What question did this study set out to answer?

To determine whether the Semliki Forest virus replicase protein nsP2 functions as an active RNA helicase capable of unwinding duplex RNA.

April 1, 1999FEBS Letters

RNA helicase activity of Semliki Forest virus replicase protein NSP2

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Authors

MCMarta Gómez de CedrónMadrid Institute for Advanced Studies NENeda EhsaniUniversity of Helsinki MMMarja L. MikkolaUniversity of Helsinki

Discussion

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Implication

In vitro assays demonstrate RNA helicase activity in Semliki Forest virus replicase protein nsP2, indicating an essential enzymatic role for viral replication in alphaviruses.

Key Points

To determine whether the Semliki Forest virus replicase protein nsP2 functions as an active RNA helicase capable of unwinding duplex RNA.
Expressed recombinant polyhistidine-tagged nsP2 in Escherichia coli and purified the protein using metal-affinity chromatography.
Conducted RNA helicase assays measuring unwinding of alpha-32P-labelled partially double-stranded RNA substrates in the presence of NTPs, Mg2+, and a K192N NTP-binding site mutant.
Purified nsP2 successfully unwound partially double-stranded RNA in a reaction strictly dependent on Mg2+ and NTP cofactors.
Introduction of the K192N mutation in the GVPGSGK192SA motif completely abolished both duplex RNA unwinding and NTPase activity.

Cite This Study

Cedrón et al. (1999) studied this question.

synapsesocial.com/papers/6a72dbccc4b6fb1e3af3ce7f https://doi.org/10.1016/s0014-5793(99)00321-x

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April 1, 1999FEBS Letters

RNA helicase activity of Semliki Forest virus replicase protein NSP2

View Full Paper

Ask AI

Bookmark

Authors

MCMarta Gómez de CedrónMadrid Institute for Advanced Studies NENeda EhsaniUniversity of Helsinki MMMarja L. MikkolaUniversity of Helsinki

Discussion

Member takes

Implication

In vitro assays demonstrate RNA helicase activity in Semliki Forest virus replicase protein nsP2, indicating an essential enzymatic role for viral replication in alphaviruses.

Key Points

To determine whether the Semliki Forest virus replicase protein nsP2 functions as an active RNA helicase capable of unwinding duplex RNA.
Expressed recombinant polyhistidine-tagged nsP2 in Escherichia coli and purified the protein using metal-affinity chromatography.
Conducted RNA helicase assays measuring unwinding of alpha-32P-labelled partially double-stranded RNA substrates in the presence of NTPs, Mg2+, and a K192N NTP-binding site mutant.
Purified nsP2 successfully unwound partially double-stranded RNA in a reaction strictly dependent on Mg2+ and NTP cofactors.
Introduction of the K192N mutation in the GVPGSGK192SA motif completely abolished both duplex RNA unwinding and NTPase activity.

Cite This Study

Cedrón et al. (1999) studied this question.

synapsesocial.com/papers/6a72dbccc4b6fb1e3af3ce7f https://doi.org/10.1016/s0014-5793(99)00321-x

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