scholarly journals G-Quadruplex-Based Fluorescent Turn-On Ligands and Aptamers: From Development to Applications

Molecules ◽  
2019 ◽  
Vol 24 (13) ◽  
pp. 2416 ◽  
Author(s):  
Mubarak I. Umar ◽  
Danyang Ji ◽  
Chun-Yin Chan ◽  
Chun Kit Kwok
Keyword(s):  
Interaction Mechanism ◽  
Rna Aptamers ◽  
Rna Sequences ◽  
Turn On ◽  
Dna And Rna ◽  
Cellular Events ◽  
Potential Applications ◽  
Micro Organisms ◽  
Nucleic Acid Structures

Guanine (G)-quadruplexes (G4s) are unique nucleic acid structures that are formed by stacked G-tetrads in G-rich DNA or RNA sequences. G4s have been reported to play significant roles in various cellular events in both macro- and micro-organisms. The identification and characterization of G4s can help to understand their different biological roles and potential applications in diagnosis and therapy. In addition to biophysical and biochemical methods to interrogate G4 formation, G4 fluorescent turn-on ligands can be used to target and visualize G4 formation both in vitro and in cells. Here, we review several representative classes of G4 fluorescent turn-on ligands in terms of their interaction mechanism and application perspectives. Interestingly, G4 structures are commonly identified in DNA and RNA aptamers against targets that include proteins and small molecules, which can be utilized as G4 tools for diverse applications. We therefore also summarize the recent development of G4-containing aptamers and highlight their applications in biosensing, bioimaging, and therapy. Moreover, we discuss the current challenges and future perspectives of G4 fluorescent turn-on ligands and G4-containing aptamers.

scholarly journals 2′-Fluoro-Pyrimidine-Modified RNA Aptamers Specific for Lipopolysaccharide Binding Protein (LBP)

2018 ◽  
Vol 19 (12) ◽  
pp. 3883 ◽  
Author(s):  
Jasmin Aldag ◽  
Tina Persson ◽  
Roland Hartmann
Keyword(s):  
In Vitro Selection ◽  
Binding Protein ◽  
High Specificity ◽  
Rna Aptamers ◽  
Pyrimidine Nucleotides ◽  
Consensus Sequences ◽  
Structure Probing ◽  
Potential Applications ◽  
Nuclease Degradation

Lipopolysaccaride binding protein (LBP), a glycosylated acute phase protein, plays an important role in the pathophysiology of sepsis. LBP binds with high affinity to the lipid part of bacterial lipopolysaccaride (LPS). Inhibition of the LPS-LBP interaction or blockage of LBP-mediated transfer of LPS monomers to CD14 may be therapeutical strategies to prevent septic shock. LBP is also of interest as a biomarker to identify septic patients at high risk for death, as LBP levels are elevated during early stages of severe sepsis. As a first step toward such potential applications, we isolated aptamers specific for murine LBP (mLBP) by in vitro selection from a library containing a 60-nucleotide randomized region. Modified RNA pools were transcribed in the presence of 2′-fluoro-modified pyrimidine nucleotides to stabilize transcripts against nuclease degradation. As verified for one aptamer experimentally, the selected aptamers adopt a “three-helix junction” architecture, presenting single-stranded 7-nt (5′-YGCTTCY) or 6-nt (5′-RTTTCY) consensus sequences in their core. The best binder (aptamer A011; Kd of 270 nM for binding to mLBP), characterized in more detail by structure probing and boundary analysis, was demonstrated to bind with high specificity to murine LBP.

scholarly journals Effect of tannin-rich leaves of oak (Quercus incana) on various microbial enzyme activities of the bovine rumen

1988 ◽  
Vol 60 (2) ◽  
pp. 287-296 ◽  
Author(s):  
H. P. S. Makkar ◽  
B. Singh ◽  
R. K. Dawra
Keyword(s):  
Enzyme Activities ◽  
Solid Matrix ◽  
Forward Reaction ◽  
Bovine Rumen ◽  
Dna And Rna ◽  
Microbial Enzyme ◽  
Micro Organisms ◽  
Oak Leaves

1. The objective of the present experiment was to study the effects of oak (Quercus incana) leaves rich in tannins on various enzyme activities of the bovine rumen.2. The procedure employed was incubation of tannin-rich, very-low-tannin or virtually tannin-free leaves in nylon-gauze bags in the rumen, and determination of enzyme activities in microbes tightly bound to the solid matrix and in microbes loosely plus tightly attached to the solid matrix.3. The activities of urease (EC3.5.1.5), carboxymethylcellulase, glutamate dehydrogenase (EC1.4.1.2) and alanine aminotransferase (glutamic-pyruvic transaminase) (EC2.6.1.2) were significantly lower in the tannin-rich group, whereas the activities of glutamate ammonia ligase (glutamine synthetase) (EC6.3.1.2; both yγ- glutamyltransferase (EC2.3.2.2) and the forward reaction) were higher in the tannin-rich group. These changes were more marked in micro-organisms tightly bound to the solid matrix than in the more complex microbial compartment.4. The protein, DNA and RNA contents, and protein: RNA ratio, were significantly lower in the tannin-rich group, whereas no difference was observed for protein: DNA between the groups.5. Effects of tannin-containing extracts of oak leaves on various rumen enzymes in vitro showed a trend similar to that observed in nylon-gauze bags, suggesting that the changes observed in various compartments were due to the tannins of oak leaves.

Antisense strategies and therapeutic applications

1996 ◽  
Vol 53 (2) ◽  
pp. 151-160 ◽  
Author(s):  
Putnam David A.
Keyword(s):  
Virus Infection ◽  
Messenger Rna ◽  
Therapeutic Agents ◽  
Water Soluble ◽  
Rna Sequences ◽  
Potential Applications ◽  
Antisense Approach ◽  
Simplex Virus

The concepts underlying the antisense approach to disease therapy are discussed, and potential applications are examined. Antisense therapeutic agents bind to DNA or RNA sequences, biocking the synthesis of cellular proteins with unparalleled specificity. Transcription and translation are the two processes with which the agents interfere. There are three major classes of antisense agents: antisense sequences, commonly called antisense oligonucleotides; antigene sequences; and ribozymes. Antisense sequences are derivatives of nucleic acids that hybridize cytosolic messenger RNA (mRNA) sense strands through hydrogen bonding to complementary nucleic acid bases. Antigene sequences hybridize double-stranded DNA in the nucleus, forming triple helixes. Ribozymes, rather than inhibiting protein synthesis simply by binding to a single targeted mRNA. combine enzymatic processes with the specificity of antisense Iwse pairing, creating a molecule that can incapacitate multiple targeted niRNAs. Anti-sense therapeutic agents are being investigated in vitro and in vivo for use in treating human immunodeficiency virus infection, hepatitis B virus infection, herpes simplex virus infection, papillomavirus infection, cancer, restenosis, rheumatoid arthritis, and allergic disorders. Although many results are preliminary, some are promising and has e led to clinical trials. A major goal in developing methods of delivering antisense agents is to reduce their susceptibility to nucleases while retaining their ability to bind to targeted sites. Modification of the phosphodiester linkages in oligonucleotides can lend the sequences enzymatic stability without affecting their binding capacities. Carrier systems designed to protect the antisense structure and improve passage through the cell membrane include liposomes, water-soluble polyrners, and nanoparticles. The pharmacokinetics of anti-sense agents are under investigation. Antisense therapeutic agents have the potential to become an integral part of medicinal regimens.

scholarly journals RNA beacons for detecting the human testis-determining factor

2021 ◽  
Author(s):  
Diego F. Joseph ◽  
Jose Alberto Nakamoto ◽  
Pohl Milón
Keyword(s):  
Specific Binding ◽  
Bioinformatic Analysis ◽  
Rna Aptamers ◽  
Alternative Methods ◽  
Human Testis ◽  
Hmg Box ◽  
Steady State Kinetics ◽  
Potential Applications ◽  
Determining Factor

AbstractThe testis-determining factor (TDF) is an essential transcriptional protein for male differentiation in mammals, expressed along spermatids to early zygotes and, to some extent, in diverse cellular lines. In this study, we developed fluorescent biosensors capable of indicating the presence of TDF. We usedin vitroevolution techniques to produce RNA aptamers that bind the recombinantly expressed HMG-box, the DNA binding domain of TDF. Bioinformatic analysis alongin vitroevolution setup suggested two predominant aptamer clusters with distinctive motifs. The top ranked aptamer from each cluster, M1 and M2, showed specific binding to TDF. Aptamers were fluorescently modified as molecular beacons. Pre-steady-state kinetics indicated the beacons bind rapidly, within 50 seconds, yet M1 showed better signal to noise ratios than M2. Structural predictions of the aptamer interaction indicated that M1 is composed by three stem loops and likely interact with the HMG-box of TDF through the pocket formed by the three loops. Molecular modelling of M1 beacon shows that binding to TDF entails a conformational change of the sensor resulting in the measured fluorescence changes. To our knowledge, this is the first work describing an RNA beacon for detecting the essential TDF. Potential applications and advantages over alternative methods are provided and discussed.

scholarly journals Tracking RNA with light: selection, structure, and design of fluorescence turn-on RNA aptamers

2019 ◽  
Vol 52 ◽  
Author(s):  
Robert J. Trachman ◽  
Adrian R. Ferré-D'Amaré
Keyword(s):  
Fluorescent Protein ◽  
Fluorescent Proteins ◽  
Structural Diversity ◽  
Rna Aptamers ◽  
Live Cells ◽  
Rna Molecules ◽  
Turn On ◽  
Green Fluorescent ◽  
Fluorescence Turn On

AbstractFluorescence turn-on aptamers,in vitroevolved RNA molecules that bind conditional fluorophores and activate their fluorescence, have emerged as RNA counterparts of the fluorescent proteins. Turn-on aptamers have been selected to bind diverse fluorophores, and they achieve varying degrees of specificity and affinity. These RNA–fluorophore complexes, many of which exceed the brightness of green fluorescent protein and their variants, can be used as tags for visualizing RNA localization and transport in live cells. Structure determination of several fluorescent RNAs revealed that they have diverse, unrelated overall architectures. As most of these RNAs activate the fluorescence of their ligands by restraining their photoexcited states into a planar conformation, their fluorophore binding sites have in common a planar arrangement of several nucleobases, most commonly a G-quartet. Nonetheless, each turn-on aptamer has developed idiosyncratic structural solutions to achieve specificity and efficient fluorescence turn-on. The combined structural diversity of fluorophores and turn-on RNA aptamers has already produced combinations that cover the visual spectrum. Further molecular evolution and structure-guided engineering is likely to produce fluorescent tags custom-tailored to specific applications.

scholarly journals Advances on Aptamers against Protozoan Parasites

Genes ◽  
2018 ◽  
Vol 9 (12) ◽  
pp. 584 ◽  
Author(s):  
Juan Ospina-Villa ◽  
César López-Camarillo ◽  
Carlos Castañón-Sánchez ◽  
Jacqueline Soto-Sánchez ◽  
Esther Ramírez-Moreno ◽  
...  
Keyword(s):  
Three Dimensional ◽  
Drug Carriers ◽  
Rna Aptamers ◽  
Health Concern ◽  
Dimensional Structure ◽  
Parasitic Infections ◽  
Attractive Alternative ◽  
Protozoan Parasites ◽  
Rna Sequences ◽  
Dna And Rna

Aptamers are single-stranded DNA or RNA sequences with a unique three-dimensional structure that allows them to recognize a particular target with high affinity. Although their specific recognition activity could make them similar to monoclonal antibodies, their ability to bind to a large range of non-immunogenic targets greatly expands their potential as tools for diagnosis, therapeutic agents, detection of food risks, biosensors, detection of toxins, drug carriers, and nanoparticle markers, among others. One aptamer named Pegaptanib is currently used for treating macular degeneration associated with age, and many other aptamers are in different clinical stages of development of evaluation for various human diseases. In the area of parasitology, research on aptamers has been growing rapidly in the past few years. Here we describe the development of aptamers raised against the main protozoan parasites that affect hundreds of millions of people in underdeveloped and developing countries, remaining a major health concern worldwide, i.e. Trypanosoma spp., Plasmodium spp., Leishmania spp., Entamoeba histolytica, and Cryptosporidium parvuum. The latest progress made in this area confirmed that DNA and RNA aptamers represent attractive alternative molecules in the search for new tools to detect and treat these parasitic infections that affect human health worldwide.

scholarly journals DDX5 helicase resolves G-quadruplex and is involved in MYC gene transcriptional activation

2019 ◽  
Vol 116 (41) ◽  
pp. 20453-20461 ◽  
Author(s):  
Guanhui Wu ◽  
Zheng Xing ◽  
Elizabeth J. Tran ◽  
Danzhou Yang
Keyword(s):  
Transcriptional Activation ◽  
Atp Hydrolysis ◽  
Proximal Promoter ◽  
Rna Sequences ◽  
Dna And Rna ◽  
Highly Active ◽  
Dead Box ◽  
Myc Gene ◽  
Cancer Intervention

G-quadruplexes (G4) are noncanonical secondary structures formed in guanine-rich DNA and RNA sequences. MYC, one of the most critical oncogenes, forms a DNA G4 in its proximal promoter region (MycG4) that functions as a transcriptional silencer. However, MycG4 is highly stable in vitro and its regulatory role would require active unfolding. Here we report that DDX5, one of the founding members of the DEAD-box RNA helicase family, is extremely proficient at unfolding MycG4-DNA. Our results show that DDX5 is a highly active G4-resolvase that does not require a single-stranded overhang and that ATP hydrolysis is not directly coupled to G4-unfolding of DDX5. The chromatin binding sites of DDX5 are G-rich sequences. In cancer cells, DDX5 is enriched at the MYC promoter and activates MYC transcription. The DDX5 interaction with the MYC promoter and DDX5-mediated MYC activation is inhibited by G4-interactive small molecules. Our results uncover a function of DDX5 in resolving DNA and RNA G4s and suggest a molecular target to suppress MYC for cancer intervention.

scholarly journals In vitro selection of l-DNA aptamers that bind a structured d-RNA molecule

2020 ◽  
Vol 48 (4) ◽  
pp. 1669-1680 ◽  
Author(s):  
Sougata Dey ◽  
Jonathan T Sczepanski
Keyword(s):  
In Vitro Selection ◽  
Rna Binding ◽  
Rna Aptamers ◽  
Dna Aptamers ◽  
Binding Properties ◽  
Rna Molecules ◽  
Dna And Rna ◽  
Selection Of

Abstract The development of structure-specific RNA binding reagents remains a central challenge in RNA biochemistry and drug discovery. Previously, we showed in vitro selection techniques could be used to evolve l-RNA aptamers that bind tightly to structured d-RNAs. However, whether similar RNA-binding properties can be achieved using aptamers composed of l-DNA, which has several practical advantages compared to l-RNA, remains unknown. Here, we report the discovery and characterization of the first l-DNA aptamers against a structured RNA molecule, precursor microRNA-155, thereby establishing the capacity of DNA and RNA molecules of the opposite handedness to form tight and specific ‘cross-chiral’ interactions with each other. l-DNA aptamers bind pre-miR-155 with low nanomolar affinity and high selectivity despite the inability of l-DNA to interact with native d-RNA via Watson–Crick base pairing. Furthermore, l-DNA aptamers inhibit Dicer-mediated processing of pre-miRNA-155. The sequence and structure of l-DNA aptamers are distinct from previously reported l-RNA aptamers against pre-miR-155, indicating that l-DNA and l-RNA interact with the same RNA sequence through unique modes of recognition. Overall, this work demonstrates that l-DNA may be pursued as an alternative to l-RNA for the generation of RNA-binding aptamers, providing a robust and practical approach for targeting structured RNAs.

scholarly journals Killing of adherent oral microbes by a non-thermal atmospheric plasma jet

2010 ◽  
Vol 59 (2) ◽  
pp. 206-212 ◽  
Author(s):  
Stefan Rupf ◽  
Antje Lehmann ◽  
Matthias Hannig ◽  
Barbara Schäfer ◽  
Andreas Schubert ◽  
...  
Keyword(s):  
Plasma Jet ◽  
Plasma Jets ◽  
Atmospheric Plasma ◽  
Killing Effect ◽  
E Coli ◽  
Potential Applications ◽  
Oral Microbes ◽  
Micro Organisms

Atmospheric plasma jets are being intensively studied with respect to potential applications in medicine. The aim of this in vitro study was to test a microwave-powered non-thermal atmospheric plasma jet for its antimicrobial efficacy against adherent oral micro-organisms. Agar plates and dentin slices were inoculated with 6 log10 c.f.u. cm−2 of Lactobacillus casei, Streptococcus mutans and Candida albicans, with Escherichia coli as a control. Areas of 1 cm2 on the agar plates or the complete dentin slices were irradiated with a helium plasma jet for 0.3, 0.6 or 0.9 s mm−2, respectively. The agar plates were incubated at 37 °C, and dentin slices were vortexed in liquid media and suspensions were placed on agar plates. The killing efficacy of the plasma jet was assessed by counting the number of c.f.u. on the irradiated areas of the agar plates, as well as by determination of the number of c.f.u. recovered from dentin slices. A microbe-killing effect was found on the irradiated parts of the agar plates for L. casei, S. mutans, C. albicans and E. coli. The plasma-jet treatment reduced the c.f.u. by 3–4 log10 intervals on the dentin slices in comparison to recovery rates from untreated controls. The microbe-killing effect was correlated with increasing irradiation times. Thus, non-thermal atmospheric plasma jets could be used for the disinfection of dental surfaces.

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