The crystal structure and RNA-binding of an orthomyxovirus nucleoprotein.
Genome packaging for viruses with segmented genomes is often a complex problem. This is particularly true for influenza viruses and other orthomyxoviruses, whose genome consists of multiple negative-sense RNAs encapsidated as ribonucleoprotein (RNP) complexes. To better understand the structural features of orthomyxovirus RNPs that allow them to be packaged, we determined the crystal structure of the nucleoprotein (NP) of a fish orthomyxovirus, the infectious salmon anemia virus (ISAV) (genus Isavirus). As the major protein component of the RNPs, ISAV-NP possesses a bi-lobular structure similar to the influenza virus NP. Because both RNA-free and RNA-bound ISAV NP forms stable dimers in solution, we were able to measure the NP RNA binding affinity as well as the stoichiometry using recombinant proteins and synthetic oligos. Our RNA binding analysis revealed that each ISAV-NP binds ~12 nts of RNA, shorter than the 24-28 nts originally estimated for the influenza A virus NP based on population average. The 12-nt stoichiometry was further confirmed by results from electron microscopy and dynamic light scattering. Considering that RNPs of ISAV and the influenza viruses have similar morphologies and dimensions, our findings suggest that NP-free RNA may exist on orthomyxovirus RNPs, and selective RNP packaging may be accomplished through direct RNA-RNA interactions.
Department of Biochemistry and Cell Biology, Rice University, Houston, Texas, United States of America.
Wenjie
W
Zheng
John
J
Olson
Vikram
V
Vakharia
Yizhi Jane
YJ
Tao
D000595:Amino Acid Sequence; D001665:Binding Sites; D019281:Dimerization; D046829:Isavirus / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009698:Nucleoproteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism / Q000648:ultrastructure; D009843:Oligoribonucleotides / Q000737:chemistry / Q000378:metabolism; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000648:ultrastructure; D011487:Protein Conformation; D054730:Protein Interaction Domains and Motifs; D055550:Protein Stability; D012313:RNA / Q000737:chemistry* / Q000378:metabolism / Q000648:ultrastructure; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure; D012261:Ribonucleoproteins / Q000737:chemistry* / Q000378:metabolism / Q000648:ultrastructure; D016415:Sequence Alignment; D012995:Solubility; D014764:Viral Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism / Q000648:ultrastructure; D014961:X-Ray Diffraction
D009698:Nucleoproteins; D009843:Oligoribonucleotides; D010446:Peptide Fragments; D011994:Recombinant Proteins; D012261:Ribonucleoproteins; D014764:Viral Proteins; D012313:RNA