DOI: 10.1128/jvi.01347-06. Cryoelectron Microscopy Structures of Rotavirus NSP2-NSP5 and NSP2-RNA Complexes: Implications for Genome Replication

Cryoelectron Microscopy Structures of Rotavirus NSP2-NSP5 and NSP2-RNA Complexes: Implications for Genome Replication

10.1128/jvi.01347-06

Crossref journal-article

American Society for Microbiology

Journal of Virology (235)

Abstract

ABSTRACT The replication and packaging of the rotavirus genome, comprising 11 segments of double-stranded RNA, take place in specialized compartments called viroplasms, which are formed during infection and involve a coordinated interplay of multiple components. Two rotavirus nonstructural proteins, NSP2 (with nucleoside triphosphatase, single-stranded RNA [ssRNA] binding and helix-destabilizing activities) and NSP5, are essential in these events. Previous structural analysis of NSP2 showed that it is an octamer in crystals, obeying 4-2-2 crystal symmetry, with a large 35-Å central hole along the fourfold axis and deep grooves at one of the twofold axes. To ascertain that the solution structure of NSP2 is the same as that in the crystals and investigate how NSP2 interacts with NSP5 and RNA, we carried out single-particle cryoelectron microscopy (cryo-EM) analysis of NSP2 alone and in complexes with NSP5 and ssRNA at subnanometer resolution. Because full-length NSP5 caused severe aggregation upon mixing with NSP2, the deletion construct NSP5 66-188 was used in these studies. Our studies show that the solution structure of NSP2 is same as the crystallographic octamer and that both NSP5 66-188 and ssRNA bind to the grooves in the octamer, which are lined by positively charged residues. The fitting of the NSP2 crystal structure to cryo-EM reconstructions of the complexes indicates that, in contrast to the binding of NSP5 66-188 , the binding of RNA induces noticeable conformational changes in the NSP2 octamer. Consistent with the observation that both NSP5 and RNA share the same binding site on the NSP2 octamer, filter binding assays showed that NSP5 competes with ssRNA binding, indicating that one of the functions of NSP5 is to regulate NSP2-RNA interactions during genome replication.

Bibliography

Jiang, X., Jayaram, H., Kumar, M., Ludtke, S. J., Estes, M. K., & Prasad, B. V. V. (2006). Cryoelectron Microscopy Structures of Rotavirus NSP2-NSP5 and NSP2-RNA Complexes: Implications for Genome Replication. Journal of Virology, 80(21), 10829â10835.

Authors 6

Xiaofang Jiang (first)
Hariharan Jayaram (additional)
Mukesh Kumar (additional)
Steven J. Ludtke (additional)
Mary K. Estes (additional)
B. V. Venkataram Prasad (additional)

References 40 Referenced 71

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Dates

Type	When
Created	18 years, 10 months ago (Oct. 15, 2006, 2:42 a.m.)
Deposited	3 years, 5 months ago (March 5, 2022, 5:34 a.m.)
Indexed	4 weeks, 2 days ago (July 25, 2025, 6:27 a.m.)
Issued	18 years, 9 months ago (Nov. 1, 2006)
Published	18 years, 9 months ago (Nov. 1, 2006)
Published Print	18 years, 9 months ago (Nov. 1, 2006)

Funders 0

None

BibTeX

@article{Jiang_2006, title={Cryoelectron Microscopy Structures of Rotavirus NSP2-NSP5 and NSP2-RNA Complexes: Implications for Genome Replication}, volume={80}, ISSN={1098-5514}, url={http://dx.doi.org/10.1128/jvi.01347-06}, DOI={10.1128/jvi.01347-06}, number={21}, journal={Journal of Virology}, publisher={American Society for Microbiology}, author={Jiang, Xiaofang and Jayaram, Hariharan and Kumar, Mukesh and Ludtke, Steven J. and Estes, Mary K. and Prasad, B. V. Venkataram}, year={2006}, month=nov, pages={10829–10835} }

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