Structural insights of the conserved “priming loop” of hepatitis B virus pre-genomic RNA

LeBlanc, Regan M.; Kasprzak, Wojciech K.; Longhini, Andrew P.; Olenginski, Lukasz T.; Abulwerdi, Fardokht A.; Ginocchio, S.; Shields, B.; Nyman, J.; Svirydava, Maryia; Vecchio, Claudia Del; Ivanic, Joseph; Schneekloth, John S.; Shapiro, Bruce A.; Dayie, T. Kwaku; Grice, Stuart F. J. Le

doi:10.1080/07391102.2021.1934544

Cited by 17 publications

(78 citation statements)

References 56 publications

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“…Palbociclib binds to HIV TAR with a KD of 100-300 nM, depending on ionic conditions, comparable to its reported affinity for cdk6 (KD ~60nM) 24 , which was only achieved following a decade-long medicinal chemistry effort. Other studies have reported binding of FDA approved drugs to RNA [42][43][44] , including kinase inhibitors 45 , but binding in these cases was 100-1,000 fold weaker than we observe 25 or non-specific 46,47 . It is sometimes stated that kinase inhibitors make good RNA binding molecules, but we have shown that RNA binding can be obtained fully independently of kinase inhibition.…”

Section: Discussioncontrasting

confidence: 89%

The kinase inhibitor Palbociclib is a potent and specific RNA-binding molecule

Shortridge

Vidalala

Varani

2022

Preprint

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The growing awareness of the role of RNA in human disease has motivated significant efforts to discover drug-like small molecules that target RNA. However, high throughput screening campaigns report very low hit rates and generally identify compounds with weak affinity, while most structures reported in Academic studies also lack the pharmacological properties of successful drugs. Even FDA-approved RNA-targeting drugs have only weak (10 uM) binding activity. Thus, it is often stated that only complex RNA structures, such as the ribosome or riboswitches, are amenable to small molecule chemistry. We report that the kinase inhibitor Palbociclib/Ibrance is a nM ligand for the HIV-1 TAR. It inhibits recruitment of the positive transcription elongation factor complex at nM concentrations and discriminates >20 fold. We further show that RNA binding can be fully decoupled from kinase inhibition, yielding a new molecule with even higher affinity for RNA. We thus demonstrate that nM affinity, specificity, and potent biochemical activity against 'undruggable' RNAs can be found in the chemical space of blockbuster drugs.

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Section: Discussioncontrasting

confidence: 89%

The kinase inhibitor Palbociclib is a potent and specific RNA-binding molecule

Shortridge

Vidalala

Varani

2022

Preprint

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“…The HBV ε RNA stem-loop that templates DNA priming [PDB: 6VAR (45)] was also docked into the full-length model of genotype B P (Figs. 5C and S29A-C).…”

Section: The Fingers Subdomain Of Thementioning

confidence: 99%

Predicted structure of the hepatitis B virus polymerase reveals an ancient conserved protein fold

Tajwar

Bradley

Ponzar

et al. 2022

Preprint

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Hepatitis B virus (HBV) replicates by protein-primed reverse transcription. It chronically infects >250 million people, and the dominant anti-HBV drugs are nucleos(t)ide analogs targeting the viral polymerase (P). P has four domains, the terminal protein (TP) that primes DNA synthesis, a spacer, the reverse transcriptase (RT), and the ribonuclease H (RNaseH). Despite being a major drug target and catalyzing a reverse transcription pathway very different from the retroviral pathway, HBV P has resisted structural analysis for decades. Here, we exploited advances in protein structure prediction to model the structure of P. The predicted HBV RT and RNaseH domains aligned to the HIV RT-RNaseH at 3.75 A RMSD, had a nucleic acid binding groove spanning the two active sites, had DNA polymerase active site motifs in their expected positions, and accommodated two Mg++ ions in both active sites. Surprisingly, the TP domain wrapped around the RT domain, with the priming tyrosine poised over the RT active site. This model was validated using published mutational analyses, and by docking RT and RNaseH inhibitors, RNA:DNA heteroduplexes, and the HBV e RNA stem-loop that templates DNA priming into the model. The HBV P fold, including the orientation of the TP domain, was conserved among hepadnaviruses from rodents to fish and in P from a fish nackednavirus, but not in other non-retroviral RTs. Therefore, this protein fold has persisted since the hepadnaviruses diverged from nackednaviruses >400 million years ago. This model will guide drug development and mechanistic studies into P function.

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“…Taken together, the techniques show the presence of flexibility of SLIV both in and out of complex. Both Hepatitis B (HBV) and HCV have also been investigated using SAXS [59,60]. SAXS data helped to support an NMR structural determination of the 6nt HBV priming loop, in which the partially stacked G16 and U17 were identified as potential residues responsible for initiating DNA synthesis [59].…”

Section: Saxsmentioning

confidence: 99%

“…Both Hepatitis B (HBV) and HCV have also been investigated using SAXS [59,60]. SAXS data helped to support an NMR structural determination of the 6nt HBV priming loop, in which the partially stacked G16 and U17 were identified as potential residues responsible for initiating DNA synthesis [59]. HCV encodes a noncoding 3 domain known as 3 X which contains a 16nt palindromic genome dimerization signal that partially overlaps with a 7nt sequence that forms a long-range interaction with a complementary site within the ORF.…”

Section: Saxsmentioning

confidence: 99%

Evaluating RNA Structural Flexibility: Viruses Lead the Way

Fairman

Lever

Kenyon

2021

Viruses

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Our understanding of RNA structure has lagged behind that of proteins and most other biological polymers, largely because of its ability to adopt multiple, and often very different, functional conformations within a single molecule. Flexibility and multifunctionality appear to be its hallmarks. Conventional biochemical and biophysical techniques all have limitations in solving RNA structure and to address this in recent years we have seen the emergence of a wide diversity of techniques applied to RNA structural analysis and an accompanying appreciation of its ubiquity and versatility. Viral RNA is a particularly productive area to study in that this economy of function within a single molecule admirably suits the minimalist lifestyle of viruses. Here, we review the major techniques that are being used to elucidate RNA conformational flexibility and exemplify how the structure and function are, as in all biology, tightly linked.

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Structural insights of the conserved “priming loop” of hepatitis B virus pre-genomic RNA

Cited by 17 publications

References 56 publications

The kinase inhibitor Palbociclib is a potent and specific RNA-binding molecule

The kinase inhibitor Palbociclib is a potent and specific RNA-binding molecule

Predicted structure of the hepatitis B virus polymerase reveals an ancient conserved protein fold

Evaluating RNA Structural Flexibility: Viruses Lead the Way

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