| Budget Amount *help |
¥1,000,000 (Direct Cost: ¥1,000,000)
Fiscal Year 1993: ¥1,000,000 (Direct Cost: ¥1,000,000)
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| Research Abstract |
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.
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