scholarly journals The Escherichia coli major exoribonuclease RNase II is a component of the RNA degradosome

2014 ◽  
Vol 34 (6) ◽  
Author(s):  
Feng Lu ◽  
Aziz Taghbalout
Keyword(s):  
Escherichia Coli ◽  
Rna Processing ◽  
Binding Sites ◽  
Rna Degradation ◽  
The Other ◽  
Polynucleotide Phosphorylase ◽  
Cell Extracts ◽  
Domains Of Life ◽  
Rna Degradosome ◽  
Rnase Ii

Multiprotein complexes that carry out RNA degradation and processing functions are found in cells from all domains of life. In Escherichia coli, the RNA degradosome, a four-protein complex, is required for normal RNA degradation and processing. In addition to the degradosome complex, the cell contains other ribonucleases that also play important roles in RNA processing and/or degradation. Whether the other ribonucleases are associated with the degradosome or function independently is not known. In the present work, IP (immunoprecipitation) studies from cell extracts showed that the major hydrolytic exoribonuclease RNase II is associated with the known degradosome components RNaseE (endoribonuclease E), RhlB (RNA helicase B), PNPase (polynucleotide phosphorylase) and Eno (enolase). Further evidence for the RNase II-degradosome association came from the binding of RNase II to purified RNaseE in far western affinity blot experiments. Formation of the RNase II–degradosome complex required the degradosomal proteins RhlB and PNPase as well as a C-terminal domain of RNaseE that contains binding sites for the other degradosomal proteins. This shows that the RNase II is a component of the RNA degradosome complex, a previously unrecognized association that is likely to play a role in coupling and coordinating the multiple elements of the RNA degradation pathways.

scholarly journals The Pyrimidine Nucleotide Reductase Step in Riboflavin and F420 Biosynthesis in Archaea Proceeds by the Eukaryotic Route to Riboflavin

2002 ◽  
Vol 184 (7) ◽  
pp. 1952-1957 ◽  
Author(s):  
Marion Graupner ◽  
Huimin Xu ◽  
Robert H. White
Keyword(s):  
Escherichia Coli ◽  
Gene Product ◽  
Pyridine Nucleotide ◽  
The Other ◽  
Riboflavin Biosynthesis ◽  
Methanosarcina Thermophila ◽  
Pyrimidine Nucleotide ◽  
Cell Extracts ◽  
Phosphate Compound ◽  
Its Gene

ABSTRACT The Methanococcus jannaschii gene MJ0671 was cloned and overexpressed in Escherichia coli, and its gene product was tested for its ability to catalyze the pyridine nucleotide-dependent reduction of either 2,5-diamino-6-ribosylamino-4(3H)-pyrimidinone 5′-phosphate (compound 3) to 2,5-diamino-6-ribitylamino-4(3H)-pyrimidinone 5′-phosphate (compound 4) or 5-amino-6-ribosylamino-2,4(1H,3H)-pyrimidinedione 5′-phosphate (compound 7) to 5-amino-6-ribitylamino-2,4(1H,3H)-pyrimidinedione 5′-phosphate (compound 5). Only compound 3 was found to serve as a substrate for the enzyme. NADPH and NADH functioned equally well as the reductants. This specificity for the reduction of compound 3 was also confirmed by using cell extracts of M. jannaschii and Methanosarcina thermophila. Thus, this step in riboflavin biosynthesis in these archaea is the same as that found in yeasts. The absence of the other genes in the biosynthesis of riboflavin in Archaea is discussed.

scholarly journals Structural elucidation of a novel mechanism for the bacteriophage-based inhibition of the RNA degradosome

eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
An Van den Bossche ◽  
Steven W Hardwick ◽  
Pieter-Jan Ceyssens ◽  
Hanne Hendrix ◽  
Marleen Voet ◽  
...  
Keyword(s):  
Rna Binding ◽  
Rna Degradation ◽  
Rnase E ◽  
Effective Inhibition ◽  
Domains Of Life ◽  
Infected Cells ◽  
Rna Degradosome ◽  
Host Infection ◽  
Structural Homologues ◽  
Unique Mechanism

In all domains of life, the catalysed degradation of RNA facilitates rapid adaptation to changing environmental conditions, while destruction of foreign RNA is an important mechanism to prevent host infection. We have identified a virus-encoded protein termed gp37/Dip, which directly binds and inhibits the RNA degradation machinery of its bacterial host. Encoded by giant phage фKZ, this protein associates with two RNA binding sites of the RNase E component of the Pseudomonas aeruginosa RNA degradosome, occluding them from substrates and resulting in effective inhibition of RNA degradation and processing. The 2.2 Å crystal structure reveals that this novel homo-dimeric protein has no identifiable structural homologues. Our biochemical data indicate that acidic patches on the convex outer surface bind RNase E. Through the activity of Dip, фKZ has evolved a unique mechanism to down regulate a key metabolic process of its host to allow accumulation of viral RNA in infected cells.

scholarly journals RNase II binds to RNase E and modulates its endoribonucleolytic activity in the cyanobacterium Anabaena PCC 7120

2020 ◽  
Vol 48 (7) ◽  
pp. 3922-3934 ◽  
Author(s):  
Cong Zhou ◽  
Juyuan Zhang ◽  
Xinyu Hu ◽  
Changchang Li ◽  
Li Wang ◽  
...  
Keyword(s):  
Catalytic Domain ◽  
Bacterial Species ◽  
Rna Degradation ◽  
Rnase E ◽  
Anabaena Pcc 7120 ◽  
Cyanobacterium Anabaena ◽  
Rna Degradosome ◽  
Rnase Ii ◽  
Pcc 7120

Abstract In Escherichia coli, the endoribonuclease E (RNase E) can recruit several other ribonucleases and regulatory proteins via its noncatalytic domain to form an RNA degradosome that controls cellular RNA turnover. Similar RNA degradation complexes have been found in other bacteria; however, their compositions are varied among different bacterial species. In cyanobacteria, only the exoribonuclease PNPase was shown to bind to the noncatalytic domain of RNase E. Here, we showed that Alr1240, a member of the RNB family of exoribonucleases, could be co-isolated with RNase E from the lysate of the cyanobacterium Anabaena PCC 7120. Enzymatic analysis revealed that Alr1240 is an exoribonuclease II (RNase II), as it only degrades non-structured single-stranded RNA substrates. In contrast to known RNase E-interacting ribonucleases, which bind to the noncatalytic domain of RNase E, the Anabaena RNase II was shown to associate with the catalytic domain of RNase E. Using a strain in which RNase E and RNase II were tagged in situ with GFP and BFP, respectively, we showed that RNase E and RNase II form a compact complex in vivo by a fluorescence resonance energy transfer (FRET) assay. RNase E activity on several synthetic substrates was boosted in the presence of RNase II, suggesting that the activity of RNase E could be regulated by RNase II-RNase E interaction. To our knowledge, Anabaena RNase II is an unusual ribonuclease that interacts with the catalytic domain of RNase E, and it may represent a new type of RNA degradosome and a novel mechanism for regulating the activity of the RNA degradosome. As Anabaena RNase E interacts with RNase II and PNPase via different regions, it is very likely that the three ribonucleases form a large complex and cooperatively regulate RNA metabolism in the cell.

scholarly journals Ancient Transcription Factors in the News

mBio ◽  
2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Irina Artsimovitch ◽  
Stefan H. Knauer
Keyword(s):  
Transcription Factors ◽  
Nucleic Acids ◽  
Rna Polymerase ◽  
Rna Processing ◽  
Binding Sites ◽  
Rna Synthesis ◽  
Mechanistic Studies ◽  
Elongation Complex ◽  
Content Type ◽  
Domains Of Life

ABSTRACTIn every cell from bacteria to mammals, NusG-like proteins bind transcribing RNA polymerase to modulate the rate of nascent RNA synthesis and to coordinate it with numerous cotranscriptional processes that ultimately determine the transcript fate. Housekeeping NusG factors regulate expression of the bulk of the genome, whereas their highly specialized paralogs control just a few targets. InEscherichiacoli, NusG stimulates silencing of horizontally acquired genes, while its paralog RfaH counters NusG action by activating a subset of these genes. Acting alone or as part of regulatory complexes, NusG factors can promote uninterrupted RNA synthesis, bring about transcription pausing or premature termination, modulate RNA processing, and facilitate translation. Recent structural and mechanistic studies of NusG homologs from all domains of life reveal molecular details of multifaceted interactions that underpin their unexpectedly diverse regulatory roles. NusG proteins share conserved binding sites on RNA polymerase and many effects on the transcription elongation complex but differ in their mechanisms of recruitment, interactions with nucleic acids and secondary partners, and regulatory outcomes. Strikingly, some can alternate between autoinhibited and activated states that possess dramatically different secondary structures to achieve exquisite target specificity.

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