Hyperphosphorylation of the Hepatitis C Virus NS5A Protein Requires an Active NS3 Protease, NS4A, NS4B, and NS5A Encoded on the Same Polyprotein
AUTOR(ES)
Neddermann, Petra
FONTE
American Society for Microbiology
RESUMO
The nonstructural protein NS5A of hepatitis c virus (HCV) has been demonstrated to be a phosphoprotein with an apparent molecular mass of 56 kDa. In the presence of other viral proteins, p56 is converted into a slower-migrating form of NS5A (p58) by additional phosphorylation events. In this report, we show that the presence of NS3, NS4A, and NS4B together with NS5A is necessary and sufficient for the generation of the hyperphosphorylated form of NS5A (p58) and that all proteins must be encoded on the same polyprotein (in cis). Kinetic studies of NS5A synthesis and pulse-chase experiments demonstrate that fully processed NS5A is the substrate for the formation of p58 and that p56 is converted to p58. To investigate the role of NS3 in NS5A hyperphosphorylation, point and deletion mutations were introduced into NS3 in the context of a polyprotein containing the proteins from NS3 to NS5A. Mutation of the catalytic serine residue into alanine abolished protease activity of NS3 and resulted in total inhibition of NS5A hyperphosphorylation, even if polyprotein processing was allowed by addition of NS3 and NS4A in trans. The same result was obtained by deletion of the first 10 or 28 N-terminal amino acids of NS3, which are known to be important for the formation of a stable complex between NS3 and its cofactor NS4A. These data suggest that the formation of p58 is closely connected to HCV polyprotein processing events. Additional data obtained with NS3 containing the 34 C-terminal residues of NS2 provide evidence that in addition to NS3 protease activity the authentic N-terminal sequence is required for NS5A hyperphosphorylation.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=113049Documentos Relacionados
- Modulation of Hepatitis C Virus NS5A Hyperphosphorylation by Nonstructural Proteins NS3, NS4A, and NS4B
- Specificity of the hepatitis C virus NS3 serine protease: effects of substitutions at the 3/4A, 4A/4B, 4B/5A, and 5A/5B cleavage sites on polyprotein processing.
- The hepatitis C virus NS4A protein: interactions with the NS4B and NS5A proteins.
- Identification of the sequence on NS4A required for enhanced cleavage of the NS5A/5B site by hepatitis C virus NS3 protease.
- NS3 is a serine protease required for processing of hepatitis C virus polyprotein.