Folding mechanisms of group I ribozymes: role of stability and contact order

2002 ◽  
Vol 30 (6) ◽  
pp. 1166-1169 ◽  
Author(s):  
S. A. Woodson
Keyword(s):  
Catalytic Activity ◽  
Rna Folding ◽  
Three Dimensional ◽  
Model System ◽  
Folding Pathway ◽  
Primary Sequence ◽  
Rna Molecules ◽  
Contact Order ◽  
Group I

The mechanism by which RNA molecules assemble into unique three-dimensional conformations is important for understanding their function, regulation and interactions with substrates. The Tetrahymena group I ribozyme is an excellent model system for understanding RNA folding mechanisms, because the catalytic activity of the native RNA is easily measured. Folding of the Tetrahymena ribozyme is dominated by intermediates in which the stable P4-P6 domain is correctly formed, but the P3-P9 domain is partially misfolded. The propensity of the RNA to misfold depends on the relative stability of native and non-native interactions. Circular permutation of the Tetrahymena ribozyme shows that the distance in the primary sequence between native interactions also influences the folding pathway.

Group I introns and RNA folding

2002 ◽  
Vol 30 (6) ◽  
pp. 1149-1152 ◽  
Author(s):  
E. Westhof
Keyword(s):  
Catalytic Activity ◽  
Self Assembly ◽  
Rna Folding ◽  
Catalytic Rna ◽  
Group I Introns ◽  
Rna Motifs ◽  
X Ray ◽  
Rna Molecules ◽  
Group I ◽  
Self Splicing

Before the discovery of catalytic RNA, tRNA molecules were the most studied RNA molecules for understanding RNA folding. Afterwards, group I introns, because of their stability and the fact that structural folding could be monitored by following their catalytic activity, became the molecule of choice for studying RNA architecture and folding. A major advantage of group I introns for studying the catalytic activity of RNA molecules is that catalytic activity is triggered by the addition of external guanosine co-factors. The self-splicing activity can therefore be precisely controlled. Using group I introns, several RNA motifs central to RNA-RNA self-assembly and folding were discovered. The analysis of the recent X-ray structures of the rRNA subunits indicates that several motifs present in the ribosome occur also in various group I introns.

scholarly journals Supramolecular organization as a factor of ribonuclease cytotoxicity

Acta Naturae ◽  
2020 ◽  
Vol 12 (3) ◽  
pp. 24-33
Author(s):  
Elena V. Dudkina ◽  
Vera V. Ulyanova ◽  
Olga N. Ilinskaya
Keyword(s):  
Catalytic Activity ◽  
Cancer Cells ◽  
Therapeutic Potential ◽  
Biological Effects ◽  
Supramolecular Organization ◽  
Cytotoxic Effects ◽  
Selective Cytotoxicity ◽  
Rna Molecules ◽  
Cell Components

One of the approaches used to eliminate tumor cells is directed destruction/modification of their RNA molecules. In this regard, ribonucleases (RNases) possess a therapeutic potential that remains largely unexplored. It is believed that the biological effects of secreted RNases, namely their antitumor and antiviral properties, derive from their catalytic activity. However, a number of recent studies have challenged the notion that the activity of RNases in the manifestation of selective cytotoxicity towards cancer cells is exclusively an enzymatic one. In this review, we have analyzed available data on the cytotoxic effects of secreted RNases, which are not associated with their catalytic activity, and we have provided evidence that the most important factor in the selective apoptosis-inducing action of RNases is the structural organization of these enzymes, which determines how they interact with cell components. The new idea on the preponderant role of non-catalytic interactions between RNases and cancer cells in the manifestation of selective cytotoxicity will contribute to the development of antitumor RNase-based drugs.

scholarly journals Three-Dimensional Structures of the A, B, and CCapsids of Rhesus Monkey Rhadinovirus: Insights into GammaherpesvirusCapsid Assembly, Maturation, and DNAPackaging

2003 ◽  
Vol 77 (24) ◽  
pp. 13182-13193 ◽  
Author(s):  
Xue-Kui Yu ◽  
Christine M. O'Connor ◽  
Ivo Atanasov ◽  
Blossom Damania ◽  
Dean H. Kedes ◽  
...  
Keyword(s):  
Rhesus Monkey ◽  
Three Dimensional ◽  
Model System ◽  
Electron Cryomicroscopy ◽  
Interacting Protein ◽  
Structural Differences ◽  
Dimensional Reconstruction ◽  
Capsid Maturation ◽  
First Time

ABSTRACT Rhesus monkey rhadinovirus (RRV) exhibits high levels of sequence homology to human gammaherpesviruses, such as Kaposi's sarcoma-associated herpesvirus, and grows to high titers in cell cultures, making it a good model system for studying gammaherpesvirus capsid structure and assembly. We have purified RRV A, B, and C capsids, thus for the first time allowing direct structure comparisons by electron cryomicroscopy and three-dimensional reconstruction. The results show that the shells of these capsids are identical and are each composed of 12 pentons, 150 hexons, and 320 triplexes. Structural differences were apparent inside the shells and through the penton channels. The A capsid is empty, and its penton channels are open. The B capsid contains a scaffolding core, and its penton channels are closed. The C capsid contains a DNA genome, which is closely packaged into regularly spaced density shells (25Å apart), and its penton channels are open. The different statuses of the penton channels suggest a functional role of the channels during capsid maturation, and the overall structural similarities of RRV capsids to alphaherpesvirus capsids suggest a common assembly and maturation pathway. The RRV A capsid reconstruction at a 15-Å resolution, the best achieved for gammaherpesvirus particles, reveals overall structural similarities to alpha- and betaherpesvirus capsids. However, the outer regions of the capsid, including densities attributed to the Ta triplex and the small capsomer-interacting protein (SCIP or ORF65), exhibit prominent differences from their structural counterparts in alphaherpesviruses. This structural disparity suggests that SCIP and the triplex, together with tegument and envelope proteins, confer structural and potentially functional specificities to alpha-, beta-, and gammaherpesviruses.

scholarly journals Distributed Biotin-Streptavidin Transcription Roadblocks for Mapping Cotranscriptional RNA Folding

2017 ◽  
Author(s):  
Eric J. Strobel ◽  
Kyle E. Watters ◽  
Julius B. Lucks
Keyword(s):  
Experimental Study ◽  
Rna Structure ◽  
Rna Folding ◽  
Folding Pathway ◽  
Decision Making Process ◽  
Important Process ◽  
Rna Structures ◽  
Rna Molecules ◽  
Fundamental Properties ◽  
Nucleotide Resolution

AbstractRNA molecules fold cotranscriptionally as they emerge from RNA polymerase. Cotranscriptional folding is an important process for proper RNA structure formation as the order of folding can determine an RNA molecule’s structure, and thus its functional properties. Despite its fundamental importance, the experimental study of RNA cotranscriptional folding has been limited by the lack of easily approachable methods that can interrogate nascent RNA structures at nucleotide resolution during transcription. We previously developed cotranscriptional selective 2’-hydroxyl acylation analyzed by primer extension sequencing (SHAPE-seq) to simultaneously probe all of the intermediate structures an RNA molecule transitions through during transcription elongation. Here, we improve the broad applicability of cotranscriptional SHAPE-Seq by developing a sequence-independent streptavidin roadblocking strategy to simplify the preparation of roadblocking transcription templates. We determine the fundamental properties of streptavidin roadblocks and show that randomly distributed streptavidin roadblocks can be used in cotranscriptional SHAPE-Seq experiments to measure the Bacillus cereus crcB fluoride riboswitch folding pathway. Comparison of EcoRIE111Q and streptavidin roadblocks in cotranscriptional SHAPE-Seq data shows that both strategies identify the same RNA structural transitions related to the riboswitch decision-making process. Finally, we propose guidelines to leverage the complementary strengths of each transcription roadblock for use in studying cotranscriptional folding.

The role of disulfide bonds in the conformational stability and catalytic activity of phytase

2004 ◽  
Vol 82 (2) ◽  
pp. 329-334 ◽  
Author(s):  
Xiao-Yun Wang ◽  
Fan-Guo Meng ◽  
Hai-Meng Zhou
Keyword(s):  
Catalytic Activity ◽  
Disulfide Bonds ◽  
Conformational Stability ◽  
Three Dimensional ◽  
Fluorescence Emission ◽  
Dimensional Structure ◽  
Enzyme Activity Assay ◽  
First Order ◽  
Far Ultraviolet

Previous studies have predicted five disulfide bonds in Aspergillus niger phytase (phy A). To investigate the role of disulfide bonds, intrinsic fluorescence spectra, far-ultraviolet circular dichroism (CD) spectra, and an enzyme activity assay were used to compare the differences of catalytic activity and conformational stability of phytase during denaturation in urea in the presence and absence of dithiothreitol (DTT). In the presence of 2 mM DTT, the inactivation and unfolding were greatly enhanced at the same concentration of denaturant. The fluorescence emission maximum red shift and decreases of ellipticity at 222 nm were in accord with the changes of catalytic activity. The kinetics of the unfolding courses were a biphasic process consisting of two first-order reactions in the absence of DTT and a monophasic process of a first-order reaction in the presence of DTT. The results suggested that the loss of enzymatic activity was most likely because of a conformational change, and that disulfide bonds played an important role in three-dimensional structure and catalytic activity.Key words: phytase, urea denaturation, inactivation, disulfide bond.

Phenotypic Traits and Regulatory Role of RNA Folding in Molecular Selection

1991 ◽  
Vol 46 (7-8) ◽  
pp. 656-662
Author(s):  
Ariel Fernández
Keyword(s):  
Rna Folding ◽  
De Novo ◽  
Phenotypic Expression ◽  
Regulatory Role ◽  
Phenotypic Traits ◽  
Intermediate Structure ◽  
Rna Sequences ◽  
Rna Molecules ◽  
Molecular Selection

Abstract We concentrate on instances in which the phenotypic expression of information encoded in an RNA primary sequence might be revealed by the folding of the RNA itself. We have discov­ered that this situation finds concrete realization in the design of RNA molecules capable of maximizing the rate of autocatalytic synthesis when incubated with viral Qβ-replicase. This requires that we introduce the notion of phenotypic traits at the molecular level. Thus, the problem of finding RNA sequences whose phenotype favorably influences propagation amounts to finding RNA sequences which fold so as to optimize enzymatic performance and are in addition endowed with the proper recognition sites. The proof that these two problems are indeed equivalent has two steps: First we predict the metastable folded structures formed as a template RNA chain grows by sequential incorporation of nucleotides. The transient folded states appear to be involved in the regulation of the enzyme activity and they occur in a manner which is “oblivious” of thermodynamic time scales. Secondly, we compute the time-dependent activation energy for relaxation of each intermediate structure. This is done to establish constraints necessary for optimization of the regulatory role of RNA folding. The search for prospective template sequences is subject to such constraints. Our results aim at elucidating an optimization process realized by molecular selection in de novo (template-free) RNA synthesis by Qβ-replicase. We argue that the phenotype which mediates selection is given by metastable folding which emerges together with the printing of the genotype, that is, within the time span of a replication turnover.

Homology Requirements for Double-Strand Break-Mediated Recombination in a Phage λ-td Intron Model System

Genetics ◽  
1996 ◽  
Vol 143 (3) ◽  
pp. 1057-1068 ◽  
Author(s):  
Monica M Parker ◽  
Deborah A Court ◽  
Karen Preiter ◽  
Marlene Belfort
Keyword(s):  
Strand Break ◽  
Double Strand Break ◽  
Model System ◽  
Group I Introns ◽  
Wild Type ◽  
Intron Insertion ◽  
Homologous Recombination Event ◽  
Group I ◽  
Intron Homing

Abstract Many group I introns encode endonucleases that promote intron homing by initiating a double-strand break-mediated homologous recombination event. A td intron-phage λ model system was developed to analyze exon homology effects on intron homing and determine the role of the λ 5′–3′ exonuclease complex (Redαβ) in the repair event. Efficient intron homing depended on exon lengths in the 35- to 50-bp range, although homing levels remained significantly elevated above nonbreak-mediated recombination with as little as 10 bp of flanking homology. Although precise intron insertion was demonstrated with extremely limiting exon homology, the complete absence of one exon produced illegitimate events on the side of heterology. Interestingly, intron inheritance was unaffected by the presence of extensive heterology at the double-strand break in wild-type λ, provided that sufficient homology between donor and recipient was present distal to the heterologous sequences. However, these events involving heterologous ends were absolutely dependent on an intact Red exonuclease system. Together these results indicate that heterologous sequences can participate in double-strand break-mediated repair and imply that intron transposition to heteroallelic sites might occur at break sites within regions of limited or no homology.

scholarly journals Understanding the Role of Three-Dimensional Topology in Determining the Folding Intermediates of Group I Introns

2013 ◽  
Vol 104 (6) ◽  
pp. 1326-1337 ◽  
Author(s):  
Chunxia Chen ◽  
Somdeb Mitra ◽  
Magdalena Jonikas ◽  
Joshua Martin ◽  
Michael Brenowitz ◽  
...  
Keyword(s):  
Three Dimensional ◽  
Group I Introns ◽  
Folding Intermediates ◽  
Group I
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