The NTPase/helicase domain of hepatitis C virus nonstructural protein 3 inhibits protein kinase C independently of its NTPase activity

Cellular & Molecular Biology Letters

Philip Hartjen⁵  Peter Borowski¹  Bastian Höchst³  Denise Heim²  Ralf Smeets⁶  Michael Reinholz⁷  Henning Kammer⁷  Henning Wege²  Andrea Baier⁴  Julian Schulze Zur Wiesch²  Judith Lucke⁷ 

[1] Department of Virology, Bernhard-Nocht-Institute for Tropical Medicine, Hamburg, Germany$$Department of Molecular Biology, The John Paul II Catholic University of Lublin, Lublin, Poland$$;I Department of Internal Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, Germany$$;Department of Virology, Bernhard-Nocht-Institute for Tropical Medicine, Hamburg, Germany$$Institutes of Molecular Medicine and Experimental Immunology, University Hospital Bonn, Bonn, Germany$$;Department of Molecular Biology, The John Paul II Catholic University of Lublin, Lublin, Poland$$;Department of Virology, Bernhard-Nocht-Institute for Tropical Medicine, Hamburg, Germany$$I Department of Internal Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, Germany$$Department of Oral and Maxillofacial Surgery, University Medical Center Hamburg-Eppendorf, Hamburg, Germany$$;Department of Oral and Maxillofacial Surgery, University Medical Center Hamburg-Eppendorf, Hamburg, Germany$$;Department of Virology, Bernhard-Nocht-Institute for Tropical Medicine, Hamburg, Germany$$

关键词: Hepatitis C virus (HCV);    NS3 protein;    Protein kinase C (PKC);    PKC isotypes;    Protein kinase inhibitors;    Pseudosubstrate inhibition;    Hepatocellular carcinoma (HCC);   

DOI  :  10.2478/s11658-013-0099-7

学科分类：分子生物学，细胞生物学和基因

来源: Uniwersytet Wroclawski * Wydzial Biotechnologii / University of Wroclaw, Faculty of Biotechnology

【 摘 要 】

Helicase motif VI is a short arginine-rich motif within the NTPase/helicase domain of the non-structural protein 3 (NS3) of the hepatitis C virus (HCV). We previously demonstrated that it reduces the catalytic activity and intracellular shuttling of protein kinase C (PKC). Thus, NS3-mediated PKC inhibition may be involved in HCV-associated hepatocellular carcinoma (HCC). In this study, we expand on our earlier results, which were obtained in experiments with short fragments of NS3, to show for the first time that the catalytically active, longer C-terminal NTPase/helicase of NS3 acts as a potent PKC inhibitor in vitro. PKC inhibition assays with the NTPase-inactive mutant NS3h-D1316A revealed a mixed type kinetic inhibition pattern. A broad range of 11 PKC isotypes was tested and all of the PKC isotypes were inhibited with IC50-values in the low micromolar range. These findings were confirmed for the wild-type NTPase/helicase domain in a non-radiometric PKC inhibition assay with ATP regeneration to rule out any effect of ATP hydrolysis caused by its NTPase activity. PKCα was inhibited with a micromolar IC50 in this assay, which compares well with our result for NS3h-D1316A (IC50 = 0.7 μM). In summary, these results confirm that catalytically active NS3 NTPase/helicase can act in an analogous manner to shorter NS3 fragments as a pseudosubstrate inhibitor of PKC.

【 授权许可】

Unknown