Differences in pressure stability of the three components of cowpea mosaic virus: implications for virus assembly and disassembly.

A comparison of pressure stability of empty capsids and ribonucleoprotein particles of cowpea mosaic virus (CPMV) is presented. A combination of high pressure and subdenaturing concentrations of urea was utilized to promote dissociation and denaturation. We found that RNA plays an important role in stabilizing the particles as well as in conferring reversibility to the pressure-induced denaturation. Dissociation and denaturation of the top component (empty capsid) was observed at 2.5 kbar and in the presence of 2.5 M urea. The pressure-dissociated state of the capsid protein had the characteristics of a denatured conformation as suggested by fluorescence spectra, lifetime of tryptophans, and binding of bis-ANS. The properties of the dissociated capsid protein were more similar to those of a molten-globule conformation, different from the more drastically unfolded state obtained using high concentrations of urea. Whereas the fluorescence of bis-ANS increased for the pressure-dissociated protein (1.5 M urea and 2.5 kbar), it decreased for the virus denatured by 6.0 M urea. Middle and bottom components underwent less than 50% change in center of spectral mass at 2.5 kbar and 2.5 M urea. The particles containing RNA could be fully affected by pressures of 2.5 kbar--as measured by the spectral shift--only in the presence of 5.0 M urea. RNA-containing capsids denatured by pressure did not bind bis-ANS, suggesting that the capsid protein continues to be bound to the RNA after the protein-protein contacts are broken by pressure. Reassembly of the nucleoprotein particles was obtained after decompression, reinforcing the idea that proteins had not dissociated from RNA.(ABSTRACT TRUNCATED AT 250 WORDS)