Identification and characterization of the NTPase activity of classical swine fever virus (CSFV) nonstructural protein 3 (NS3) expressed in bacteria

Identification and characterization of the NTPase activity of classical swine fever virus (CSFV)... The nonstructural protein 3 (NS3) of members of the family Flaviviridae possesses multiple enzyme activities that are likely to be essential for viral replication. Here, we cloned and expressed full-length CSFV NS3 protein (NS3FL) and its N-terminal truncated version (ntNS3) in E. coli . NTPase activities of the purified NS3FL and ntNS3 proteins and their reaction conditions were investigated. The results showed that CSFV NS3FL and ntNS3 proteins contained a specific polynucleotide-stimulated NTPase acitivity. Characterization of ntNS3 NTPase activity showed that optimal reaction conditions with respect to pH, MgCl 2 and monovalent cations were similar to those of bovine viral diarrhea virus (BVDV) and hepatitis C virus (HCV). Site-directed mutagenesis analysis demonstrated that the GxG K 232 T to GxG A T mutation in the conserved motif I abolished the NTPase activity of ntNS3, whereas substitution of TATP A 354 for TATP V in the motif III had no effect on the enzyme activity. Moreover, the kinetic properties ( K m and k cat ) of CSFV NS3 were more similar to those of BVDV. Our results provide insight into the structure-function relationship of CSFV NS3 and facilitate our understanding of its role in the replication cycle of CSFV. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Archives of Virology Springer Journals

Identification and characterization of the NTPase activity of classical swine fever virus (CSFV) nonstructural protein 3 (NS3) expressed in bacteria

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Publisher
Springer-Verlag
Copyright
Copyright © 2007 by Springer-Verlag
Subject
Biomedicine; Virology; Medical Microbiology; Infectious Diseases
ISSN
0304-8608
eISSN
1432-8798
D.O.I.
10.1007/s00705-007-0969-2
Publisher site
See Article on Publisher Site

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