scholarly journals RNA 3’-terminal phosphate cyclases and cyclase-like proteins

2016 ◽  
Vol 62 (3) ◽  
pp. 327-334
Author(s):  
Witold Filipowicz
Keyword(s):  
Rna Cleavage ◽  
Rna Molecules ◽  
Metabolic Reactions ◽  
Cyclic Phosphate ◽  
Cleavage Reactions

RNA molecules bearing terminal 2’,3’-cyclic phosphate are quite common in nature. For example, 2’,3’-cyclic phosphate termini are produced during RNA cleavage by many en-doribonucleases either as intermediates or final products. Many RNA-based nucleases (ribo-zymes) also generate cyclic phosphate termini. However, cleavage reactions are not the only way in which RNAs bearing cyclic phosphate ends are produced. They can also be generated by RNA 3’-terminal phosphate cyclases (RtcA), a family of enzymes conserved in eukaryotes, bacteria, and archaea. These enzymes catalyze the ATP-dependent conversion of the 3’-phos-phate to a 2’,3’-cyclic phosphodiester at the end of RNA. In this article, I review knowledge about the biochemistry and structure of RNA 3’-phosphate cyclases and also proteins of the RNA cyclase-like (Rcl1) family, and discuss their documented or possible roles in different RNA metabolic reactions.

scholarly journals Cyclases of the 3'-terminal phosphate in RNA: a new family of RNA processing enzymes conserved in eucarya, bacteria and archaea.

1998 ◽  
Vol 45 (4) ◽  
pp. 895-906 ◽  
Author(s):  
W Filipowicz ◽  
E Billy ◽  
K Drabikowski ◽  
P Genschik
Keyword(s):  
Escherichia Coli ◽  
Rna Processing ◽  
Rna Cleavage ◽  
Biological Functions ◽  
Rna Molecules ◽  
Cyclic Groups ◽  
Processing Enzymes ◽  
New Family ◽  
Genes Encoding ◽  
Cyclic Phosphate

The 2',3'-cyclic phosphate termini are produced, as either intermediates or final products, during RNA cleavage by many different endoribonucleases. Likewise, ribozymes such as hammerheads, hairpins, or the hepatitis delta ribozyme, generate 2',3'-cyclic phosphate ends. Discovery of the RNA 3'-terminal phosphate cyclase has indicated that cyclic phosphate termini in RNA can also be produced by an entirely different mechanism. The RNA 3'-phosphate cyclase converts the 3'-terminal phosphate in RNA into the 2',3'-cyclic phosphodiester in the ATP-dependent reaction which involves formation of the covalent cyclase-AMP and the RNA-N3' pp5' A intermediates. The findings that several eukaryotic and prokaryotic RNA ligases require the 2',3'-cyclic phosphate for the ligation of RNA molecules raised a possibility that the RNA 3'-phosphate cyclase may have an anabolic function in RNA metabolism by generating terminal cyclic groups required for ligation. Recent cloning of a cDNA encoding the human cyclase indicated that genes encoding cyclase-like proteins are conserved among Eucarya, Bacteria, and Archaea. The protein encoded by the Escherichia coli gene was overexpressed and shown to have the RNA 3'-phosphate cyclase activity. This article reviews properties of the human and bacterial cyclases, their mechanism of action and substrate specificity. Possible biological functions of the enzymes are also discussed.

scholarly journals The nonenzymatic template-directed ligation of oligonucleotides

2006 ◽  
Vol 3 (3) ◽  
pp. 243-249 ◽  
Author(s):  
A. V. Lutay ◽  
E. L. Chernolovskaya ◽  
M. A. Zenkova ◽  
V. V. Vlassov
Keyword(s):  
Metal Ion ◽  
Divalent Cations ◽  
Bound Water ◽  
Transesterification Reaction ◽  
Water Molecules ◽  
Pka Values ◽  
Rna Molecules ◽  
High Reaction ◽  
Cyclic Phosphate ◽  
Cyclic Phosphates

Abstract. The nonenzymatic template-directed ligation of oligonucleotides containing 2', 3'-cyclic phosphate was investigated in the presence of divalent cations. Ligation of the oligonucleotides readily occurred in the presence of Mg2+, Mn2+, Co2+, Zn2+, Pb2+. Efficacy of the metal ion catalysts inversely correlated with pKa values of the metal-bound water molecules. The intermolecular transesterification reaction yielded at least 95metal ion. Relatively high reaction yields (up to 15fragmentation to oligonucleotides with 2',3'-cyclic phosphates, followed by reactions of those oligonucleotides could provide a source of new RNA molecules under prebiotic conditions.

scholarly journals The nonenzymatic template-directed ligation of oligonucleotides

2006 ◽  
Vol 3 (1) ◽  
pp. 1-21 ◽  
Author(s):  
A.V. Lutay ◽  
E. L. Chernolovskaya ◽  
M. A. Zenkova ◽  
V. V. Vlasso
Keyword(s):  
Metal Ion ◽  
Divalent Cations ◽  
Bound Water ◽  
Transesterification Reaction ◽  
Water Molecules ◽  
Pka Values ◽  
Rna Molecules ◽  
Phosphodiester Bonds ◽  
Cyclic Phosphate ◽  
Cyclic Phosphates

Abstract. The nonenzymatic template-directed ligation of oligonucleotides containing 2',3'-cyclic phosphate was investigated in the presence of divalent cations. Ligation of the oligonucleotides readily occurred in the presence of Mg2+, Mn2+, Co2+, Zn2+, Pb2+. Efficacy of the metal ion catalysts inversely correlated with pKa values of the metal-bound water molecules. The intermolecular transesterification reaction yielded at least 95% of 2',5'-phosphodiester bonds independently on the nature of the metal ion. Relatively high reaction yields (up to 15%) suggest, that RNA fragmentation to oligonucleotides with 2',3'-cyclic phosphates, followed by reactions of those oligonucleotides could provide a source of new RNA molecules under prebiotic conditions.

scholarly journals Classification of the nucleolytic ribozymes based upon catalytic mechanism

F1000Research ◽  
2019 ◽  
Vol 8 ◽  
pp. 1462
Author(s):  
David M.J. Lilley
Keyword(s):  
Catalytic Mechanism ◽  
Base Catalysis ◽  
Nucleophilic Attack ◽  
Rna Cleavage ◽  
Hydroxyl Groups ◽  
Acid Base ◽  
General Base ◽  
Cleavage Reactions ◽  
Mechanistic Basis

The nucleolytic ribozymes carry out site-specific RNA cleavage reactions by nucleophilic attack of the 2′-oxygen atom on the adjacent phosphorus with an acceleration of a million-fold or greater. A major part of this arises from concerted general acid–base catalysis. Recent identification of new ribozymes has expanded the group to a total of nine and this provides a new opportunity to identify sub-groupings according to the nature of the general base and acid. These include nucleobases, hydrated metal ions, and 2′-hydroxyl groups. Evolution has selected a number of different combinations of these elements that lead to efficient catalysis. These differences provide a new mechanistic basis for classifying these ribozymes.

Toehold mediated strand displacement to measure released product from self-cleaving ribozymes

RNA ◽  
2021 ◽  
pp. rna.078823.121
Author(s):  
Jay Bhakti Kapadia ◽  
Nawwaf Kharma ◽  
Alen Nellikulam Davis ◽  
Nicolas Kamel ◽  
Jonathan Perreault
Keyword(s):  
Hammerhead Ribozyme ◽  
Polyacrylamide Gels ◽  
Strand Displacement ◽  
Rna Molecules ◽  
Starting Point ◽  
Dna Strands ◽  
Regular Sampling ◽  
Method Of Measurement ◽  
Cleavage Reactions ◽  
Labeled Rna

This paper presents a probe comprising a fluorophore and a quencher, enabling measurement of released product from self-cleaving hammerhead ribozyme, without labeled RNA molecules, regular sampling or use of polyacrylamide gels. The probe is made of two DNA strands; one strand is labelled with a fluorophore at its 5′-end, while the other strand is labelled with a quencher at its 3′-end. These two DNA strands are perfectly complementary, but with a 3′-overhang of the fluorophore strand. These unpaired nucleotides act as a toehold, which is utilized by a detached cleaved fragment (coming from a self-cleaving hammerhead ribozyme) as the starting point for a strand displacement reaction. This reaction causes the separation of the fluorophore strand from the quencher strand, culminating in fluorescence, detectable in a plate reader. Notably, the emitted fluorescence is proportional to the amount of detached cleaved-off RNAs, displacing the DNA quencher strand. This method can replace or complement radio-hazardous unstable 32P as a method of measurement of the product release from ribozyme cleavage reactions; it also eliminates the need for polyacrylamide gels, for the same purpose. Critically, this method allows to distinguish between the total amount of cleaved ribozymes and the amount of detached fragments, resulting from that cleavage reaction.

Characterization of Two RNA-catalyzed RNA Cleavage Reactions

1987 ◽  
Vol 52 (0) ◽  
pp. 267-275 ◽  
Author(s):  
J.R. Sampson ◽  
F.X. Sullivan ◽  
L.S. Behlen ◽  
A.B. DiRenzo ◽  
O.C. Uhlenbeck
Keyword(s):  
Rna Cleavage ◽  
Cleavage Reactions

scholarly journals Reconstitution of mammalian Cleavage Factor II involved in 3’ processing of mRNA precursors

2018 ◽  
Author(s):  
Peter Schäfer ◽  
Christian Tüting ◽  
Lars Schönemann ◽  
Uwe Kühn ◽  
Thomas Treiber ◽  
...  
Keyword(s):  
Protein Complex ◽  
Kinase Activity ◽  
Zinc Fingers ◽  
Rna Cleavage ◽  
Affinity Binding ◽  
Sequence Specificity ◽  
Factor Ii ◽  
Sequence Elements ◽  
Cleavage And Polyadenylation ◽  
Cleavage Reactions

AbstractCleavage factor II (CF II) is a poorly characterized component of the multi-protein complex catalyzing 3’ cleavage and polyadenylation of mammalian mRNA precursors. We have reconstituted CF II as a heterodimer of hPcf11 and hClp1. The heterodimer is active in partially reconstituted cleavage reactions, whereas hClp1 by itself is not. Pcf11 moderately stimulates the RNA 5’ kinase activity of hClp1; the kinase activity is dispensable for RNA cleavage. CF II binds RNA with nanomolar affinity. Binding is mediated mostly by the two zinc fingers in the C-terminal region of hPcf11. RNA is bound without pronounced sequence-specificity, but extended G-rich sequences appear to be preferred. We discuss the possibility that CF II contributes to the recognition of cleavage/polyadenylation substrates through interaction with G-rich far-downstream sequence elements.

scholarly journals Measurement of Kinetics of Hammerhead Ribozyme Cleavage Reactions using Toehold Mediated Strand Displacement

2020 ◽  
Author(s):  
Jay Bhakti Kapadia ◽  
Nawwaf Kharma ◽  
Alen Nellikulam Davis ◽  
Nicolas Kamel ◽  
Jonathan Perreault
Keyword(s):  
Hammerhead Ribozyme ◽  
Polyacrylamide Gels ◽  
Strand Displacement ◽  
Rna Molecules ◽  
Starting Point ◽  
Dna Strands ◽  
Regular Sampling ◽  
Cleavage Reactions ◽  
Labeled Rna ◽  
Kinetics Of

ABSTRACTThis paper presents a probe comprising a fluorophore and a quencher, enabling measurement of hammerhead ribozyme cleavage reactions, without labeled RNA molecules, regular sampling or use of polyacrylamide gels. The probe is made of two DNA strands; one strand is labelled with a fluorophore at its 5’-end, while the other strand is labelled with a quencher at its 3’-end. These two DNA strands are perfectly complementary, but with a 3’-overhang of the fluorophore strand. These unpaired nucleotides act as a toehold, which is utilized by a detached cleaved fragment (coming from a self-cleaving hammerhead ribozyme) as the starting point for a strand displacement reaction. This reaction causes the separation of the fluorophore strand from the quencher strand, culminating in fluorescence, detectable in a plate reader. Notably, the emitted fluorescence is proportional to the amount of detached cleaved-off RNAs, displacing the DNA quencher strand. This method can replace or complement radio-hazardous unstable 32P as a method of measurement of the kinetics of ribozyme cleavage reactions; it also eliminates the need for polyacrylamide gels, for the same purpose. Critically, this method allows to distinguish between the total amount of cleaved ribozymes and the amount of detached fragments, resulting from that cleavage reaction.

scholarly journals The Old and New RNA World

2014 ◽  
Vol 83 (4) ◽  
pp. 441-448
Author(s):  
Zofia Szweykowska-Kulińska
Keyword(s):  
Gene Expression ◽  
Genetic Information ◽  
Environmental Changes ◽  
Rna World ◽  
Chromatin Condensation ◽  
Feedback Regulation ◽  
Information Storage ◽  
Rna Molecules ◽  
Metabolic Reactions ◽  
Almost All

Among the numerous hypotheses offering a scenario for the origin of life on Earth, the one called “The RNA World” has gained the most attention. According to this hypothesis RNA acted as a genetic information storage material, as a catalyst of all metabolic reactions, and as a regulator of all processes in the primordial world. Various experiments show that RNA molecules could have been synthesized abiotically, with the potential to mediate a whole repertoire of metabolic reactions. Ribozymes carrying out aminoacyl-tRNA reactions have been found in SELEX (systematic evolution of ligands by exponential enrichment) approaches and the development of a ribosome from a RNA-built protoribosome is easy to imagine. Transfer RNA aminoacylation, protoribosome origin, and the availability of amino acids on early Earth allowed the genetic code to evolve. Encoded proteins most likely stabilized RNA molecules and were able to create channels across membranes. In the modern cell, DNA replaced RNA as the main depositor of genetic information and proteins carry out almost all metabolic reactions. However, RNA is still playing versatile, crucial roles in the cell. Apart from its classical functions in the cell, a huge small RNA world is controlling gene expression, chromatin condensation, response to environmental cues, and protecting the cell against the invasion of various nucleic acids forms. Long non-coding RNAs act as crucial gene expression regulators. Riboswitches act at the level of transcription, splicing or translation and mediate feedback regulation on biosynthesis and transport of the ligand they sense. Alternative splicing generates genetic variability and increases the protein repertoire in response to developmental or environmental changes. All these regulatory functions are essential in shaping cell plasticity in the changing milieu. Recent discoveries of new, unexpected and important functions of RNA molecules support the hypothesis that we live in a New RNA World.

scholarly journals DNA-dependent RNA cleavage by the Natronobacterium gregoryi Argonaute

2017 ◽  
Author(s):  
Sunghyeok Ye ◽  
Taegeun Bae ◽  
Kyoungmi Kim ◽  
Omer Habib ◽  
Seung Hwan Lee ◽  
...  
Keyword(s):  
Physiological Role ◽  
Eukaryotic Cells ◽  
Rna Cleavage ◽  
Rna Molecules ◽  
Rna Targets

AbstractWe show here that, unlike most other prokaryotic Argonaute (Ago) proteins, which are DNA-guided endonucleases, the Natronobacterium gregoryi-derived Ago (NgAgo) can function as a DNA-guided endoribonuclease, cleaving RNA, rather than DNA, in a targeted manner. The NgAgo protein, in complex with 5’-hydroxylated or 5’-phosphrylated oligodeoxyribonucleotides (ODNs) of variable lengths, split RNA targets into two or more fragments in vitro, suggesting its physiological role in bacteria and demonstrating a potential for degrading RNA molecules such as mRNA or lncRNA in eukaryotic cells in a targeted manner.

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