Analysis of Immumoreactivity of Heterologously Expressed Non-structural Protein 4B (NS4B) from Hepatitis C Virus (HCV) Genotype 1a

Authors

  • Amir Savardashtaki Department of Medical Biotechnology, School of Allied Medical Sciences, Iran University of Medical Sciences, Tehran, Iran
  • Mohammad M Farajollahi Department of Medical Biotechnology, School of Allied Medical Sciences, Iran University of Medical Sciences, Tehran, Iran
  • Sepideh Hamzehlou Department of Medical Biotechnology, School of Allied Medical Sciences, Iran University of Medical Sciences, Tehran, Iran
  • Zohreh Sharifi Department of Virology, Iranian blood Transfusion Organization, Tehran, Iran.
Abstract:

Background: Detection of hepatitis C virus specific antibodies is the initial step in chronic HCV diagnosis.  HCV NS4B is among the most immunogenic HCV antigens and has been widely used in commercial Enzyme Immunoassays (EIA). Additionally, NS4B, a key protein in the virus replication, can be an alternative target for antiviral therapy. Objectives: Development of a new method for high-level expression and purification of NS4B coding region was the aim of the report. Materials and Methods: Viral RNA was purified from the serum of an HCV positive patient and NS4B coding region was amplified using nested RT-PCR. PCR products were cloned into pET102/D-TOPO expression vector and transformed into E. coli BL21. Induction was performed by adding 1mM isopropyl-b-D-thiogalactopyranoside (IPTG) to the culture medium. Immunoreactivity of the purified recombinant proteins was evaluated by immunoblotting and indirect enzyme-linked immunosorbent assay (ELISA). Results: The recombinant NS4B protein was expressed and its immunoreactivity was confirmed by ELISA and western blotting. Conclusions: The directional TOPO cloning provides an efficient and easy platform for heterologous expression of immunoreactiveHCV NS4B.

Upgrade to premium to download articles

Sign up to access the full text

Already have an account?login

similar resources

analysis of immumoreactivity of heterologously expressed non-structural protein 4b (ns4b) from hepatitis c virus (hcv) genotype 1a

background: detection of hepatitis c virus specific antibodies is the initial step in chronic hcv diagnosis.  hcv ns4b is among the most immunogenic hcv antigens and has been widely used in commercial enzyme immunoassays (eia). additionally, ns4b, a key protein in the virus replication, can be an alternative target for antiviral therapy.objectives: development of a new method for high-level exp...

full text

Gene expression profiles of HeLa Cells impacted by hepatitis C virus non-structural protein NS4B.

By a cDNA array representing 2308 signal transduction-related genes, we studied the expression profiles of HeLa cells stably transfected by Hepatitis C virus nonstructural protein 4B (HCV-NS4B). The alterations of the expression of four genes were confirmed by real-time quantitative RTPCR; and the aldo-keto reductase family 1, member C1 (AKR1C1) enzyme activity was detected in HCV-NS4B transien...

full text

Interferon Resistance of Hepatitis C Virus Genotypes 1a/1b: Relationship to Structural E2 Gene Quasispecies Mutations

Hepatitis C virus (HCV) envelope glycoprotein-2 (E2) inhibits the interferon (IFN)–induced, double –stranded RNA activated protein kinase (PKR) via PKR eukaryotic initiation factor-2α phosphorylation homology domain (PePHD). Present study examined the genetic variability of the PePHD in patients receiving interferon therapy. The PePHD region from HCV genotype 1a/1b infected patients receiving I...

full text

Screening of hepatitis C virus inhibitors using genotype 1a HCV replicon cell lines.

Hepatitis C virus (HCV) replicons are the primary tools for identifying and evaluating anti-HCV compounds during drug discovery research. Genotype 1a and 1b are the most common genotypes in North America and Europe. These genotypes display significant genetic divergence (∼20% at the nucleotide level and ∼14% at the amino acid level), which can translate into marked differences in drug susceptib...

full text

Isolation and Characterization of Highly Replicable Hepatitis C Virus Genotype 1a Strain HCV-RMT

Multiple genotype 1a clones have been reported, including the very first hepatitis C virus (HCV) clone called H77. The replication ability of some of these clones has been confirmed in vitro and in vivo, although this ability is somehow compromised. We now report a newly isolated genotype 1a clone, designated HCV-RMT, which has the ability to replicate efficiently in patients, chimeric mice wit...

full text

construction and immunogenicity analysis of hepatitis c virus (hcv) truncated non-structural protein 3 (ns3) plasmid vaccine

conclusions the pc-ns3 possesses the capacity to express ns3 in the mammalian cell line and demonstrated strong immunogenicity in a murine model. our primary results demonstrated that the immunogenic truncated region of ns3 could be used as a potential vaccine candidate against hepatitis c. results the pcdna3.1 plasmid harboring the coding sequence of ns3 (pc-ns3) was constructed and confirmed ...

full text

My Resources

Save resource for easier access later

Save to my library Already added to my library

{@ msg_add @}


Journal title

volume 13  issue 4

pages  32- 37

publication date 2015-12-01

By following a journal you will be notified via email when a new issue of this journal is published.

Hosted on Doprax cloud platform doprax.com

copyright © 2015-2023