scholarly journals Antiviral peptides targeting the helicase activity of enterovirus nonstructural protein 2C

2021 ◽  
Author(s):  
Yuan Fang ◽  
Chang Wang ◽  
Chong Wang ◽  
Ruyi Yang ◽  
Peng Bai ◽  
...  
Keyword(s):  
Broad Spectrum ◽  
Nonstructural Protein ◽  
Rna Helicase ◽  
Coxsackievirus B3 ◽  
Human Enterovirus ◽  
Helicase Activity ◽  
Coxsackievirus A16 ◽  
Enterovirus A71 ◽  
In Cells

Enteroviruses belong to the genus Enterovirus of the family Picornaviridae and include four human enterovirus groups (EV A-D), and the epidemic of enteroviruses such as human enterovirus A71 (EV-A71) and Coxsackievirus-A16 (CVA16) is a threat to global public health. Enteroviral 2C is the most conserved nonstructural protein among all enteroviruses and possesses the RNA helicase activity that plays pivotal roles during enteroviral life cycles, which makes 2C an attractive target for developing the anti-enteroviral drugs. In this study, we designed a peptide, named 2CL, based on the structure of EV-A71 2C. This peptide effectively impaired the oligomerization of EV-A71 2C protein, and inhibited the RNA helicase activities of 2C proteins encoded by EV-A71 and CVA16, and both of which belong to EV-A, and showed potent antiviral efficacy against EV-A71 and CVA16 in cells. Moreover, the 2CL treatment elicited a strong in vivo protective efficacy against lethal EV-A71 challenge. Besides, the antiviral strategy of targeting the 2C helicase activity can be applied to inhibit the replication of EV-B. Either 2CL or B-2CL, the peptide redesigned based on the 2CL-corresponding sequence of EV-Bs, exerted effective antiviral activity against two important EV-Bs, Coxsackievirus B3 and Echovirus 11. Together, our findings demonstrated that targeting the helicase activity of 2C by rationally designed peptide is an efficient antiviral strategy against enteroviruses, and the 2CL and B-2CL showed promising clinical potentials to be further developed as broad-spectrum anti-enteroviral drugs. Importance Enteroviruses are a large group of positive-sense single-stranded RNA viruses, and include numerous human pathogens, such as enterovirus A71 (EV-A71), coxsackieviruses, and echoviruses. However, no approved antiviral drug is available. Enteroviral 2C is the most conserved nonstructural protein among all enteroviruses and contains the RNA helicase activity critical for the viral life cycle. Herein, according to the structure of EV-A71 2C, we designed a peptide that effectively inhibited the RNA helicase activities of EV-A71-and coxsackievirus A16 (CVA16)-encoded 2C proteins. Moreover, this peptide exerted potent antiviral effects against EV-A71 and CVA16 in cells and elicited therapeutic efficacy against lethal EV-A71 challenge in vivo. Furthermore, we demonstrated that the strategy of targeting the 2C helicase activity can be used to other relevant enteroviruses, including coxsackievirus B3 and echovirus 11. In summary, our findings provide compelling evidence that the designed peptides targeting the helicase activity of 2C could be broad-spectrum antiviral for enteroviruses.

scholarly journals Viruses harness YxxØ motif to interact with host AP2M1 for replication: A vulnerable broad-spectrum antiviral target

2020 ◽  
Vol 6 (35) ◽  
pp. eaba7910
Author(s):  
Shuofeng Yuan ◽  
Hin Chu ◽  
Jingjing Huang ◽  
Xiaoyu Zhao ◽  
Zi-Wei Ye ◽  
...  
Keyword(s):  
Broad Spectrum ◽  
Influenza A ◽  
Host Protein ◽  
Influenza A Viruses ◽  
Protein Protein Interaction ◽  
Enterovirus A71 ◽  
Evolutionarily Conserved ◽  
Immunodeficiency Virus

Targeting a universal host protein exploited by most viruses would be a game-changing strategy that offers broad-spectrum solution and rapid pandemic control including the current COVID-19. Here, we found a common YxxØ-motif of multiple viruses that exploits host AP2M1 for intracellular trafficking. A library chemical, N-(p-amylcinnamoyl)anthranilic acid (ACA), was identified to interrupt AP2M1-virus interaction and exhibit potent antiviral efficacy against a number of viruses in vitro and in vivo, including the influenza A viruses (IAVs), Zika virus (ZIKV), human immunodeficiency virus, and coronaviruses including MERS-CoV and SARS-CoV-2. YxxØ mutation, AP2M1 depletion, or disruption by ACA causes incorrect localization of viral proteins, which is exemplified by the failure of nuclear import of IAV nucleoprotein and diminished endoplasmic reticulum localization of ZIKV-NS3 and enterovirus-A71-2C proteins, thereby suppressing viral replication. Our study reveals an evolutionarily conserved mechanism of protein-protein interaction between host and virus that can serve as a broad-spectrum antiviral target.

scholarly journals Structure-Based Mutational Analysis of the Hepatitis C Virus NS3 Helicase

2001 ◽  
Vol 75 (17) ◽  
pp. 8289-8297 ◽  
Author(s):  
Chun-Ling Tai ◽  
Wen-Ching Pan ◽  
Shwu-Huey Liaw ◽  
Ueng-Cheng Yang ◽  
Lih-Hwa Hwang ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Atpase Activity ◽  
Rna Binding ◽  
Mutational Analysis ◽  
Nonstructural Protein ◽  
Rna Helicase ◽  
Helicase Domain ◽  
Helicase Activity ◽  
Conserved Sequence

ABSTRACT The carboxyl terminus of the hepatitis C virus (HCV) nonstructural protein 3 (NS3) possesses ATP-dependent RNA helicase activity. Based on the conserved sequence motifs and the crystal structures of the helicase domain, 17 mutants of the HCV NS3 helicase were generated. The ATP hydrolysis, RNA binding, and RNA unwinding activities of the mutant proteins were examined in vitro to determine the functional role of the mutated residues. The data revealed that Lys-210 in the Walker A motif and Asp-290, Glu-291, and His-293 in the Walker B motif were crucial to ATPase activity and that Thr-322 and Thr-324 in motif III and Arg-461 in motif VI significantly influenced ATPase activity. When the pairing between His-293 and Gln-460, referred to as gatekeepers, was replaced with the Asp-293/His-460 pair, which makes the NS3 helicase more like the DEAD helicase subgroup, ATPase activity was not restored. It thus indicated that the whole microenvironment surrounding the gatekeepers, rather than the residues per se, was important to the enzymatic activities. Arg-461 and Trp-501 are important residues for RNA binding, while Val-432 may only play a coadjutant role. The data demonstrated that RNA helicase activity was possibly abolished by the loss of ATPase activity or by reduced RNA binding activity. Nevertheless, a low threshold level of ATPase activity was found sufficient for helicase activity. Results in this study provide a valuable reference for efforts under way to develop anti-HCV therapeutic drugs targeting NS3.

scholarly journals Human Enterovirus Nonstructural Protein 2CATPase Functions as Both an RNA Helicase and ATP-Independent RNA Chaperone

2015 ◽  
Vol 11 (7) ◽  
pp. e1005067 ◽  
Author(s):  
Hongjie Xia ◽  
Peipei Wang ◽  
Guang-Chuan Wang ◽  
Jie Yang ◽  
Xianlin Sun ◽  
...  
Keyword(s):  
Nonstructural Protein ◽  
Rna Helicase ◽  
Human Enterovirus ◽  
Rna Chaperone

scholarly journals RNase R is associated in a functional complex with the RhpA DEAD-box RNA helicase in Helicobacter pylori

2021 ◽  
Author(s):  
Alejandro Tejada-Arranz ◽  
Rute G Matos ◽  
Yves Quentin ◽  
Maxime Bouilloux-Lafont ◽  
Eloïse Galtier ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Gene Expression Regulation ◽  
Rna Helicase ◽  
Minor Role ◽  
Helicase Activity ◽  
Rnase R ◽  
Dead Box ◽  
H Pylori

Abstract Ribonucleases are central players in post-transcriptional regulation, a major level of gene expression regulation in all cells. Here, we characterized the 3′-5′ exoribonuclease RNase R from the bacterial pathogen Helicobacter pylori. The ‘prototypical’ Escherichia coli RNase R displays both exoribonuclease and helicase activities, but whether this latter RNA unwinding function is a general feature of bacterial RNase R had not been addressed. We observed that H. pylori HpRNase R protein does not carry the domains responsible for helicase activity and accordingly the purified protein is unable to degrade in vitro RNA molecules with secondary structures. The lack of RNase R helicase domains is widespread among the Campylobacterota, which include Helicobacter and Campylobacter genera, and this loss occurred gradually during their evolution. An in vivo interaction between HpRNase R and RhpA, the sole DEAD-box RNA helicase of H. pylori was discovered. Purified RhpA facilitates the degradation of double stranded RNA by HpRNase R, showing that this complex is functional. HpRNase R has a minor role in 5S rRNA maturation and few targets in H. pylori, all included in the RhpA regulon. We concluded that during evolution, HpRNase R has co-opted the RhpA helicase to compensate for its lack of helicase activity.

scholarly journals The NS4A Protein of Hepatitis C Virus Promotes RNA-Coupled ATP Hydrolysis by the NS3 Helicase

2009 ◽  
Vol 83 (7) ◽  
pp. 3268-3275 ◽  
Author(s):  
Rudolf K. F. Beran ◽  
Brett D. Lindenbach ◽  
Anna Marie Pyle
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Serine Protease ◽  
Protease Activity ◽  
Rna Binding ◽  
Nonstructural Protein ◽  
Helicase Activity ◽  
Protease Domain

ABSTRACT Nonstructural protein 3 (NS3) is an essential replicative component of the hepatitis C virus (HCV) and a member of the DExH/D-box family of proteins. The C-terminal region of NS3 (NS3hel) exhibits RNA-stimulated NTPase and helicase activity, while the N-terminal serine protease domain of NS3 enhances RNA binding and unwinding by NS3hel. The nonstructural protein 4A (NS4A) binds to the NS3 protease domain and serves as an obligate cofactor for NS3 serine protease activity. Given its role in stimulating protease activity, we sought to determine whether NS4A also influences the activity of NS3hel. Here we show that NS4A enhances the ability of NS3hel to bind RNA in the presence of ATP, thereby acting as a cofactor for helicase activity. This effect is mediated by amino acids in the C-terminal acidic domain of NS4A. When these residues are mutated, one observes drastic reductions in ATP-coupled RNA binding and duplex unwinding by NS3. These same mutations are lethal in HCV replicons, thereby establishing in vitro and in vivo that NS4A plays an important role in the helicase mechanism of NS3 and its function in replication.

scholarly journals Inhibition of RNA Helicase Activity Prevents Coxsackievirus B3-Induced Myocarditis in Human iPS Cardiomyocytes

2020 ◽  
Vol 21 (9) ◽  
pp. 3041
Author(s):  
Soo-Hyeon Yun ◽  
Ha-Hyeon Shin ◽  
Eun-Seon Ju ◽  
You-Jung Lee ◽  
Byung-Kwan Lim ◽  
...  
Keyword(s):  
Heart Function ◽  
Myocardial Damage ◽  
Rna Helicase ◽  
Coxsackievirus B3 ◽  
Host Protein ◽  
Mouse Heart ◽  
Vitro Assay ◽  
Chronic Myocarditis

Aims: Coxsackievirus B3 (CVB3) is known to be an important cause of myocarditis and dilated cardiomyopathy. Enterovirus-2C (E2C) is a viral RNA helicase. It inhibits host protein synthesis. Based on these facts, we hypothesize that the inhibition of 2C may suppress virus replication and prevent enterovirus-mediated cardiomyopathy. Methods and Results: We generated a chemically modified enterovirus-2C inhibitor (E2CI). From the in vitro assay, E2CI was showed strong antiviral effects. For in vivo testing, mice were treated with E2CI intraperitoneally injected daily for three consecutive days at a dose of 8 mg/kg per day, after CVB3 post-infection (p.i) (CVB3 + E2CI, n = 33). For the infected controls (CVB3 only, n = 35), mice were injected with PBS (phosphate buffered saline) in a DBA/2 strain to establish chronic myocarditis. The four-week survival rate of E2CI-treated mice was significantly higher than that of controls (92% vs. 71%; p < 0.05). Virus titers and myocardial damage were significantly reduced in the E2CI treated group. In addition, echocardiography indicated that E2CI administration dramatically maintained mouse heart function compared to control at day 28 p.i chronic stage (LVIDD, 3.1 ± 0.08 vs. 3.9 ± 0.09, p < 0.01; LVDS, 2.0 ± 0.07 vs. 2.5 ± 0.07, p < 0.001; FS, 34.8 ± 1.6% vs. 28.5 ± 1.5%; EF, 67. 9 ± 2.9% vs. 54.7 ± 4.7%, p < 0.05; CVB3 + E2CI, n = 6 vs. CVB3, n = 4). Moreover, E2CI is effectively worked in human iPS (induced pluripotent stem cell) derived cardiomyocytes. Conclusion: Enterovirus-2C inhibitor (E2CI) was significantly reduced viral replication, chronic myocardium damage, and CVB3-induced mortality in DBA/2 mice. These results suggested that E2CI is a novel therapeutic agent for the treatment of enterovirus-mediated diseases.

Effect of Some Metabolic Inhibitors on Vinblastine-Induced Ribosomal-Granular Material Complexes

1971 ◽  
Vol 29 ◽  
pp. 548-549
Author(s):  
Awtar Krishan ◽  
Dora Hsu
Keyword(s):  
Granular Material ◽  
Rna Synthesis ◽  
Culture Medium ◽  
Actinomycin D ◽  
Metabolic Inhibitors ◽  
Protein And Rna Synthesis ◽  
Apparent Reduction ◽  
Plant Alkaloids ◽  
In Cells

Cells exposed to antitumor plant alkaloids, vinblastine and vincristine sulfate have large proteinacious crystals and complexes of ribosomes, helical polyribosomes and electron-dense granular material (ribosomal complexes) in their cytoplasm, Binding of H3-colchicine by the in vivo crystals shows that they contain microtubular proteins. Association of ribosomal complexes with the crystals suggests that these structures may be interrelated.In the present study cultured human leukemic lymphoblasts (CCRF-CEM), were incubated with protein and RNA-synthesis inhibitors, p. fluorophenylalanine, puromycin, cycloheximide or actinomycin-D before the addition of crystal-inducing doses of vinblastine to the culture medium. None of these compounds could completely prevent the formation of the ribosomal complexes or the crystals. However, in cells pre-incubated with puromycin, cycloheximide, or actinomycin-D, a reduction in the number and size of the ribosomal complexes was seen. Large helical polyribosomes were absent in the ribosomal complexes of cells treated with puromycin, while in cells exposed to cycloheximide, there was an apparent reduction in the number of ribosomes associated with the ribosomal complexes (Fig. 2).

Targeting RNA Helicase DHX33 Blocks Ras Driven Lung Tumorigenesis in vivo

2019 ◽  
Author(s):  
Xingshun Wang ◽  
Weimin Feng ◽  
Cheng Peng ◽  
Shiyun Chen ◽  
Kin Yip Tam ◽  
...  
Keyword(s):  
Rna Helicase ◽  
Lung Tumorigenesis
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