Expression and purification of untagged full-length HCV NS5B RNA-dependent RNA polymerase.
Protein Expr Purif
; 35(2): 304-12, 2004 Jun.
Article
em En
| MEDLINE
| ID: mdl-15135407
ABSTRACT
The NS5B encoded by the hepatitis C virus genome is a RNA-dependent RNA polymerase essential to viral replication. The entire NS5B protein contains a catalytic domain followed by a regulatory motif and a membrane-anchor domain at its C-terminus. Reported here is the molecular cloning and expression of the full-length NS5B polymerase (NS5B-FL) in bacterial cells as a non-fusion protein. The non-tagged NS5B-FL was purified to homogeneity using sequential chromatographic columns and its identity was confirmed using anti-NS5B peptide antibodies and amino acid sequencing. Purified NS5B-FL demonstrated RNA-dependent RNA polymerase activity and was able to replicate a HCV RNA genome fragment through both copy-back and de novo mechanisms. Its biochemical properties were further characterized in comparison with a truncated form of NS5B polymerase with a deletion of 51 residues from its C-terminus.
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Base de dados:
MEDLINE
Assunto principal:
Proteínas não Estruturais Virais
Idioma:
En
Ano de publicação:
2004
Tipo de documento:
Article