scholarly journals Dissecting and predicting different types of binding sites in nucleic acids based on structural information

2021 ◽  
Author(s):  
Zheng Jiang ◽  
Si-Rui Xiao ◽  
Rong Liu
Keyword(s):  
Nucleic Acid ◽  
Nucleic Acids ◽  
Binding Sites ◽  
Structural Information ◽  
Machine Learning Algorithms ◽  
Nucleic Acid Binding ◽  
Integrative Framework ◽  
Dna And Rna ◽  
Feature Spaces ◽  
Critical Binding

Abstract The biological functions of DNA and RNA generally depend on their interactions with other molecules, such as small ligands, proteins and nucleic acids. However, our knowledge of the nucleic acid binding sites for different interaction partners is very limited, and identification of these critical binding regions is not a trivial work. Herein, we performed a comprehensive comparison between binding and nonbinding sites and among different categories of binding sites in these two nucleic acid classes. From the structural perspective, RNA may interact with ligands through forming binding pockets and contact proteins and nucleic acids using protruding surfaces, while DNA may adopt regions closer to the middle of the chain to make contacts with other molecules. Based on structural information, we established a feature-based ensemble learning classifier to identify the binding sites by fully using the interplay among different machine learning algorithms, feature spaces and sample spaces. Meanwhile, we designed a template-based classifier by exploiting structural conservation. The complementarity between the two classifiers motivated us to build an integrative framework for improving prediction performance. Moreover, we utilized a post-processing procedure based on the random walk algorithm to further correct the integrative predictions. Our unified prediction framework yielded promising results for different binding sites and outperformed existing methods.

scholarly journals Novel cyanine dye as competitive ligand for probing the drug–nucleic acid interactions

2020 ◽  
Keyword(s):  
Nucleic Acid ◽  
Nucleic Acids ◽  
Binding Sites ◽  
Lanthanide Complexes ◽  
Rna Binding ◽  
Cyanine Dye ◽  
Dna And Rna ◽  
Dna And Rna Binding ◽  
Drug Displacement ◽  
Potential Antitumor

Background: During the past decades, increasing attention has been given to elucidating the molecular details of interactions between the pharmacological agents and nucleic acids since the drug–DNA complexation may lead to impairment of DNA replication, strand breaking and mutations. A variety of techniques have been developed to characterize the drug-nucleic acid binding, among which the fluorescence dye displacement assay is one of the most informative approaches. Recently, it was demonstrated that cyanine dyes can be successfully employed for the high throughput screening of the interactions between nucleic acids and drugs. To the best of our knowledge, so far, the potential application of cyanine dyes for the drug-displacement studies remains insufficiently evaluated. Objectives: The aim of the present study was to investigate the ability of a novel cyanine dye to serve as a competitor for the potential antitumor compounds, lanthanide complexes bearing europium (III) tris-β-diketonate (EC) for the DNA and RNA binding sites. Materials and methods: Calf thymus DNA, yeast RNA, trimethine cyanine dye and lanthanide complexes bearing europium (III) tris-β-diketonate were used for sample preparation. The fluorescence data were acquired using Perkin-Elmer LS-55 spectrofluorimeter. Results: Using the fluorescence spectroscopy technique we conducted the displacement reaction trimethine cyanine dye/europium coordination complexes in the presence of double stranded DNA and single-stranded RNA. An increase of the EC concentration in the systems AK3-5/DNA or AK3-5/RNA was followed by a gradual reduction in the AK3-5 fluorescence intensity, indicating that europium (III) tris-β-diketonate compounds can serve as competitors for the trimethine cyanine dye on the nucleic acids. Both the drug chemical structure and the type of nucleic acid proved to control the extent of EC-induced decrease of AK3-5 fluorescence in the presence of the DNA or RNA. Conclusion: By recruiting the potential antitumor agents europium chelate complexes as the competitive ligands for the cyanine dye for the DNA and RNA binding sites, we found that a novel trimethine compound can be effectively used in the fluorescence drug displacement assays.

Snapshot blotting: The transfer of nucleic acids and nucleoprotein complexes from electrophoresis gels to EM grids

1992 ◽  
Vol 50 (1) ◽  
pp. 762-763
Author(s):  
Stephen D. Jett
Keyword(s):  
Nucleic Acid ◽  
Nucleic Acids ◽  
Nucleic Acid Binding ◽  
Mobility Shift ◽  
Carbon Coated ◽  
Nucleoprotein Complexes ◽  
Mobility Shift Assay ◽  
Protein Nucleic Acid ◽  
Gel Mobility Shift ◽  
Nucleic Acid Interactions

The electrophoresis gel mobility shift assay is a popular method for the study of protein-nucleic acid interactions. The binding of proteins to DNA is characterized by a reduction in the electrophoretic mobility of the nucleic acid. Binding affinity, stoichiometry, and kinetics can be obtained from such assays; however, it is often desirable to image the various species in the gel bands using TEM. Present methods for isolation of nucleoproteins from gel bands are inefficient and often destroy the native structure of the complexes. We have developed a technique, called “snapshot blotting,” by which nucleic acids and nucleoprotein complexes in electrophoresis gels can be electrophoretically transferred directly onto carbon-coated grids for TEM imaging.

An Investigation of the Anomalous Staining of Chromatin by the Acid Dyes, Methyl Blue and Aniline Blue

1962 ◽  
Vol s3-103 (64) ◽  
pp. 519-530
Author(s):  
R. B. McKAY
Keyword(s):  
Nucleic Acid ◽  
Nucleic Acids ◽  
Methyl Blue ◽  
Aniline Blue ◽  
Acid Dyes ◽  
Dna And Rna ◽  
Basic Dyes ◽  
Blue Stain ◽  
High Degree ◽  
Mechanism Of The Reaction

Methyl blue and aniline blue, though acid dyes, stain the chromatin of the spermatogenetic cells of the mouse (especially of the primary spermatocytes) strongly. Extraction of the basiphil nucleic acid constituents from the chromatin causes loss of this property, while destruction of acidophilia in the protein constituents does not. It has been concluded that the dyes interact with the nucleic acids. Further, they appear to react with both DNA and RNA in the chromatin, although they show no affinity for the cytoplasm of the exocrine cells in sections of pancreas, which is rich in RNA. The mechanism of the reaction has not been fully elucidated, although apparently the dyes do not behave as basic dyes towards the nucleic acids, and the interaction is non-ionic. Methyl blue and aniline blue stain strongly other ‘acidic’ substrates, such as cellulose and nitrocellulose, and attempts have been made to relate the staining of nucleic acids to the staining of these substrates, particularly cellulose; for the staining properties of this substrate have been intensively investigated elsewhere. No satisfactory correlation, however, has been obtained, for nitrocellulose has been found to be less strongly stained at pH 3.0 than at pH 7.1, while the reverse is true for cellulose. Further, only one of 3 direct cotton dyes used appears to have any affinity for the chromatin of the spermatogenetic cells. Direct cotton dyes have large flat molecules with a high degree of conjugation. It is suggested that these characteristics are essential for interaction with nucleic acids, and also that the molecule must be reasonably compact. Finally, it has been shown that methyl blue, aniline blue, and 3 direct cotton dyes of the azo type have no ability to stain the glycogen in liver cells, yet glycogen is very closely related to cellulose.

Binding Of Immune Serine Proteases To Nucleic Acids Enhances Their Nuclear Localization and Promotes Their Cleavage Of Nucleic Acid-Binding Protein Substrates

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 3471-3471
Author(s):  
Jennifer Whangbo ◽  
Marshall Thomas ◽  
Geoffrey McCrossan ◽  
Aaron Deutsch ◽  
Kimberly Martinod ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Nucleic Acids ◽  
Nuclear Localization ◽  
Serine Proteases ◽  
Rna Binding ◽  
Nuclear Accumulation ◽  
Target Cells ◽  
Nucleic Acid Binding ◽  
High Affinity ◽  
Nucleic Acid Binding Proteins

Abstract When released from cytotoxic T lymphocytes and natural killer cells, Granzyme (Gzm) serine proteases induce programmed cell death of pathogen-infected cells and tumor cells. The Gzms rapidly accumulate in the target cell nucleus by an unknown mechanism. Many of the known substrates of GzmA and GzmB, the most abundant killer cell proteases, bind to DNA or RNA. Gzm substrates predicted by unbiased proteomics studies are also highly enriched for nucleic acid binding proteins. Here we show by fluorescence polarization assays that Gzms bind DNA and RNA with nanomolar affinity. We hypothesized that Gzm binding to nucleic acids enhances nuclear accumulation in target cells and facilitates their cleavage of nucleic acid-binding substrates. In fact, RNase treatment of cell lysates reduced cleavage of RNA binding protein (RBP) targets by GzmA and GzmB. Moreover, adding RNA to recombinant RBP substrates greatly enhanced in vitro cleavage by GzmB, but adding RNA to non-nucleic acid binding proteins did not. For example, exogenous RNA enhanced GzmB cleavage of recombinant hnRNP C1 (an RBP) but not LMNB1 (a non-RBP). In addition, GzmB cleaved the RNA-binding HuR protein efficiently only when it was bound to an HuR-binding RNA oligonucleotide, but not in the presence of an equal amount of non-binding RNA. Thus, nucleic acids facilitate Gzm cleavage of nucleic acid binding substrates. To evaluate whether nucleic acid binding influences Gzm trafficking in target cells, we incubated fixed target cells with RNase and then added Gzms. RNA degradation in target cells reduced Gzm cytosolic localization and increased nuclear accumulation. Similarly, pre-incubating Gzms with exogenous competitor DNA reduced Gzm nuclear localization. The Gzms form a monophyletic clade with other immune serine proteases including neutrophil elastase (NE) and cathepsin G (CATG). Upon neutrophil activation, NE translocates to the nucleus to drive the formation of neutrophil extracellular traps (NETs). NE and CATG, but not non-immune serine proteases such as trypsin and pancreatic elastase, also bind DNA with high affinity and localize to the nucleus of permeabilized cells. Consistent with this finding, competitor DNA also blocks the nuclear localization of NE. Moreover NE and CATG localization to NETs depends on DNA binding. Thus the antimicrobial activity of NETs may depend in part upon the affinity of these proteases for DNA. Our findings indicate that high affinity nucleic acid binding is a conserved and functionally important property of serine proteases involved in cell-mediated immunity. Disclosures: Lieberman: Alnylam Pharmaceuticals: Membership on an entity’s Board of Directors or advisory committees.

scholarly journals NABP1, a novel RORγ-regulated gene encoding a single-stranded nucleic-acid-binding protein

2006 ◽  
Vol 397 (1) ◽  
pp. 89-99 ◽  
Author(s):  
Hong Soon Kang ◽  
Ju Youn Beak ◽  
Yong-Sik Kim ◽  
Robert M. Petrovich ◽  
Jennifer B. Collins ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Nucleic Acids ◽  
Binding Protein ◽  
Size Exclusion ◽  
Binding Motif ◽  
Nucleic Acid Binding ◽  
Wild Type ◽  
Gene Encoding ◽  
Nucleic Acid Binding Protein ◽  
Acid Binding Protein

RORγ2 (retinoid-related orphan receptor γ2) plays a critical role in the regulation of thymopoiesis. Microarray analysis was performed in order to uncover differences in gene expression between thymocytes of wild-type and RORγ−/− mice. This analysis identified a novel gene encoding a 22 kDa protein, referred to as NABP1 (nucleic-acid-binding protein 1). This subsequently led to the identification of an additional protein, closely related to NABP1, designated NABP2. Both proteins contain an OB (oligonucleotide/oligosaccharide binding) motif at their N-terminus. This motif is highly conserved between the two proteins. NABP1 is highly expressed in the thymus of wild-type mice and is greatly suppressed in RORγ−/− mice. During thymopoiesis, NABP1 mRNA expression is restricted to CD4+CD8+ thymocytes, an expression pattern similar to that observed for RORγ2. These observations appear to suggest that NABP1 expression is regulated either directly or indirectly by RORγ2. Confocal microscopic analysis showed that the NABP1 protein localizes to the nucleus. Analysis of nuclear proteins by size-exclusion chromatography indicated that NABP1 is part of a high molecular-mass protein complex. Since the OB-fold is frequently involved in the recognition of nucleic acids, the interaction of NABP1 with various nucleic acids was examined. Our results demonstrate that NABP1 binds single-stranded nucleic acids, but not double-stranded DNA, suggesting that it functions as a single-stranded nucleic acid binding protein.

scholarly journals Oligonucleotide-based systems: DNA, microRNAs, DNA/RNA aptamers

2016 ◽  
Vol 60 (1) ◽  
pp. 27-35 ◽  
Author(s):  
Pawan Jolly ◽  
Pedro Estrela ◽  
Michael Ladomery
Keyword(s):  
Nucleic Acid ◽  
Nucleic Acids ◽  
Rna Aptamers ◽  
Locked Nucleic Acid ◽  
Synthetic Analogue ◽  
Dna And Rna ◽  
Naturally Occurring ◽  
Wide Range ◽  
Synthetic Oligonucleotides ◽  
Shed Light

There are an increasing number of applications that have been developed for oligonucleotide-based biosensing systems in genetics and biomedicine. Oligonucleotide-based biosensors are those where the probe to capture the analyte is a strand of deoxyribonucleic acid (DNA), ribonucleic acid (RNA) or a synthetic analogue of naturally occurring nucleic acids. This review will shed light on various types of nucleic acids such as DNA and RNA (particularly microRNAs), their role and their application in biosensing. It will also cover DNA/RNA aptamers, which can be used as bioreceptors for a wide range of targets such as proteins, small molecules, bacteria and even cells. It will also highlight how the invention of synthetic oligonucleotides such as peptide nucleic acid (PNA) or locked nucleic acid (LNA) has pushed the limits of molecular biology and biosensor development to new perspectives. These technologies are very promising albeit still in need of development in order to bridge the gap between the laboratory-based status and the reality of biomedical applications.

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