Studying the Effect of Amino Acid Substitutions in the M2 Ion Channel of the Influenza Virus on the Antiviral Activity of the Aminoadamantane Derivative In Vitro and In Silico.

Timur Mansurovich Garaev,Sergey Vadimovich Rubinsky,Alexei Gennadievich Prilipov,Alexander Aleksandrovich Lashkov,Marina Mikhailovna Zhuravleva,Marina Pavlovna Finogenova,Tatiana Anatol'Evna Timofeeva,Galina Kadymovna Sadykova,Artyom Irorevich Odnovorov,Svetlana Nikolaevna Norkina,Tatiana Vladimirovna Grebennikova

doi:10.34172/apb.2021.079

Timur Mansurovich Garaev, Sergey Vadimovich Rubinsky + Show 9 more

Open Access

https://doi.org/10.34172/apb.2021.079

Copy DOI

Abstract

Purpose: The aminoadamantane derivative of L-histidyl-1-adamantayl ethylamine hydrochloride (HCl*H-His-Rim) has showed a high inhibition level against influenza A virus strains in vitro. The aim of this work is to search and establish evidence of the direct effect of the drug on influenza A virus proton channel M2. Methods: The compound HCl*H-His-Rim was obtained by classical peptide synthesis methods. Influenza A virus mutants of A/PuertoRico/8/34(H1N1) strain were obtained by reverse genetics methods. The mutant samples of the virus were cultured on chicken embryos with a virus titer in the hemagglutination test. ELISA was carried out on Madin-Darby canine kidney (MDCK) monolayer cells when multiplying the virus 10-4-10-6. The binding stability of HCl*H-His-Rim was compared to those of M2 (S31N) and M2 (S31N_A30T) channels by molecular dynamic (MD) modeling. The calculation was performed taking into account the interaction with the model lipid bilayer (1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine) in the presence of water molecules in accordance with the three-center model. Results: It was found that HCl*H-His-Rim is a direct action drug against influenza A. The most likely conformation of drug binding to target protein has been shown. It has been found that the A30T mutation reduces the binding energy of the drug, and the results obtained in vitro have confirmed the data calculated in silico. Conclusion: The mechanism of action of HCl*H-His-Rim is directly related to the suppression of the function of the proton channel M2 of influenza A virus.

Highlights

The M2 proteins of Influenza A virus are transmembrane proteins that form proton-selective channels in the lipid membrane
The function of the ion channels can be blocked by small inhibitor molecules, which leads to a significant decline in the reproduction of viral particles
Before the appearance of high-resolution crystallographic structures, the drug binding site was predicted from the position of mutations that led to the emergence of resistant strains of influenza A virus

Summary

Introduction

The M2 proteins of Influenza A virus are transmembrane proteins that form proton-selective channels in the lipid membrane. Proton-selective M2 channels regulate pH inside the viral particle during virion penetration into the endosome of the cell by acidification the inner space of the viral particle, which results in dissociation of the matrix protein M1 complex with ribonucleotide and leads to the release of genetic material into the cytoplasm of the host cell.[2,3] Another important role of M2 protein appears at a later stage during viral assembly, when newly synthesized viral proteins are transported to the cell membrane surface. At low pH values, the virus proteins, in particular, hemagglutinin (HA), can undergo premature and undesirable transformation; in this case the M2 proton channels pump hydrogen ions from the Golgi complex to maintain a sufficiently high pH value and preserve the transport form of the newly formed HA.[4,5]

Objectives

Methods

Results

Conclusion