Recombinant Dengue virus protein NS2B alters membrane permeability in different membrane models
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RESEARCH
Open Access
Recombinant Dengue virus protein NS2B alters membrane permeability in different membrane models Moisés León-Juárez1,3, Macario Martínez-Castillo1, Gaurav Shrivastava1, Julio García-Cordero1, Nicolás Villegas-Sepulveda1, Mónica Mondragón-Castelán2, Ricardo Mondragón-Flores2 and Leticia Cedillo-Barrón1*
Abstract Background: One of the main phenomena occurring in cellular membranes during virus infection is a change in membrane permeability. It has been observed that numerous viral proteins can oligomerize and form structures known as viroporins that alter the permeability of membranes. Previous findings have identified such proteins in cells infected with Japanese encephalitis virus (JEV), a member of the same family that Dengue virus (DENV) belongs to (Flaviviridae). In the present work, we investigated whether the small hydrophobic DENV protein NS2B serves a viroporin function. Methods: We cloned the DENV NS2B sequence and expressed it in a bacterial expression system. Subsequently, we evaluated the effect of DENV NS2B on membranes when NS2B was overexpressed, measured bacterial growth restriction, and evaluated changes of permeability to hygromycin. The NS2B protein was purified by affinity chromatography, and crosslinking assays were performed to determine the presence of oligomers. Hemolysis assays and transmission electron microscopy were performed to identify structures involved in permeability changes. Results: The DENV-2 NS2B protein showed similitude with the JEV viroporin. The DENV-2 NS2B protein possessed the ability to change the membrane permeability in bacteria, to restrict bacterial cell growth, and to enable membrane permeability to hygromycin B. The NS2B protein formed trimers that could participate in cell lysis and generate organized structures on eukaryotes membranes. Conclusions: Our data suggest that the DENV-2 NS2B viral protein is capable of oligomerizing and organizing to form pore-like structures in different lipid environments, thereby modifying the permeability of cell membranes. Keywords: Dengue virus, NS2B, Membrane permeability, Virus, Viroporin, Oligomerize, Flavivirus
Background Dengue viruses (DENVs) are enveloped (+) sense RNA viruses belonging to the Flaviviridae family. The DENV replication cycle initiates following receptor binding to the host cell membrane, which is followed by internalization via endocytosis and subsequent release into the cytoplasm. The genome is then translated into a large polyprotein, which is proteolytically processed to yield 3 * Correspondence: [email protected] 1 Departmento de Biomedicina Molecular, Centro de Investigacion y Estudios avanzados IPN, Av. Instituto Politecnico 2508 Col. San Pedro Zacatenco, 07360 México, Mexico Full list of author information is available at the end of the article
structural proteins (envelope, membrane precursor, and capsid) and 7 non-structural (NS) proteins (NS1, NS2A, NS2B, NS3, NS4A, NS4B, and NS5) [1, 2]. Cells infected with DENV undergo a series of detrimental functional and structural
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