Cloning, expression and analysis of the interaction of the M2-1 protein of HRSV with Flavonides: investigation for targets in blocking viral replication

Abstract

Human Respiratory Syncytial Virus (hRSV) is the major causative agent of acute respiratory infections (ARIs) such as pneumonia and bronchiolitis. This is a virus of the Paramyxoviridae family with helical symmetry and lipid envelope. It has a single strand RNA genome, not segmented, with 10 coding genes. One of the factors that contribute to success in viral replication is the interaction M2-1 protein with P, M protein and RNA viral. This protein may act inhibiting or neutralizing various steps of the pathway of interferons or to assist in silencing ribonucleoprotein complex (RNP) of RSV. Thus a linkage that render interaction with M2-1 is a potential candidate to prevent interaction of M2-1 with other proteins, thereby preventing the success of viral replication. The aim of the research is to clone, express and purify the M2-1 protein of hRSV and then determine physicochemical characteristics of the interaction with flavonoids in temperature different to investigate the interaction of a potential candidate for inhibiting the activity of the M2-1. The M2-1 gene will be cloned in vector pET28A and expressed in Escherichia coli strain BL21. The expressed protein will be purified on affinity chromatography, thermal stability and interaction with flavonoids will be analyzed by spectroscopy. The results of this study will be unveil protein stability conditions and the interaction site of small ligands, allowing to obtain interaction model, which could lead to potent therapeutic models. As a long term scope, the protein structure determination will be used for refined experiments that will provide better understanding of the flavonoids binding sites. (AU)