Divalent ion-dependent reversible swelling of tomato bushy stunt virus and organization of the expanded virion.

Abstract The divalent ion-dependent swelling of tomato bushy stunt virus at neutral pH has been investigated by a combination of analytical ultracentrifugation, fluorescence techniques and small-angle X-ray and neutron scattering. In the presence of EDTA, the virions swell by about 12% (in radius) in a narrow pH interval centered at about pH 7.0. Native virus is progressively changed into swollen particles with increasing pH, and no intermediate component could be observed. Upon swelling, virions exhibit a twofold increase of their intrinsic fluorescence; they remain resistant to RNAase but the viral RNA becomes accessible to ethidium bromide, which intercalates at about the same number of sites as in free RNA. The radial distribution of RNA and protein resembles that of the native virus, all radii being increased by 22 A. The protein remains clustered into two concentric shells separated by a 30 A thick shell containing most of the viral RNA. Swelling of native tomato bushy stunt virus at pH 7 is under the specific control of very strongly bound calcium ions, which may be antagonized by magnesium ions. Swollen virons can be recompacted in the presence of calcium ions at pH > 7.0, or by dialysis against EDTA-containing buffers at pH ≤ 6. Recompaction is progressive, from about pH 6.6 to 6.2. Throughout the transition, small-angle X-ray scattering curves are demonstrably not linear combinations of the two extremes, showing that intermediate particles must exist. Neutron and X-ray small-angle scattering curves of recompacted particles are identical to those of native virus, providing evidence that the gross features of the architecture have been recovered. It is only if recompaction takes place at pH 5 that virions behave.