2-Aminobenzoxazole ligands of the hepatitis C virus internal ribosome entry site

Rynearson, Kevin D.; Charrette, Brian Paul; Gabriel, Christopher; Moreno, Jesus; Boerneke, Mark A.; Dibrov, Sergey M.; Hermann, Thomas

doi:10.1016/j.bmcl.2014.05.088

Cited by 31 publications

(41 citation statements)

References 18 publications

(20 reference statements)

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“…4F, translocation of the HCV subdomain IIa RNA construct is sterically hindered when using a ~3 nm diameter nanopore, while passage of the construct after it adopts its drug-induced linear conformation is largely unhindered. To sense this conformational shift, nanopore experiments were conducted with the RNA construct in its natively bent conformation, as well in the presence of two benzimidazole derived drugs that bind with different affinities ( 1 : K d = 3.4 μM, 2 : K d = 64 μM, measured by an independent FRET assay 42,44 ). Dwell-time histograms in the presence of different drug concentrations are shown in Fig.…”

Section: Point-of-care Applicationsmentioning

confidence: 99%

Studies of RNA Sequence and Structure Using Nanopores

Henley

Carson

Wanunu

2016

Progress in Molecular Biology and Translational Science

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Nanopores are powerful single-molecule sensors with nanometer scale dimensions suitable for detection, quantification, and characterization of nucleic acids and proteins. Beyond sequencing applications, both biological and solid-state nanopores hold great promise as tools for studying the biophysical properties of RNA. In this review, we highlight selected landmark nanopore studies with regards to RNA sequencing, microRNA detection, RNA/ligand interactions, and RNA structural/conformational analysis.

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Section: Point-of-care Applicationsmentioning

confidence: 99%

Studies of RNA Sequence and Structure Using Nanopores

Henley

Carson

Wanunu

2016

Progress in Molecular Biology and Translational Science

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“…5 Two drugs were used in this study, as shown in Figure 2a: positive control drug 1 has been shown using FRET to bind to the elbow region of the IRES domain with a dissociation constant of ~3 μ M, 5 while negative control 2 is an analogue drug with a much weaker affinity of ~64 μ M. 26 Therefore, in this study compound 1 serves as a positive control and compound 2 as a negative control. Binding and conformational change induced by compound 1 to the HCV RNA was confirmed by repeating the FRET assay performed by Parsons and co-workers (Figure 2b).…”

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confidence: 99%

Nanopore-Based Conformational Analysis of a Viral RNA Drug Target

et al. 2014

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Nanopores are single-molecule sensors that show exceptional promise as a biomolecular analysis tool by enabling label-free detection of small amounts of sample. In this paper, we demonstrate that nanopores are capable of detecting the conformation of an antiviral RNA drug target. The hepatitis C virus uses an internal ribosome entry site (IRES) motif in order to initiate translation by docking to ribosomes in its host cell. The IRES is therefore a viable and important drug target. Drug-induced changes to the conformation of the HCV IRES motif, from a bent to a straight conformation, have been shown to inhibit HCV replication. However, there is presently no straightforward method to analyze the effect of candidate small-molecule drugs on the RNA conformation. In this paper, we show that RNA translocation dynamics through a 3 nm diameter nanopore is conformation-sensitive by demonstrating a difference in transport times between bent and straight conformations of a short viral RNA motif. Detection is possible because bent RNA is stalled in the 3 nm pore, resulting in longer molecular dwell times than straight RNA. Control experiments show that binding of a weaker drug does not produce a conformational change, as consistent with independent fluorescence measurements. Nanopore measurements of RNA conformation can thus be useful for probing the structure of various RNA motifs, as well as structural changes to the RNA upon small-molecule binding.

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“…The IRES subdomain IIa switch has been exploited as a therapeutic target to selectively suppress viral translation with synthetic inhibitors that capture specific conformations of the RNA (5,7,(9)(10)(11)(12). High levels of conservation in HCV clinical isolates and the presence of a deep solvent-excluded ligand binding pocket render the IIa RNA switch a target for the development of antiviral drugs.…”

Section: Significance and Applications Of The Iia Switchesmentioning

confidence: 99%

Ligand-responsive RNA mechanical switches

Boerneke

Hermann

2015

RNA Biology

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Ligand-responsive RNA mechanical switches represent a new class of simple switching modules that adopt well-defined ligand-free and bound conformational states, distinguishing them from metabolite-sensing riboswitches. Initially discovered in the internal ribosome entry site (IRES) of hepatitis C virus (HCV), these RNA switch motifs were found in the genome of diverse other viruses. Although large variations are seen in sequence and local secondary structure of the switches, their function in viral translation initiation that requires selective ligand recognition is conserved. We recently determined the crystal structure of an RNA switch from Seneca Valley virus (SVV) which is able to functionally replace the switch of HCV. The switches from both viruses recognize identical cognate ligands despite their sequence dissimilarity. Here, we describe the discovery of 7 new switches in addition to the previously established 5 examples. We highlight structural and functional features unique to this class of ligand-responsive RNA mechanical switches and discuss implications for therapeutic development and the construction of RNA nanostructures.

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2-Aminobenzoxazole ligands of the hepatitis C virus internal ribosome entry site

Cited by 31 publications

References 18 publications

Studies of RNA Sequence and Structure Using Nanopores

Studies of RNA Sequence and Structure Using Nanopores

Nanopore-Based Conformational Analysis of a Viral RNA Drug Target

Ligand-responsive RNA mechanical switches

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