Biological function of hepatitis C virus nonstructural protein NS3

丙型肝炎病毒（HCV）是慢性丙型肝炎的病原体。利用重组DNA技术克隆了该病毒，推测其属黄病毒科。然而，病毒非结构蛋白NS3在病毒复制中的活性和意义尚不清楚。为此，我们提取了HCV-NS3相关蛋白，并对其功能进行了如下测试：制备含有NS3 5′（pHCN3-N′）区或3′（pHCN3-C′）区的HCV-NS3表达载体，在1mM IPTG存在下在细菌中表达HCV-NS3相关蛋白。当表达的蛋白与人血清反应时，发现在含有pHCN3-N‘的细菌中产生的HCN3-N’蛋白大小为32 kDa， pHCN3-C‘的HCN3-C’大小为49 kDa。此外，该蛋白仅与丙型肝炎患者血清反应阳性，与健康对照组血清无反应。在丙型肝炎患者中检测HCN3-C′的反应性，发现干扰素治疗后HCN3-C′变为阴性。此外，随着hcv诱导的肝脏疾病的进展，反应性增加。用该蛋白免疫家兔获得抗HCN3-C′血清。肝组织均质化，这是采取了针活检患者的慢性丙型肝炎。在抗血清与肝组织之间，发现了一个大小为70kda的蛋白的明显免疫反应。采用SDS-PAGE和电洗脱法纯化HCN3-C′蛋白。在纯化的蛋白中，可以检测到显著的atp酶活性。目前的研究表明，HCV-NS3至少部分地在丙型肝炎的进展中起作用。此外，还提示了HCV-NS3在病毒复制中的意义。

Hepatitis C virus (HCV) is the causative agent of chronic type C hepatitis. It is cloned by the recombinant DNA techniques and suggested to be a member of the family Flaviviridae. However, the activity and significance of virus nonstructural protein NS3 on the replication of virus are not known. To clarify these, HCV-NS3 related proteins were extracted and their function was tested as follows :1. HCV-NS3 expression vectors were prepared containing either 5'(pHCN3-N') region of NS3 or 3'(pHCN3-C') and HCV-NS3 related proteins were expressed in the bacteria in the presence of 1mM IPTG.When the expressed proteins were reacted with human sera, HCN3-N' protein which was produced in the bacteria containing pHCN3-N' was found to be the size of 32 kDa and HCN3-C' from pHCN3-C' was of 49 kDa, respectively. In addition, the reaction was positive only with sera of patients with hepatitis C.No reaction was seen between the extracted proteins and sera of healthy controls.2. The reactivity of HCN3-C' was tested in patients with hepatitis C and found that it became to be negative after the treatment of the patients with interferon. In addition, the reactivity was increased as progressed in the HCV-induced liver disease.3. Antisera against HCN3-C' were obtained following the immunization of rabbits with the protein. Liver tissue, which was taken by needle biopsy of a patient with chronic type C hepatitis, was homogenized. Between the antisera and the liver tissue, a clear immunoreaction was found in a size of 70 kDa protein.4. The protein HCN3-C' was purified by SDS-PAGE and the electro-elution method. In the purified protein, it was possible to detect a remarkable activity of ATPase.The present study suggests that HCV-NS3 plays, at least in part, a role in the progession of type C hepatitis. In addition, the significance of HCV-NS3 in the replication of virus is suggested.