scholarly journals Structural-profiling of low molecular weight RNAs by nanopore trapping/translocation using Mycobacterium smegmatis porin A

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Yuqin Wang ◽  
Xiaoyu Guan ◽  
Shanyu Zhang ◽  
Yao Liu ◽  
Sha Wang ◽  
...  
Keyword(s):  
Single Molecule ◽  
Mycobacterium Smegmatis ◽  
Tertiary Structure ◽  
Learning Algorithm ◽  
Biological Activities ◽  
Rna Structures ◽  
Native Form ◽  
Tertiary Structures ◽  
Structure Report ◽  
Interfering Rna

AbstractFolding of RNA can produce elaborate tertiary structures, corresponding to their diverse roles in the regulation of biological activities. Direct observation of RNA structures at high resolution in their native form however remains a challenge. The large vestibule and the narrow constriction of a Mycobacterium smegmatis porin A (MspA) suggests a sensing mode called nanopore trapping/translocation, which clearly distinguishes between microRNA, small interfering RNA (siRNA), transfer RNA (tRNA) and 5 S ribosomal RNA (rRNA). To further profit from the acquired event characteristics, a custom machine learning algorithm is developed. Events from measurements with a mixture of RNA analytes can be automatically classified, reporting a general accuracy of ~93.4%. tRNAs, which possess a unique tertiary structure, report a highly distinguishable sensing feature, different from all other RNA types tested in this study. With this strategy, tRNAs from different sources are measured and a high structural conservation across different species is observed in single molecule.

scholarly journals A Phylogenetically Conserved Hairpin-Type 3′ Untranslated Region Pseudoknot Functions in Coronavirus RNA Replication

1999 ◽  
Vol 73 (10) ◽  
pp. 8349-8355 ◽  
Author(s):  
Gwyn D. Williams ◽  
Ruey-Yi Chang ◽  
David A. Brian
Keyword(s):  
Untranslated Region ◽  
Tertiary Structure ◽  
Mutational Analysis ◽  
Rna Replication ◽  
Regulatory Control ◽  
Control Element ◽  
Tertiary Structures ◽  
Pseudoknotted Structure ◽  
Type 3 ◽  
Interfering Rna

ABSTRACT Secondary and tertiary structures in the 3′ untranslated region (UTR) of plus-strand RNA viruses have been postulated to function as control elements in RNA replication, transcription, and translation. Here we describe a 54-nucleotide (nt) hairpin-type pseudoknot within the 288-nt 3′ UTR of the bovine coronavirus genome and show by mutational analysis of both stems that the pseudoknotted structure is required for the replication of a defective interfering RNA genome. The pseudoknot is phylogenetically conserved among coronaviruses both in location and in shape but only partially in nucleotide sequence, and evolutionary covariation of bases to maintain G · U pairings indicates that it functions in the plus strand. RNase probing of synthetic transcripts provided additional evidence of its tertiary structure and also identified the possible existence of two conformational states. These results indicate that the 3′ UTR pseudoknot is involved in coronavirus RNA replication and lead us to postulate that it functions as a regulatory control element.

scholarly journals Base-intercalated and base-wedged stacking elements in 3D-structure of RNA and RNA–protein complexes

2020 ◽  
Vol 48 (15) ◽  
pp. 8675-8685
Author(s):  
Eugene Baulin ◽  
Valeriy Metelev ◽  
Alexey Bogdanov
Keyword(s):  
Tertiary Structure ◽  
Protein Complexes ◽  
3D Structure ◽  
Data Bank ◽  
Rna Structures ◽  
Tertiary Structures ◽  
Long Range Interactions ◽  
Heterocyclic Bases ◽  
Rna Protein Complexes

Abstract Along with nucleobase pairing, base-base stacking interactions are one of the two main types of strong non-covalent interactions that define the unique secondary and tertiary structure of RNA. In this paper we studied two subfamilies of nucleobase-inserted stacking structures: (i) with any base intercalated between neighboring nucleotide residues (base-intercalated element, BIE, i + 1); (ii) with any base wedged into a hydrophobic cavity formed by heterocyclic bases of two nucleotides which are one nucleotide apart in sequence (base-wedged element, BWE, i + 2). We have exploited the growing database of natively folded RNA structures in Protein Data Bank to analyze the distribution and structural role of these motifs in RNA. We found that these structural elements initially found in yeast tRNAPhe are quite widespread among the tertiary structures of various RNAs. These motifs perform diverse roles in RNA 3D structure formation and its maintenance. They contribute to the folding of RNA bulges and loops and participate in long-range interactions of single-stranded stretches within RNA macromolecules. Furthermore, both base-intercalated and base-wedged motifs participate directly or indirectly in the formation of RNA functional centers, which interact with various ligands, antibiotics and proteins.

Torus Circumference and Thickness Parameters From a Linear DNA Size Series Suggest How Toruses Self-Assemble

1985 ◽  
Vol 43 ◽  
pp. 522-523
Author(s):  
George C. Ruben ◽  
Kenneth A. Marx
Keyword(s):  
Tertiary Structure ◽  
Dna Complexes ◽  
Tertiary Structures ◽  
Self Assembling ◽  
Double Stranded Dna ◽  
Calf Thymus ◽  
Dna Organization ◽  
Condensed Dna ◽  
Dna Size

Certain double stranded DNA bacteriophage and viruses are thought to have their DNA organized into large torus shaped structures. Morphologically, these poorly understood biological DNA tertiary structures resemble spermidine-condensed DNA complexes formed in vitro in the total absence of other macromolecules normally synthesized by the pathogens for the purpose of their own DNA packaging. Therefore, we have studied the tertiary structure of these self-assembling torus shaped spermidine- DNA complexes in a series of reports. Using freeze-etch, low Pt-C metal (10-15Å) replicas, we have visualized the microscopic DNA organization of both calf Thymus( CT) and linear 0X-174 RFII DNA toruses. In these structures DNA is circumferentially wound, continuously, around the torus into a semi-crystalline, hexagonal packed array of parallel DNA helix sections.

scholarly journals Calcium-induced tertiary structure modifications of endo-β-1,3-glucanase from Pyrococcus furiosus in 7.9 M guanidinium chloride

2005 ◽  
Vol 386 (3) ◽  
pp. 515-524 ◽  
Author(s):  
Roberta CHIARALUCE ◽  
Giulio GIANESE ◽  
Sebastiana ANGELACCIO ◽  
Rita FLORIO ◽  
Johan F. T. van LIESHOUT ◽  
...  
Keyword(s):  
Tertiary Structure ◽  
Pyrococcus Furiosus ◽  
Glycoside Hydrolases ◽  
Guanidinium Chloride ◽  
Absorption Region ◽  
Native Form ◽  
Carboxylate Groups ◽  
The Family ◽  
Notable Increase ◽  
Putative Binding Site

The family 16 endo-β-1,3 glucanase from the extremophilic archaeon Pyrococcus furiosus is a laminarinase, which in 7.9 M GdmCl (guanidinium chloride) maintains a significant amount of tertiary structure without any change of secondary structure. The addition of calcium to the enzyme in 7.9 M GdmCl causes significant changes to the near-UV CD and fluorescence spectra, suggesting a notable increase in the tertiary structure which leads to a state comparable, but not identical, to the native state. The capability to interact with calcium in 7.9 M GdmCl with a consistent recovery of native tertiary structure is a unique property of this extremely stable endo-β-1,3 glucanase. The effect of calcium on the thermodynamic parameters relative to the GdmCl-induced equilibrium unfolding has been analysed by CD and fluorescence spectroscopy. The interaction of calcium with the native form of the enzyme is studied by Fourier-transform infrared spectroscopy in the absorption region of carboxylate groups and by titration in the presence of a chromophoric chelator. A homology-based model of the enzyme is generated and used to predict the putative binding site(s) for calcium and the structural interactions potentially responsible for the unusual stability of this protein, in comparison with other family 16 glycoside hydrolases.

scholarly journals Rapid and multiplex preparation of engineered Mycobacterium smegmatis porin A (MspA) nanopores for single molecule sensing and sequencing

2021 ◽  
Author(s):  
Shuanghong Yan ◽  
Liying Wang ◽  
Xiaoyu Du ◽  
Shanyu Zhang ◽  
Sha Wang ◽  
...  
Keyword(s):  
Single Molecule ◽  
Mycobacterium Smegmatis ◽  
Recent Developments

Acknowledging its unique conical lumen structure, Mycobacterium smegmatis porin A (MspA) was the first type of nanopore that has successfully sequenced DNA. Recent developments of nanopore single molecule chemistry have...

scholarly journals Preparation of Mycobacterium smegmatis porin A (MspA) nanopores for single molecule sensing of nucleic acids

2021 ◽  
Vol 7 (5) ◽  
pp. 355-364
Author(s):  
Wang Yuqin ◽  
◽  
Fan Pingping ◽  
Zhang Shanyu ◽  
Yan Shuanghong ◽  
...  
Keyword(s):  
Nucleic Acids ◽  
Single Molecule ◽  
Mycobacterium Smegmatis

scholarly journals An NMR-based approach reveals the core structure of the functional domain of SINEUP lncRNAs

2020 ◽  
Vol 48 (16) ◽  
pp. 9346-9360
Author(s):  
Takako Ohyama ◽  
Hazuki Takahashi ◽  
Harshita Sharma ◽  
Toshio Yamazaki ◽  
Stefano Gustincich ◽  
...  
Keyword(s):  
Nuclear Magnetic Resonance ◽  
Computational Prediction ◽  
Functional Domain ◽  
Rna Structures ◽  
Tertiary Structures ◽  
Functional Roles ◽  
The Core ◽  
Non Coding Rnas ◽  
Dynamic Domain

Abstract Long non-coding RNAs (lncRNAs) are attracting widespread attention for their emerging regulatory, transcriptional, epigenetic, structural and various other functions. Comprehensive transcriptome analysis has revealed that retrotransposon elements (REs) are transcribed and enriched in lncRNA sequences. However, the functions of lncRNAs and the molecular roles of the embedded REs are largely unknown. The secondary and tertiary structures of lncRNAs and their embedded REs are likely to have essential functional roles, but experimental determination and reliable computational prediction of large RNA structures have been extremely challenging. We report here the nuclear magnetic resonance (NMR)-based secondary structure determination of the 167-nt inverted short interspersed nuclear element (SINE) B2, which is embedded in antisense Uchl1 lncRNA and upregulates the translation of sense Uchl1 mRNAs. By using NMR ‘fingerprints’ as a sensitive probe in the domain survey, we successfully divided the full-length inverted SINE B2 into minimal units made of two discrete structured domains and one dynamic domain without altering their original structures after careful boundary adjustments. This approach allowed us to identify a structured domain in nucleotides 31–119 of the inverted SINE B2. This approach will be applicable to determining the structures of other regulatory lncRNAs.

scholarly journals Investigating Serum and Tissue Expression Identified a Cytokine/Chemokine Signature as a Highly Effective Melanoma Marker

Cancers ◽  
2020 ◽  
Vol 12 (12) ◽  
pp. 3680
Author(s):  
Marco Cesati ◽  
Francesca Scatozza ◽  
Daniela D’Arcangelo ◽  
Gian Carlo Antonini-Cappellini ◽  
Stefania Rossi ◽  
...  
Keyword(s):  
Single Molecule ◽  
Learning Algorithm ◽  
Profile Analysis ◽  
Pearson Correlation ◽  
Tissue Expression ◽  
Support Vector ◽  
Expression Data ◽  
Serum Samples ◽  
Tissue Samples ◽  
Mortality And Morbidity

The identification of reliable and quantitative melanoma biomarkers may help an early diagnosis and may directly affect melanoma mortality and morbidity. The aim of the present study was to identify effective biomarkers by investigating the expression of 27 cytokines/chemokines in melanoma compared to healthy controls, both in serum and in tissue samples. Serum samples were from 232 patients recruited at the IDI-IRCCS hospital. Expression was quantified by xMAP technology, on 27 cytokines/chemokines, compared to the control sera. RNA expression data of the same 27 molecules were obtained from 511 melanoma- and healthy-tissue samples, from the GENT2 database. Statistical analysis involved a 3-step approach: analysis of the single-molecules by Mann–Whitney analysis; analysis of paired-molecules by Pearson correlation; and profile analysis by the machine learning algorithm Support Vector Machine (SVM). Single-molecule analysis of serum expression identified IL-1b, IL-6, IP-10, PDGF-BB, and RANTES differently expressed in melanoma (p < 0.05). Expression of IL-8, GM-CSF, MCP-1, and TNF-α was found to be significantly correlated with Breslow thickness. Eotaxin and MCP-1 were found differentially expressed in male vs. female patients. Tissue expression analysis identified very effective marker/predictor genes, namely, IL-1Ra, IL-7, MIP-1a, and MIP-1b, with individual AUC values of 0.88, 0.86, 0.93, 0.87, respectively. SVM analysis of the tissue expression data identified the combination of these four molecules as the most effective signature to discriminate melanoma patients (AUC = 0.98). Validation, using the GEPIA2 database on an additional 1019 independent samples, fully confirmed these observations. The present study demonstrates, for the first time, that the IL-1Ra, IL-7, MIP-1a, and MIP-1b gene signature discriminates melanoma from control tissues with extremely high efficacy. We therefore propose this 4-molecule combination as an effective melanoma marker.

BIOLOGICAL ACTIONS OF HUMAN SOMATOTROPHIN AND ITS DERIVATIVES ON MOUSE MAMMARY GLAND AND TELEOST URINARY BLADDER

1977 ◽  
Vol 73 (2) ◽  
pp. 377-383 ◽  
Author(s):  
B. A. DONEEN ◽  
H. A. BERN ◽  
CHOH HAO LI
Keyword(s):  
Mammary Gland ◽  
Urinary Bladder ◽  
Tertiary Structure ◽  
Biological Activities ◽  
Mouse Mammary Gland ◽  
Performic Acid ◽  
Acid Oxidation ◽  
Peptide Fragments ◽  
Ovine Prolactin

SUMMARY These studies are concerned with the structural and functional evolution of the ancestrally related pituitary prolactins and somatotrophins. Prolactin-like biological activities of human somatotrophin (hGH) and its peptide fragments were bioassayed in vitro on the mouse mammary gland and the teleost urinary bladder. Plasmin modified-hGH was as active as hGH in both bioassays. The NH2-terminal 134-residue fragment possessed about 10% of the lactogenic and urinary bladder potency of hGH, whereas the CO2H-terminal 51-residue fragment was inactive at the concentrations observed. These results suggest that the same regions of primary structure are responsible for the prolactin-like actions of hGH on the target organs of lower and higher vertebrates. Alteration of the tertiary structure of hGH, human chorionic somatomammotrophin, and ovine prolactin by performic acid oxidation destroys the mammary gland activities of these hormones.

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