Structural studies of the enveloped dsRNA bacteriophage phi 6 of Pseudomonas syringae by Raman spectroscopy. II. Nucleocapsid structure and thermostability of the virion, nucleocapsid and polymerase complex.
Structures and thermostabilities of the double-stranded (ds) RNA bacteriophage phi 6 and of its isolated nucleocapsid-polymerase complex (nucleocapsid core) and dsRNA components have been investigated by Raman spectroscopy. The spectra show that proteins of the phi 6 virion are collectively deficient in beta-sheet secondary structure. In particular, the major protein (P8) of the outer spherical shell of the phi 6 nucleocapsid exhibits a secondary structure dominated largely by alpha-helix and irregular conformations. The absence of appreciable beta-structure in the P8 subunit suggests a tertiary conformation lacking the beta-barrel motif common to subunits of most other spherical viral capsids. In addition, the Raman spectra show that subunits of the dodecahedral nucleocapsid core are also predominantly alpha-helical. The results thus indicate a largely alpha-helical secondary structure for the major subunit (P1) of the phi 6 nucleocapsid core, as well as for the P8 subunit of the outer spherical shell. Using Raman difference spectroscopy, we demonstrate that proteins of the nucleocapsid core (P1, P2, P4 and P7) interact extensively with the packaged phi 6 RNA genome, and further, that conformational stability of the packaged RNA is reduced upon removal from the core. Also, we find that proteins of the phi 6 nucleocapsid are significantly more thermostable than proteins of the viral membrane envelope, which are reported in the accompanying paper (Li et al., 1993). The present results suggest that both the architectural principles and modes of protein-RNA interaction in the phi 6 virion differ fundamentally from those of icosahedral single-stranded RNA viruses. Both Raman and circular dichroism spectra indicate that the dsRNA genome of phi 6 is an A-form structure. The Raman marker bands signify the presence only of C3'-endo/anti nucleoside conformers. The Raman signature of dsRNA, revealed in the spectrum of the phi 6 genome, is discussed here as a model for assessing base-pairing and base-stacking interactions in other ribonucleoprotein assemblies.
Bamford
J K
JK
Department of Genetics, University of Helsinki, Finland.
Bamford
D H
DH
Li
T
T
Thomas
G J
GJ
D017126:Bacteriophage phi 6 / Q000737:chemistry* / Q000201:enzymology / Q000648:ultrastructure; D002213:Capsid / Q000737:chemistry*; D012321:DNA-Directed RNA Polymerases / Q000737:chemistry*; D016679:Genome, Viral; D046911:Macromolecular Substances; D009690:Nucleic Acid Conformation; D017433:Protein Structure, Secondary; D011549:Pseudomonas; D012330:RNA, Double-Stranded / Q000737:chemistry*; D013059:Spectrum Analysis, Raman / Q000379:methods; D013816:Thermodynamics; D014771:Virion / Q000737:chemistry* / Q000648:ultrastructure
D046911:Macromolecular Substances; D012330:RNA, Double-Stranded; D012321:DNA-Directed RNA Polymerases