-
The Journal of Biological Chemistry Feb 2020The ribosome and RNase P are cellular ribonucleoprotein complexes that perform peptide bond synthesis and phosphodiester bond cleavage, respectively. Both are ancient... (Review)
Review
The ribosome and RNase P are cellular ribonucleoprotein complexes that perform peptide bond synthesis and phosphodiester bond cleavage, respectively. Both are ancient biological assemblies that were already present in the last universal common ancestor of all life. The large subunit rRNA in the ribosome and the RNA subunit of RNase P are the ribozyme components required for catalysis. Here, we explore the idea that these two large ribozymes may have begun their evolutionary odyssey as an assemblage of RNA "fragments" smaller than the contemporary full-length versions and that they transitioned through distinct stages along a pathway that may also be relevant for the evolution of other non-coding RNAs.
Topics: Evolution, Molecular; Models, Molecular; RNA, Catalytic; RNA, Ribosomal; Ribonuclease P
PubMed: 31953324
DOI: 10.1074/jbc.REV119.009929 -
The Journal of Investigative... Dec 2002Since their initial discovery, ribozymes have shown great promise not just as a tool in the manipulation of gene expression, but also as a novel therapeutic agent. This... (Review)
Review
Since their initial discovery, ribozymes have shown great promise not just as a tool in the manipulation of gene expression, but also as a novel therapeutic agent. This review discusses the promises and pitfalls of ribozyme technology, with a special emphasis on cancer-related applications, though relevance to skin disease will also be discussed.
Topics: Animals; Drug Delivery Systems; Drug Industry; Humans; RNA, Catalytic; Skin Diseases; Skin Neoplasms
PubMed: 12518796
DOI: 10.1046/j.1523-1747.2002.19642.x -
Current Opinion in Structural Biology Dec 1998Our understanding of the relationship between the structure of RNA and its catalytic activity has advanced significantly in the past year. These advances include... (Review)
Review
Our understanding of the relationship between the structure of RNA and its catalytic activity has advanced significantly in the past year. These advances include time-resolved crystallographic studies on the hammerhead ribozyme, as well as new structures of a group I intron, a lead(II)-cleavage ribozyme, a hepatitis delta virus ribozyme, and components of the spliceosome machinery and the peptidyl transferase center of the ribosome and, most significantly, the structure of the ribosome itself.
Topics: Catalysis; Crystallography, X-Ray; Magnetic Resonance Spectroscopy; Protein Conformation; RNA Splicing; RNA, Catalytic
PubMed: 9914252
DOI: 10.1016/s0959-440x(98)80091-2 -
RNA (New York, N.Y.) May 2012The hammerhead ribozyme is a small catalytic RNA motif capable of endonucleolytic (self-) cleavage. It is composed of a catalytic core of conserved nucleotides flanked... (Review)
Review
The hammerhead ribozyme is a small catalytic RNA motif capable of endonucleolytic (self-) cleavage. It is composed of a catalytic core of conserved nucleotides flanked by three helices, two of which form essential tertiary interactions for fast self-scission under physiological conditions. Originally discovered in subviral plant pathogens, its presence in several eukaryotic genomes has been reported since. More recently, this catalytic RNA motif has been shown to reside in a large number of genomes. We review the different approaches in discovering these new hammerhead ribozyme sequences and discuss possible biological functions of the genomic motifs.
Topics: Genetic Variation; Genome; Nucleic Acid Conformation; Nucleotide Motifs; RNA, Catalytic; Sequence Homology; Tandem Repeat Sequences
PubMed: 22454536
DOI: 10.1261/rna.031401.111 -
Methods (San Diego, Calif.) May 2019A large number of catalytic RNAs, or ribozymes, have been identified in the genomes of various organisms and viruses. Ribozymes are involved in biological processes such... (Review)
Review
A large number of catalytic RNAs, or ribozymes, have been identified in the genomes of various organisms and viruses. Ribozymes are involved in biological processes such as regulation of gene expression and viral replication, but biological roles of many ribozymes still remain unknown. Ribozymes have also inspired researchers to engineer synthetic ribozymes that function as sensors or gene switches. To gain deeper understanding of the sequence-function relationship of ribozymes and to efficiently engineer synthetic ribozymes, a large number of ribozyme variants need to be examined which was limited to hundreds of sequences by Sanger sequencing. The advent of high-throughput sequencing technologies, however, has allowed us to sequence millions of ribozyme sequences at low cost. This review focuses on the recent applications of high-throughput sequencing to both characterize and engineer ribozymes, to highlight how the large-scale sequence data can advance ribozyme research and engineering.
Topics: Animals; Genetic Engineering; High-Throughput Nucleotide Sequencing; Humans; Mutation; RNA, Catalytic
PubMed: 30738128
DOI: 10.1016/j.ymeth.2019.02.001 -
Progress in Molecular Biology and... 2013The hammerhead ribozyme has long been considered a prototype for understanding RNA catalysis, but discrepancies between the earlier crystal structures of a minimal... (Review)
Review
The hammerhead ribozyme has long been considered a prototype for understanding RNA catalysis, but discrepancies between the earlier crystal structures of a minimal hammerhead self-cleaving motif and various biochemical investigations frustrated attempt to understand hammerhead ribozyme catalysis in terms of structure. With the discovery that a tertiary contact distal from the ribozyme's active site greatly enhances its catalytic prowess, and the emergence of new corresponding crystal structures of full-length hammerhead ribozymes, a unified understanding of catalysis in terms of the structure is now possible. A mechanism in which the invariant residue G12 functions as a general base, and the 2'-OH moiety of the invariant G8, itself forming a tertiary base pair with the invariant C3, is the general acid, appears consistent with both the crystal structure and biochemical experimental results. Originally discovered in the context of plant satellite RNA viruses, the hammerhead more recently has been found embedded in the 3'-untranslated region of mature mammalian mRNAs, suggesting additional biological roles in genetic regulation.
Topics: Animals; Biocatalysis; Humans; Nucleic Acid Conformation; RNA, Catalytic
PubMed: 24156940
DOI: 10.1016/B978-0-12-381286-5.00001-9 -
Viruses Mar 2019Current cellular facts allow us to follow the link from chemical to biochemical metabolites, from the ancient to the modern world. In this context, the "RNA world"...
Current cellular facts allow us to follow the link from chemical to biochemical metabolites, from the ancient to the modern world. In this context, the "RNA world" hypothesis proposes that early in the evolution of life, the ribozyme was responsible for the storage and transfer of genetic information and for the catalysis of biochemical reactions. Accordingly, the hammerhead ribozyme (HHR) and the hairpin ribozyme belong to a family of endonucleolytic RNAs performing self-cleavage that might occur during replication. Furthermore, regarding the widespread occurrence of HHRs in several genomes of modern organisms (from mammals to small parasites and elsewhere), these small ribozymes have been regarded as living fossils of a primitive RNA world. They fold into 3D structures that generally require long-range intramolecular interactions to adopt the catalytically active conformation under specific physicochemical conditions. By studying viroids as plausible remains of ancient RNA, we recently demonstrated that they replicate in non-specific hosts, emphasizing their adaptability to different environments, which enhanced their survival probability over the ages. All these results exemplify ubiquitous features of life. Those are the structural and functional versatility of small RNAs, ribozymes, and viroids, as well as their diversity and adaptability to various extreme conditions. All these traits must have originated in early life to generate novel RNA populations.
Topics: Nucleic Acid Conformation; RNA, Catalytic; RNA, Viral; Viroids
PubMed: 30901893
DOI: 10.3390/v11030283 -
Accounts of Chemical Research Dec 2020Ribozymes and deoxyribozymes are catalytic RNA and DNA, respectively, that catalyze chemical reactions such as self-cleavage or ligation reactions. While some ribozymes...
Ribozymes and deoxyribozymes are catalytic RNA and DNA, respectively, that catalyze chemical reactions such as self-cleavage or ligation reactions. While some ribozymes are found in nature, a larger variety of ribozymes and deoxyribozymes have been discovered by in vitro selection from random sequences. These catalytic nucleic acids, especially ribozymes, are of fundamental interest because they are crucial for the RNA world hypothesis, which suggests that RNA played a central role in both the propagation of genetic information and catalyzing metabolic reactions in primordial life prior to the emergence of proteins and DNA. On the practical side, catalytic nucleic acids have been extensively engineered for various applications, such as biosensors and genetic devices for synthetic biology. Therefore, it is important to gain a deeper understanding of the sequence-function relationships of ribozymes and deoxyribozymes.Mutational analysis, or measurements of activities of catalytic nucleic acid mutants, is one of the most fundamental approaches for that purpose. Mutations that abolish, reduce, retain, or even increase activity provide useful information about nucleic acid catalysts for engineering and other purposes. However, methods for mutational analysis of ribozymes and deoxyribozymes have not evolved much for decades, requiring tedious and low-throughput assays (e.g., gel electrophoresis) of individually prepared mutants. This has prevented researchers from performing quantitative mutational analysis of ribozymes and deoxyribozymes on a large scale.To address this limitation, we developed a massively parallel ribozyme and deoxyribozyme assay strategy that allows >10 assays using high-throughput sequencing (HTS). We used HTS to literally count the number of cleaved (or ligated) and uncleaved (or unligated) ribozyme (or deoxyribozyme) sequences and calculated the activities of each mutant in a reaction mixture. This simple yet powerful strategy was applied to analyze the mutational effects of various natural and synthetic ribozymes and deoxyribozymes at scales impossible for conventional mutational analysis. These large-scale sequence-function data sets were used to better understand the functional consequences of mutations and to engineer ribozymes for practical applications. Furthermore, these newly available data are motivating researchers to employ more rigorous computational methods to extract additional insights such as structural information and nonlinear effects of multiple mutations. The new HTS-based assay strategy is distinct from and complementary to a related strategy that uses HTS to analyze ribozyme and deoxyribozyme populations subjected to in vitro selection. Postselection sequencing can cover a larger sequence space, although it does not directly quantify the activities of ribozyme and deoxyribozyme mutants. With further advances in DNA sequencing technologies and computational methods, there should be more opportunities to harness the power of HTS to deepen our understanding of catalytic nucleic acids and enhance our ability to engineer them for even more applications.
Topics: Aptamers, Nucleotide; DNA, Catalytic; High-Throughput Nucleotide Sequencing; High-Throughput Screening Assays; Kinetics; Mutation; Nucleic Acid Conformation; RNA, Catalytic
PubMed: 33164502
DOI: 10.1021/acs.accounts.0c00546 -
PloS One 2015In support of the RNA world hypothesis, previous studies identified trimetaphosphate (Tmp) as a plausible energy source for RNA world organisms. In one of these studies,...
In support of the RNA world hypothesis, previous studies identified trimetaphosphate (Tmp) as a plausible energy source for RNA world organisms. In one of these studies, catalytic RNAs (ribozymes) that catalyze the triphosphorylation of RNA 5'-hydroxyl groups using Tmp were obtained by in vitro selection. One ribozyme (TPR1) was analyzed in more detail. TPR1 catalyzes the triphosphorylation reaction to a rate of 0.013 min-1 under selection conditions (50 mM Tmp, 100 mM MgCl2, 22°C). To identify a triphosphorylation ribozyme that catalyzes faster triphosphorylation, and possibly learn about its secondary structure TPR1 was subjected to a doped selection. The resulting ribozyme, TPR1e, contains seven mutations relative to TPR1, displays a previously unidentified duplex that constrains the ribozyme's structure, and reacts at a 24-fold faster rate than the parent ribozyme. Under optimal conditions (150 mM Tmp, 650 mM MgCl2, 40°C), the triphosphorylation rate of TRP1e reaches 6.8 min-1.
Topics: Base Sequence; Evolution, Molecular; Kinetics; Models, Molecular; Molecular Sequence Data; Mutagenesis, Site-Directed; Nucleic Acid Conformation; Phosphorylation; RNA, Catalytic
PubMed: 26545116
DOI: 10.1371/journal.pone.0142559 -
Molecules (Basel, Switzerland) Jan 2017Small nucleolytic ribozymes are a family of naturally occurring RNA motifs that catalyse a self-transesterification reaction in a highly sequence-specific manner. The... (Review)
Review
Small nucleolytic ribozymes are a family of naturally occurring RNA motifs that catalyse a self-transesterification reaction in a highly sequence-specific manner. The hammerhead ribozyme was the first reported and the most extensively studied member of this family. However, and despite intense biochemical and structural research for three decades since its discovery, the history of this model ribozyme seems to be far from finished. The hammerhead ribozyme has been regarded as a biological oddity typical of small circular RNA pathogens of plants. More recently, numerous and new variations of this ribozyme have been found to inhabit the genomes of organisms from all life kingdoms, although their precise biological functions are not yet well understood.
Topics: Animals; Base Pairing; Base Sequence; Biocatalysis; Catalytic Domain; History, 20th Century; History, 21st Century; Hydrolysis; Models, Molecular; Nucleic Acid Conformation; Plants; RNA; RNA, Catalytic; RNA, Circular; Schistosoma mansoni
PubMed: 28054987
DOI: 10.3390/molecules22010078