NS2 VS NS3 PATCH
The regulation of NS2/3 cleavage could constitute a novel mechanism of switching between viral RNA replication and other processes of the hepatitis C virus life cycle. A conserved NS3 surface patch orchestrates NS2 protease stimulation, NS5A hyperphosphorylation and HCV genome replication. We therefore propose that NS2/3 processing is a critical step in the viral life cycle and is required to permit the accumulation of sufficient NS3 for RNA replication to occur. Finally, we demonstrate that uncleaved NS2/3 degradation can be prevented by the addition of a proteasome inhibitor. Molecular Docking DENV has four serotypes 12 but the binding site of NS2B- Acknowledgment: NS3 has same substrate specificities thus, any inhibitor against The authors would like to acknowledge Government College the binding pocket of NS2/NS3 protease could work against University, Faisalabad (GCUF), 38000, Punjab, Pakistan. 2.NS3 is not backward compatible with NS2 its built from the scratch to replace NS2. Notably, we show that fusion with NS2 leads to the rapid degradation of NS3, whose activity is essential for RNA replication. 1.NS2 is used for wired and wireless simulation wheras ns3 is used for internet simulation. Interestingly, NS3 was still capable of efficient processing of the polyprotein expressed from a subgenomic replicon in Huh-7 cells in the presence of uncleaved NS2. This subsequently resulted in reduced kinetics in an in vitro NS3 protease assay with the unprocessed NS2/3 protein. Unprocessed NS2 had no significant effect on the in vitro ATPase and helicase activities of NS3, whereas immunoprecipitation experiments demonstrated a decreased affinity of NS4A for uncleaved NS2/3 as compared with NS3.
The effect of uncleaved NS2 on the various activities of NS3 was therefore explored. We show here that mutation of three highly conserved residues in NS2 (His 952, Glu 972, and Cys 993) abrogates NS2/3 protease activity and that introduction of any of these mutations into subgenomic NS2-5B replicons results in complete inactivation of NS2/3 processing and RNA replication in both stable and transient replication assays. The hepatitis C virus NS2/3 protease is responsible for cleavage of the viral polyprotein between nonstructural proteins NS2 and NS3.