Hepatitis C virus polyprotein processing: kinetics and mutagenic analysis of serine proteinase-dependent cleavage.
Hepatitis C virus (HCV) serine proteinase (Cpro-2) is responsible for the processing of HCV nonstructural (NS) protein processing. To clarify the mechanism of Cpro-2-dependent processing, pulse-chase and mutation analyses were performed by using a transient protein production system in cultured cells. Pulse-chase study revealed the sequential production of HCV-NS proteins. Production of p70(NS3) and p66(NS5B) were rapid. An 89-kDa processing intermediate protein (p89) was observed during the early part of the chase. p89 seemed to be cleaved first into a 31-kDa protein (p31) and a p58/56(NS5A). p31 was further processed into p4(NS4A) and p27(NS4B). Mutation analysis of cleavage sites of NS4A/4B, NS4B/5A, and NS5A/5B revealed that cleavage at each site is essentially independent from cleavage occurring at the other site.
Virology Division, National Cancer Center Research Institute, Tokyo, Japan.
Y
Y
Tanji
M
M
Hijikata
Y
Y
Hirowatari
K
K
Shimotohno
D000818:Animals; D002460:Cell Line; D002522:Chlorocebus aethiops; D004591:Electrophoresis, Polyacrylamide Gel; D016174:Hepacivirus / Q000235:genetics / Q000378:metabolism*; D007668:Kidney; D007700:Kinetics; D008715:Methionine / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D016366:Open Reading Frames; D011499:Protein Processing, Post-Translational; D012697:Serine Endopeptidases / Q000378:metabolism*; D013997:Time Factors; D014162:Transfection; D017361:Viral Nonstructural Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*
D017361:Viral Nonstructural Proteins; D008715:Methionine; D012697:Serine Endopeptidases