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，初步认为该病毒属于黄病毒科。然而，病毒非结构蛋白NS 3对病毒复制的活性和意义尚不清楚。为了阐明这些，提取HCV-NS 3相关蛋白并对其功能进行如下测试：1.制备HCV-NS 3表达载体，其含有5'端或NS 3或3'的（pHCN 3-N'）区域（pHCN 3-C ′）和HCV-NS 3相关蛋白在1 mM IPTG存在下在细菌中表达。在pHCN 3-N'和pHCN 3-C'中分别产生32 kDa和49 kDa的HCN 3-N'和HCN 3-C'蛋白。此外，该蛋白仅与丙型肝炎患者血清呈阳性反应，与健康对照血清无反应.对丙型肝炎患者进行了HCN_3-C'反应性检测，经干扰素治疗后，HCN_3-C'反应性转为阴性。此外，在HCV诱导的肝脏疾病中，反应性随着进展而增加.用该蛋白免疫家兔后获得抗HCN 3-C'的抗血清。对一例慢性丙型肝炎患者的肝组织进行穿刺活检，取肝组织匀浆。抗血清与肝组织之间有明显的免疫反应，约70 kDa的蛋白质存在.通过SDS-PAGE和电洗脱法纯化了HCN 3-C'蛋白。在纯化的蛋白质中，可以检测到明显的ATP酶活性。本研究表明，HCV-NS 3在丙型肝炎的发生中起作用，至少部分。此外，还提出了HCV-NS 3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.