Hepatitis C Virus (HCV) is a significant global health concern, infecting millions of people worldwide. It is a positive, single-stranded RNA virus belonging to the Flaviviridae family. The HCV genome encodes a single polyprotein of approximately 3,000 amino acids, which is processed by host and viral proteases into structural and non-structural proteins. Among these, the non-structural protein 3 (NS3) is particularly noteworthy due to its multifunctional enzymatic activities and its role in viral replication and pathogenesis.
The NS3 protein of HCV is a multifunctional enzyme with three distinct activities: serine protease, NTPase, and RNA helicase. The serine protease activity of NS3 is responsible for the proteolytic processing of the HCV polyprotein, which is essential for the maturation of other non-structural proteins, including NS4A, NS4B, NS5A, and NS5B . The NTPase and RNA helicase activities are crucial for viral RNA replication.
The NS3 protein spans amino acids 1027 to 1657 of the HCV polyprotein. The recombinant fragment of NS3, encompassing amino acids 1450 to 1643, retains the essential enzymatic activities and is often used in research to study the protein’s function and to develop antiviral drugs .
The recombinant NS3 protein (1450-1643 a.a.) is typically expressed in Escherichia coli (E. coli) systems. The gene encoding the NS3 fragment is cloned into an expression vector, which is then introduced into E. coli cells. The bacteria are cultured, and the expression of the recombinant protein is induced. Following induction, the cells are lysed, and the NS3 protein is purified using chromatographic techniques to achieve high purity levels, often exceeding 95% .
The recombinant NS3 protein is widely used in various research applications, including: