient variation of pnp transcription profile: within the first hour after temperature downshift the amount of pnp transcripts detectable by Northern blotting increased more than 10-fold and new mRNA species that cover pnp and the downstream region, including the cold shock gene deaD, appeared; 2 h after temperature downshift the transcription profile reverted to a preshift-like pattern in a PNPase-independent manner. The higher amount of pnp transcripts appeared to be mainly due to an increased stability of the RNAs. The abundance of pnp transcripts was not paralleled by comparable variation of the protein: PNPase steadily increased about twofold during the first 3 h at low temperature, as determined both by Western blotting and enzymatic activity assay, suggesting that PNPase, unlike other known cold shock proteins, is not efficiently translated in the acclimation phase. In experiments aimed at assessing the role of PNPase in autogenous control during cold shock, we detected a Rho-dependent termination site within pnp. In the cold acclimation phase, termination at this site depended upon the presence of PNPase, suggesting that during cold shock pnp is autogenously regulated at the level of transcription elongation.
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PMID:Transcriptional and post-transcriptional control of polynucleotide phosphorylase during cold acclimation in Escherichia coli. 1093 Dec 96

When Escherichia coli cells are shifted to low temperatures (e.g. 15 degrees C), growth halts while the 'cold shock response' (CSR) genes are induced, after which growth resumes. One CSR gene, pnp, encodes polynucleotide phosphorylase (PNPase), a 3'-exoribonuclease and component of the RNA degradosome. At 37 degrees C, ribonuclease III (RNase III, encoded by rnc) cleaves the pnp untranslated leader, whereupon PNPase represses its own translation by an unknown mechanism. Here, we show that PNPase cold-temperature induction involves several post-transcriptional events, all of which require the intact pnp mRNA leader. The bulk of induction results from reversal of autoregulation at a step subsequent to RNase III cleavage of the pnp leader. We also found that pnp translation occurs throughout cold-temperature adaptation, whereas lacZ(+) translation was delayed. This difference is striking, as both mRNAs are greatly stabilized upon the shift to 15 degrees C. However, unlike the lacZ(+) mRNA, which remains stable during adaptation, pnp mRNA decay accelerates. Together with other evidence, these results suggest that mRNA is generally stabilized upon a shift to cold temperatures, but that a CSR mRNA-specific decay process is initiated during adaptation.
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PMID:Cold-temperature induction of Escherichia coli polynucleotide phosphorylase occurs by reversal of its autoregulation. 1112 93

Upon cold shock, Escherichia coli cell growth transiently stops. During this acclimation phase, specific cold shock proteins (CSPs) are highly induced. At the end of the acclimation phase, their synthesis is reduced to new basal levels, while the non-cold shock protein synthesis is resumed, resulting in cell growth reinitiation. Here, we report that polynucleotide phosphorylase (PNPase) is required to repress CSP production at the end of the acclimation phase. A pnp mutant, upon cold shock, maintained a high level of CSPs even after 24 h. PNPase was found to be essential for selective degradation of CSP mRNAs at 15 degrees C. In a poly(A) polymerase mutant and a CsdA RNA helicase mutant, CSP expression upon cold shock was significantly prolonged, indicating that PNPase in concert with poly(A) polymerase and CsdA RNA helicase plays a critical role in cold shock adaptation.
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PMID:Selective mRNA degradation by polynucleotide phosphorylase in cold shock adaptation in Escherichia coli. 1129

Escherichia coli contains a large CspA family, CspA to CspI. Here, we demonstrate that E. coli is highly protected against cold-shock stress, as these CspA homologues existed at approximately a total of two million molecules per cell at low temperature and growth defect was not observed until four csp genes (cspA, cspB, cspE and cspG) were deleted. The quadruple-deletion strain acquired cold sensitivity and formed filamentous cells at 15 degrees C although chromosomes were normally segregated. The cold-sensitivity and filamentation phenotypes were suppressed by all members of the CspA family except for CspD, which causes lethality upon overexpression. Interestingly, the cold sensitivity of the mutant was also suppressed by the S1 domain of polynucleotide phosphorylase (PNPase), which also folds into a beta-barrel structure similar to that of CspA. The present results show that cold-shock proteins and S1 domains share not only the tertiary structural similarity but also common functional properties, suggesting that these seemingly distinct protein categories may have evolved from a common primordial RNA-binding protein.
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PMID:Acquirement of cold sensitivity by quadruple deletion of the cspA family and its suppression by PNPase S1 domain in Escherichia coli. 1129 85


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