Viral Polymerases

Structures, Functions and Roles as Antiviral Drug Targets

Satya Prakash Gupta

• Publisher: Academic Press
• Year: 2018
• Print ISBN: 9780128154229
• eBook ISBN: 9780128154236
• Topic: Biological Sciences
• Subtopic: Microbiology
• Index: Biological Sciences

Viral Polymerases: Structures, Functions and Roles as Antiviral Drug Targets presents in-depth study information on the structure and functions of polymerases and their roles in the lifecycle of viruses, and as drug targets. Viral polymerases constitute a vital component in the lifecycle of many viruses, such as human immunodeficiency virus (HIV), hepatitis viruses, influenza virus, and several others. They are essentially required for the replication of viruses. Thus, the polymerases that can be found in viruses (called viral polymerases) represent favorable targets for the design and development of antiviral drugs.- Provides comprehensive, state-of-the-art coverage on virus infections, the virus lifecycle, and mechanisms of polymerase inhibition- Analyzes the structure-activity relationships of inhibitors of each viral polymerase- Presents a consistent and comprehensive coverage of all aspects of viral polymerases, including structure, function and their role as antiviral drug targets

Chapter 1

RNA-Dependent RNA Polymerases and Their Emerging Roles in Antiviral Therapy

Ankit Ganeshpurkar, Gopichand Gutti and Sushil Kumar Singh, Department of Pharmaceutical Engineering & Technology, Indian Institute of Technology (Banaras Hindu University), Varanasi, Uttar Pradesh, India

Abstract

Viruses are the causative agents leading to variety of diseases. The viral RdRPs are the attractive targets for antiviral therapy. This chapter presents a collective view on the structure and ligand-based drug design for the inhibition of viral RdRPs. It also highlights the structural aspects of various RdRPs. Interestingly, all these enzymes share structural similarities but certain significantly different structural features may be vital to design their selective inhibitors. The chapter also introduces the readers with different classes of RdRP inhibitors, i.e., nucleoside and nonnucleoside inhibitors involving various scaffolds such as Anthranilic acid, Benzimidazole, Benzothiadiazine Indole, Proline sulfonamide, Pyrrolidine diacids, and Pyridobenzothiazole. This chapter is thus prelude to the remaining chapters in this book, which deal with the viral RdRPs.

Keywords

RNA-dependent RNA polymerase; nucleoside inhibitors; allosteric inhibitors; structure-based drug design

1.1 Introduction

Viruses are pestilential agents, typically made up of one kind of nucleic acid, either RNA or DNA. They attach to the host cell to invade it and replicate inside the cell by utilizing host biochemicals and processes. Among the two types of viruses, RNA and DNA viruses, the former cause a critical burden on world healthcare systems (Cameron et al., 2009; Claverie, 2006). Viruses escape the host immune response by integrating their genome into the host DNA and due to this ability they play the crucial roles of causative agents for plentiful human diseases (Hemmi et al., 2002; Hancioglu et al., 2007; Haseltine et al., 2008; Jensen and Thomsen, 2012; Ahlquist et al., 2003). RNA viruses use well-organized replication and transcription strategies to amplify their genetic material (Díez et al., 2000). The evolutionary dynamics of RNA viruses complicate the management of viral diseases including hepatitis, dengue, yellow fever, Ebola, etc., and the mutation rate in these viruses is perilously equal to the highest tolerable error rate (Hiscox, 2007; Holmes, 2011). RNA viruses have a compact and well-organized genome sequences to maintain mutational robustness (Franco et al., 1995). The theory, “Quasispecies,” explains about the viral population, which is flexible with diverse group of variants and different replicative capacities will emerge with spontaneous mutations after each round of replication. The theory also has been used to describe the evolutionary dynamics of RNA viruses (Lauring and Andino, 2010; Lauring et al., 2013). A few examples of medically important RNA viruses include Zika virus (ZIKV), which spreads by daytime-active Aedes mosquitoes; Ebola virus, which causes deadly hemorrhagic fevers; influenza virus, which causes notorious pandemic “Spanish flu” infection; and hepatitis C virus, a blood-borne virus, which causes both acute and chronic infections. Globally, an estimated 71 million people have chronic hepatitis C infection. Although, several viral drug targets are known and well established, no single agent has been discovered for complete cure of viral infections. Several challenges still exist in the drug discovery cascade of the antiviral. Despite of continuous advances, RNA viruses remain major causes of human disease that costs in mortality, morbidity, and monetary terms (Neumann et al., 2002; Ahlquist, 2006; Cattaneo et al., 1988). The RNA viruses are classified into three different classes, i.e., double-stranded (dsRNA), positive sense single-strand (+ssRNA), and negative sense single-strand (–ssRNA) based on type of genome (Ahlquist, 2002; Baltimore, 1971).

1.2 RNA Polymerase

RNA, a polynucleotide molecule, is one of the major essential constituents of life. It is often present in single-stranded form and performs a variety of functions not limited to only peptide bond formation but can also be involved in replication, intron splicing, translational regulation, and gene regulation. RNAs are of three types, viz. messenger RNA (mRNA), transfer RNA (tRNA), and ribosomal RNA (rRNA). The mRNA conveys information stored in genetic material to direct protein synthesis, rRNA helps to form peptide linkage between amino acids while tRNA helps to supply amino acids to the ribosomes. An RNA molecule having conserved base pair regions, plays many important roles. The most stable helix may have important biological roles. RNA molecules replicate through a class of enzymes termed as RNA polymerases. The RNA polymerases belong to two different classes as described below.

1.2.1 DNA-Dependent RNA Polymerases

The RNA polymerases that catalyze the formation of phosphodiester bonds between ribonucleotides to form an ssRNA polymer using DNA as a template are called as DNA-dependent RNA polymerases (DdRPs). They are found in various life forms including viruses. They uncoil the double-stranded DNA, which acts as a template for the synthesis of RNA.

1.2.2 RNA-Dependent RNA Polymerases

The RNA-dependent RNA polymerases (RdRPs), also called RNA replicases, catalyze the replication of RNA from an RNA template and are essential proteins encoded in the genomes of all RNA-containing viruses with no DNA stage (Poltronieri et al., 2015; Ahlquist, 2006). An RdRP is involved in a pathway outside the “central dogma” of early molecular biology. RdRPs, present in a wide variety of RNA viruses, are involved in genome replication, mRNA synthesis, or RNA recombination, etc. They are essential for the survival of viruses. They also play an important role in evolution when RNA was primary genetic material. The RdRPs have multiple conserved sequences and motifs across the length. They share structural similarity with DNA-dependent DNA polymerase, DdRP, and reverse transcriptase (RT). The higher error rate (in order of 10⁻⁴) in transcription process by RdRPs leads to genomic variations in the RNA virus population. The switching mechanism during RdRPs copying process may lead to RNA recombination and also helps viruses to repair deleterious mutations and acquire new genes and genetic rearrangements.

1.3 Structure of Viral RNA-Dependent RNA Polymerases

All RdRPs share structural resemblance, having fingers, palm, and thumb domains, which provide cupped right-hand-like appearance. The three domains together constitute the binding site for RNA and nucleotide phosphates (NTPs). Although, the low sequence similarity is observed in various polymerases of different families, they have similar structural features and thus indicate a high evolutionary similarity in distinct species. A peculiar N-terminal sequence is found between fingers and thumb domain that appears as a bridge in all RdRPs. Cystoviridae, Flaviviridae, and Reoviridae family of viruses show the presence of C-terminal domain encircling the central cleft of the enzyme. The representative RdRP structure (EV71) has seven motifs or specific regions, i.e., G, F1–3, A, B, C, D, and E from N- to C-termini. Each motif has different sequences, but their folds are conserved and adopt specific conformation, which is similar in different RdRPs (homomorphs). Loops that connect the finger domains are called fingertips and are responsible for overall unique closed-hand conformation of the enzyme. They are not present in other types of polymerases. The fingertip residues pack the major groove of the template RNA and unstack the RNA strand at +3 position. These residues are not conserved but play an important role in holding the template RNA without much conformational changes. The variations in finger subdomain also help in performing functions like nuclear import, oligomerization, phosphorylation, and protein–protein interactions.

The thumb domain residues are involved in RNA binding and surround the minor groove of the template RNA while other residues also contribute to the formation of “NTP tunnel.” The entire domain has α-helices except the N-terminal that bears β-sheet only (Hansen et al., 1997). This region has small sequence protrusions that stabilize template channel and initiate binding of NTPs on the template. These regions have higher conformational flexibility, which facilitates translocation of the template after each NTP addition (condensation reaction). The NTP tunnels are lined by positively charged amino acid residues, though not sufficient, but still provide guidance to NTPs to enter the catalytic site (Fig. 1.1A).

Figure 1.1 (A) Structure of RdRP, representing the active site (EV71); (B) structural representation of major cavities of RdRP (EV71); (C) ribbon structure of EV71 RdRP depicting the various subdomains of RdRPs (PDB–3N6M); and (D) two metal ions mechanism for stabilization of incoming NTP.

The palm domain consists of A, C, and D motifs bearing catalytic site. The three β-antiparallel sheets lined by three α-helices form the core of RdRPs, which is conserved in all the enzymes. The core region also has highly conserved N-terminal aspartate...

Table of contents

• Cover image
• Title page
• Table of Contents
• Copyright
• List of Contributors
• Preface
• Chapter 1. RNA-Dependent RNA Polymerases and Their Emerging Roles in Antiviral Therapy
• Chapter 2. Structure–Function Relationship of Negative-Stranded Viral RNA Polymerases: Prospectives for Antiviral Therapy
• Chapter 3. RNA-Dependent RNA Polymerase of Alphaviruses: A Potential Target for the Design of Drugs Against Alphaviruses
• Chapter 4. DNA-Dependent DNA Polymerases as Drug Targets in Herpesviruses and Poxviruses
• Chapter 5. Poliovirus Polymerase: An Effective Target for Design and Development of Antipolio Drugs
• Chapter 6. Studies on HIV-1 Polymerase and Its Inhibitors
• Chapter 7. A Focus on Ebola Virus Polymerase: Structure, Functions and Antiviral Therapies
• Chapter 8. Hepatitis C Virus NS5B RNA-Dependent RNA Polymerase Inhibitor: An Integral Part of HCV Antiviral Therapy
• Chapter 9. HBV Polymerase as a Target for Development of Anti-HBV Drugs
• Chapter 10. Polymerases of Coronaviruses: Structure, Function, and Inhibitors
• Chapter 11. Rhinovirus RNA Polymerase: Structure, Function, and Inhibitors
• Chapter 12. Herpesvirus Polymerase Inhibitors
• Chapter 13. Zika Virus Polymerase: Structure, Function, and Inhibitors
• Chapter 14. Dengue Virus Polymerase: A Crucial Target for Antiviral Drug Discovery
• Chapter 15. Adenovirus DNA Polymerase: Structure, Function, and Prospects in Diagnostics and Therapeutics
• Index