Sequence specificity for uridylylation of the viral peptide linked to the genome (VPg) of enteroviruses

Schein, Catherine H.; Ye, Mengyi; Paul, Aniko V.; Oberste, M. Steven; Chapman, Nora M.; Noort, Gerbrand J. van der Heden van; Filippov, Dmitri V.; Choi, Kyung Hyun

doi:10.1016/j.virol.2015.05.016

Cited by 17 publications

(15 citation statements)

References 52 publications

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“…This reaction uses the normal 3D pol active site in a macromolecular complex where the VPg tyrosine hydroxyl is presumably positioned in the active site to mimic a RNA 3′ hydroxyl group. In vitro uridylylation is possible using a mixture of 3D pol , VPg, poly(A) RNA, and UTP, and for this reaction there is significant cross-reactivity between different viral 3D pol and VPg proteins (Schein et al, 2015). However, uridylylation is much more efficient with cre RNA and 3CD pro as cofactors to form a complex with a significantly lower K m for UTP (Paul et al, 2000; Steil and Barton, 2008; Yin et al, 2003).…”

Section: 3dpol–vpg Complexesmentioning

confidence: 99%

Picornaviral polymerase structure, function, and fidelity modulation

Peersen

2017

Virus Research

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Like all positive strand RNA viruses, the picornaviruses replicate their genomes using a virally encoded RNA-dependent RNA polymerase enzyme known as 3Dpol. Over the past decade we have made tremendous advances in our understanding of 3Dpol structure and function, including the discovery of a novel mechanism for closing the active site that allows these viruses to easily fine tune replication fidelity and quasispecies distributions. This review summarizes current knowledge of picornaviral polymerase structure and how the enzyme interacts with RNA and other viral proteins to form stable and processive elongation complexes. The picornaviral RdRPs are among the smallest viral polymerases, but their fundamental molecular mechanism for catalysis appears to be generally applicable as a common feature of all positive strand RNA virus polymerases.

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Section: 3dpol–vpg Complexesmentioning

confidence: 99%

Picornaviral polymerase structure, function, and fidelity modulation

Peersen

2017

Virus Research

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“…Previous work showed that the CVA24 3D pol can uridylylate VPg peptides from even very distantly related EV 28 . As Figures 3 and 5 indicate, it is a very active protein.…”

Section: Discussionmentioning

confidence: 99%

“…CVA24 and the PCP consensus VPgs were synthesized and CVA24 3D pol and Dengue virus (DENV) NS5 polymerase (used for control assays not shown) were purified as described elsewhere 28, 35 . The CVA24 polymerase was concentrated to about 3 mg/ml and stored at −20°C in PB with additional glycerol added to a final concentration of 18%.…”

Section: Methodsmentioning

confidence: 99%

“…All compounds were initially docked to a grid limited to the VPg binding site (white box, Figure 2). The sequence in this area is conserved in EV-3D pol of species C and B 28 .The “small” docking grid, which covered the conserved allosteric site, was centered on five residues (F377, R379, A380, D381 and E382) shown by mutagenesis to be important for uridylylation but not RNA elongation 32 . This location was determined based on docking 33 and mutagenesis studies 37 , and comparison to the CVB3 co-crystal structure 38 (Figure 1, right).…”

Section: Methodsmentioning

confidence: 99%

“…Although all four EV-3D pol tested showed preference for their cognate VPg, they could also uridylylate a PCP-consensus 24–27 VPg designed to represent the p hysical c hemical p roperties of 31 different EV-VPgs 28 . Thus, the residues and mechanism required for uridylylation must be similar in most, if not all, EV, indicating that specific inhibitors of this reaction could be promising pan-EV therapeutics.…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Allosteric inhibitors of Coxsackie virus A24 RNA polymerase

Schein

Rowold

Choi

2016

Bioorganic & Medicinal Chemistry

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Coxsackie virus A24 (CVA24), a causative agent of acute hemorrhagic conjunctivitis, is a prototype of enterovirus (EV) species C. The RNA polymerase (3Dpol) of CVA24 can uridylylate the viral peptide linked to the genome (VPg) from distantly related EV, and is thus, a good model for studying this reaction. Once UMP is bound, VPgpU primes RNA elongation. Structural and mutation data have identified a conserved binding surface for VPg on the RNA polymerase (3Dpol), located about 20Å from the active site. Here, computational docking of over 60,000 small compounds was used to select those with the lowest (best) specific binding energies (BE) for this allosteric site. Compounds with varying structures and low BE were assayed for their effect on formation of VPgU by CVA24-3Dpol. Two compounds with the lowest specific BE for the site inhibited both uridylylation and formation of VPgpolyU at 10–20 µM. These small molecules can be used to probe the role of this allosteric site in polymerase function, and may be the basis for novel antiviral compounds.

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Repurposing approved drugs on the pathway to novel therapies

Schein

2019

Medicinal Research Reviews

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The time and cost of developing new drugs have led many groups to limit their search for therapeutics to compounds that have previously been approved for human use. Many “repurposed” drugs, such as derivatives of thalidomide, antibiotics, and antivirals have had clinical success in treatment areas well beyond their original approved use. These include applications in treating antibiotic‐resistant organisms, viruses, cancers and to prevent burn scarring. The major theoretical justification for reusing approved drugs is that they have known modes of action and controllable side effects. Coadministering antibiotics with inhibitors of bacterial toxins or enzymes that mediate multidrug resistance can greatly enhance their activity. Drugs that control host cell pathways, including inflammation, tumor necrosis factor, interferons, and autophagy, can reduce the “cytokine storm” response to injury, control infection, and aid in cancer therapy. An active compound, even if previously approved for human use, will be a poor clinical candidate if it lacks specificity for the new target, has poor solubility or can cause serious side effects. Synergistic combinations can reduce the dosages of the individual components to lower reactivity. Preclinical analysis should take into account that severely ill patients with comorbidities will be more sensitive to side effects than healthy trial subjects. Once an active, approved drug has been identified, collaboration with medicinal chemists can aid in finding derivatives with better physicochemical properties, specificity, and efficacy, to provide novel therapies for cancers, emerging and rare diseases.

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Sequence specificity for uridylylation of the viral peptide linked to the genome (VPg) of enteroviruses

Cited by 17 publications

References 52 publications

Picornaviral polymerase structure, function, and fidelity modulation

Picornaviral polymerase structure, function, and fidelity modulation

Allosteric inhibitors of Coxsackie virus A24 RNA polymerase

Repurposing approved drugs on the pathway to novel therapies

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