Biophysical and atomic force microscopy characterization of the RNA from satellite tobacco mosaic virus

Agarose gel electrophoresis, circular dichroism and differential scanning calorimetry showed that single-stranded RNA from satellite tobacco mosaic virus transforms from a conformationally ‘closed state’ at 4°C to a more conformationally ‘open state’ at 65°C. The transition is reversible and shows no hysteresis. Atomic force microscopy (AFM) allowed visualization of the two states and indicated that the conformationally ‘closed state’ probably corresponds to the native encapsidated conformation, and that the ‘open state’ represents a conformation, characterized as short, thick chains of domains, as a consequence of the loss of tertiary interactions. Heating from 75°C to 85°C in the presence of EDTA was necessary to further unravel the ‘open’ conformation RNA into extended chains of lengths >280 nm. Virus exposed to low concentrations of phenol at 65°C, extruded RNA as distinctive ‘pigtails’ in a synchronous fashion, and these ‘pigtails’ then elongated, as the RNA was further discharged by the particles. Moderate concentrations of phenol at 65°C produced complete disruption of virions and only remains of decomposed particles and disordered RNA were evident. AFM images of RNA emerging from disrupted virions appear most consistent with linear arrangements of structural domains.