Carbon-13 and proton nuclear magnetic resonance spectroscopy of plant viruses: evidence for protein-nucleic acid interactions in belladonna mottle virus and detection of polyamines in turnip yellow mosaic virus.
Belladonna mottle virus (BDMV) enriched in carbon-13 was isolated from infected tobacco plants grown in a 2% 13C02, 8% 12CO2 atmosphere. The enrichment led to a fivefold improvement in the signal to noise ratio of the (13)C NMR spectrum of BDMV. 1H and 13C NMR peaks of the intact virion are broad. However, upon removal of the RNA, many sharp peaks appear in spectra of the empty capsid which are attributed to aliphatic amino acid side chains at the inner surface of the protein shell that have gained segmental mobility. These include resonances assigned to the side chain of glutamate. Internal motions are necessary to account for the experimental 13C NMR linewidths. Aliphatic 13CHn (n = 1-3) groups must have correlation times on the order of 10 nsec and quaternary carbons must have correlation times between 50 and 300 nsec to explain the narrow line widths. No sharp peaks were observed in the aromatic regions of 1H or 13C NMR spectra; thus all the aromatic side chains seem to be tightly packed in the capsid as well as the virion. 1H and 13C NMR have been used for the first time to detect polyamines in virus particles. 1H NMR studies confirmed the presence of polyamines, apparently packaged during assembly, in turnip yellow mosaic virus (TYMV) and demonstrated their absence in BDMV. Sedimentation analysis has shown that the RNA is released upon increasing the pH above neutrality for BDMV but only above pH 11.5 for TYMV (R. Virudachalam, K. Sitaraman, K. L. Heuss, J. L. Markley, and P. Argos, Virology 130, 351-359, 1983). 1H NMR studies demonstrated that the BDMV capsid is permeable to polyamines and that the pH stability of BDMV with added spermidine is comparable to that of TYMV.
Virudachalam
R
R
Purdue University Biochemical Magnetic Resonance Laboratory, Department of Chemistry, Purdue University, West Lafayette, Indiana 47907, USA.
Sitaramant
K
K
Heusst
K L
KL
Argost
P
P
Markley
J L
JL