Structure-based mutagenesis identifies important novel determinants of the NS2B cofactor of the West Nile Virus two-component NS2B-NS3 proteinase

Inflammatory and Infectious Disease Center, Burnham Institute for Medical Research, La Jolla, CA 92037, USA.
Journal of General Virology (Impact Factor: 3.18). 04/2008; 89(Pt 3):636-41. DOI: 10.1099/vir.0.83359-0
Source: PubMed


West Nile virus (WNV) is an emerging mosquito-borne flavivirus that causes neuronal damage in the absence of treatment. In many flaviviruses, including WNV, the NS2B cofactor promotes the productive folding and the functional activity of the two-component NS3 (pro)teinase. Based on an analysis of the NS2B-NS3pro structure, we hypothesized that the G(22) residue and the negatively charged patch D(32)DD(34) of NS2B were part of an important configuration required for NS2B-NS3pro activity. Our experimental data confirmed that G(22) and D(32)DD(34) substitution for S and AAA, respectively, inactivated NS2B-NS3pro. An additional D42G mutant, which we designed as a control, had no dramatic effect on either the catalytic activity or self-proteolysis of NS2B-NS3pro. Because of the significant level of homology in flaviviral NS2B-NS3pro, our results will be useful for the development of specific allosteric inhibitors designed to interfere with the productive interactions of NS2B with NS3pro.

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    • "Processing of the polyprotein precursor by the NS2B/NS3 protease is essential in the flavivirus life cycle (Chambers et al., 1990b), and as such, it has been one of the prime antiviral targets for WNV therapy. Typically, in vitro substrate assays are used to assess WNV NS2B/NS3 protease activity using the truncated bacterial-expressed recombinant form of the protease, NS2B 40 -G 4 SG 4 -NS3pro (Nall et al., 2004; Chappell et al., 2005, 2006, 2007, 2008a,b; Shiryaev et al., 2006, 2007a,b,c; Bera et al., 2007; Radichev et al., 2008). Nall et al. (2004) investigated the enzymatic characterization of the WNV recombinant NS2B 40 -G 4 SG 4 -NS3pro protease in vitro. "
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