STRUCTURAL STUDIES OF THE ENVELOPED DSRNA BACTERIOPHAGE PHI-6 OF PSEUDOMONAS-SYRINGAE BY RAMAN-SPECTROSCOPY .2. NUCLEOCAPSID STRUCTURE AND THERMOSTABILITY OF THE VIRION, NUCLEOCAPSID AND POLYMERASE COMPLEX

Structures and thermostabilities of the double-stranded (ds) RNA bacteriophage φ6 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 φ6 virion are collectively deficient in β-sheet secondary structure. In particular, the major protein (P8) of the outer spherical shell of the φ6 nucleocapsid exhibits a secondary structure dominated largely by α-helix and irregular conformations. The absence of appreciable β-structure in the P8 subunit suggests a tertiary conformation lacking the β-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 α-helical. The results thus indicate a largely α-helical secondary structure for the major subunit (P1) of the φ6 nucleocapsid core, as well as for the PS 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 φ6 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 φ6 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 φ6 virion differ fundamentally from those of icosahedral single-stranded RNA viruses. Both Raman and circular dichroism spectra indicate that the dsRNA genome of φ6 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 φ6 genome, is discussed here as a model for assessing base-pairing and base-stacking interactions in other ribonucleoprotein assemblies.