Binding mode of the activity-modulating C-terminal segment of NS2B to NS3 in the dengue virus NS2B-NS3 protease

Laura de la Cruz, Wan-Na Chen, Bimbil Graham, Gottfried Otting

Research output: Contribution to journalArticleResearchpeer-review

28 Citations (Scopus)

Abstract

The two-component dengue virus NS2B-NS3 protease (NS2B-NS3pro) is an established drug target but inhibitor design is hampered by uncertainties about its 3D structure in solution. Crystal structures reported very different conformations for the functionally important C-terminal segment of the NS2B cofactor (NS2Bc), indicating open and closed conformations in the absence and presence of inhibitors, respectively. An earlier NMR study in solution indicated that a closed state is the preferred conformation in the absence of an artificial linker engineered between NS2B and NS3pro. To obtain direct structural information on the fold of unlinked NS2B-NS3pro in solution, we tagged NS3pro with paramagnetic tags and measured pseudocontact shifts by NMR to position NS2Bc relative to NS3pro. NS2Bc was found to bind to NS3pro in the same way as reported in a previously published model and crystal structure of the closed state. The structure is destabilized, however, by high ionic strength and basic pH, showing the importance of electrostatic forces to tie NS2Bc to NS3pro. Narrow NMR signals previously thought to represent the open state are associated with protein degradation. In conclusion, the closed conformation of the NS2B-NS3 protease is the best model for structure-guided drug design.
Original languageEnglish
Pages (from-to)1517 - 1533
Number of pages17
JournalFEBS Journal
Volume281
Issue number6
DOIs
Publication statusPublished - 2014

Cite this

de la Cruz, Laura ; Chen, Wan-Na ; Graham, Bimbil ; Otting, Gottfried. / Binding mode of the activity-modulating C-terminal segment of NS2B to NS3 in the dengue virus NS2B-NS3 protease. In: FEBS Journal. 2014 ; Vol. 281, No. 6. pp. 1517 - 1533.
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title = "Binding mode of the activity-modulating C-terminal segment of NS2B to NS3 in the dengue virus NS2B-NS3 protease",
abstract = "The two-component dengue virus NS2B-NS3 protease (NS2B-NS3pro) is an established drug target but inhibitor design is hampered by uncertainties about its 3D structure in solution. Crystal structures reported very different conformations for the functionally important C-terminal segment of the NS2B cofactor (NS2Bc), indicating open and closed conformations in the absence and presence of inhibitors, respectively. An earlier NMR study in solution indicated that a closed state is the preferred conformation in the absence of an artificial linker engineered between NS2B and NS3pro. To obtain direct structural information on the fold of unlinked NS2B-NS3pro in solution, we tagged NS3pro with paramagnetic tags and measured pseudocontact shifts by NMR to position NS2Bc relative to NS3pro. NS2Bc was found to bind to NS3pro in the same way as reported in a previously published model and crystal structure of the closed state. The structure is destabilized, however, by high ionic strength and basic pH, showing the importance of electrostatic forces to tie NS2Bc to NS3pro. Narrow NMR signals previously thought to represent the open state are associated with protein degradation. In conclusion, the closed conformation of the NS2B-NS3 protease is the best model for structure-guided drug design.",
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Binding mode of the activity-modulating C-terminal segment of NS2B to NS3 in the dengue virus NS2B-NS3 protease. / de la Cruz, Laura; Chen, Wan-Na; Graham, Bimbil; Otting, Gottfried.

In: FEBS Journal, Vol. 281, No. 6, 2014, p. 1517 - 1533.

Research output: Contribution to journalArticleResearchpeer-review

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