Elsevier

Virology

Volume 367, Issue 1, 10 October 2007, Pages 41-50
Virology

Mutational analysis of a helicase motif-based RNA 5′-triphosphatase/NTPase from bamboo mosaic virus

https://doi.org/10.1016/j.virol.2007.05.013Get rights and content
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Abstract

The helicase-like domain of BaMV replicase possesses NTPase and RNA 5′-triphosphatase activities. In this study, mutational effects of the helicase signature motifs and residue L543 on the two activities were investigated. Either activity was inactivated by K643A-S644A, D702A, D730A, R855A, or L543P mutations. On the other hand, Q826A, D858A and L543A had activities, in terms of kcat/Km, reduced by 5- to 15-fold. AMPPNP, a nonhydrolyzable ATP analogue, competitively inhibited RNA 5′-triphosphatase activity. Analogies of mutational effects on the two activities and approximation of Ki(AMPPNP) and Km(ATP) suggest that the catalytic sites of the activities are overlapped. Mutational effects on the viral accumulation in Chenopodium quinoa indicated that the activities manifested by the domain are required for BaMV survival. Results also suggest that Q826 in motif V plays an additional role in preventing tight binding to ATP, which would otherwise decrease further RNA 5′-triphosphatase, leading to demise of the virus in plant.

Keywords

RNA 5′-triphosphatase
NTPase
ATPase
Helicase-like protein
Bamboo mosaic virus
Potexvirus
Alphavirus-like superfamily

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