Birth and Growth Kinetics of Brome Mosaïc Virus Microcrystals Marina Casselyn, Annette Tardieu, Hervé Delacroix, Stéphanie Finet  Biophysical Journal  Volume 87, Issue 4, Pages 2737-2748 (October 2004) DOI: 10.1529/biophysj.104.042036 Copyright © 2004 The Biophysical Society Terms and Conditions

Figure 1 (Top) Characteristic two-dimensional patterns. (Bottom) Corresponding normalized intensity scattering curves. (a) Static x-ray experiment: average of 20 two-dimensional scattering patterns (each 50ms) of a 2.5mg/ml BMV sample. (b) Experimental form factor I(0,s) of the BMV obtained from the less concentrated sample (2.5mg/ml) for 1.17 10−3<s<4.62 10−3Å−1, and from the 10 and the 20mg/ml for the s>4.62 10−3Å−1 (average of 20 frames of 50ms). (c) Example of a kinetic experiment with a sample containing 10mg/ml BMV and 5% PEG 20K (w/v). The three frames, chosen from a series of 20 frames (50ms of acquisition time), were recorded respectively as 180ms, 5s, and 1511s (∼25min) after the mixing. (Top) Progressive formation of diffraction rings. (Bottom) The corresponding normalized intensity scattering curves show the appearance of Bragg peaks. Biophysical Journal 2004 87, 2737-2748DOI: (10.1529/biophysj.104.042036) Copyright © 2004 The Biophysical Society Terms and Conditions

Figure 2 Diffraction peaks formation as a function of time (indicated in seconds) and of PEG molecular weight at concentration 10% (w/v). The labels (a), (b), and (c) represent, respectively, the diffracting data of samples containing PEG 3K in the presence of 20mg/ml BMV, PEG 8K in presence of 10mg/ml BMV, and PEG 20K in presence of 10mg/ml BMV. Note that for PEG 20K and to a lesser extent for PEG 8K a few peaks are already present at the beginning of the kinetic experiment, because of microcrystals stuck to the capillary from the previous run. The characteristic lobes of the scattering of soluble viruses are still visible. The labels (d), (e), and (f) are the corresponding structure factors (the scattered intensity divided by the form factor, to remove the scattering signal of the virus shape and improve the visibility of the peaks). Noteworthy is the fact that the width of the diffraction peaks decreases with increasing the PEG molecular weight. Biophysical Journal 2004 87, 2737-2748DOI: (10.1529/biophysj.104.042036) Copyright © 2004 The Biophysical Society Terms and Conditions

Figure 3 Evolution of the diffraction peaks as a function of time, for samples containing (top left) 10% PEG 3K, 20mg/ml BMV; (top right) 10% PEG 8K, 10mg/ml BMV; (bottom left) 5% PEG 20K, 10mg/ml BMV; and (bottom right) 5% PEG 20K, 20mg/ml BMV. The refinement of the peaks is visible on top-left panel. The peaks are numbered as a function of their position on bottom-left panel (see text). Biophysical Journal 2004 87, 2737-2748DOI: (10.1529/biophysj.104.042036) Copyright © 2004 The Biophysical Society Terms and Conditions

Figure 4 Evolution as a function of time of the intensity of peak #1, of the concentration of soluble virus (measured from the plateau level) and of the microcrystal radii of gyration (calculated from the peak width according to Eq. 13) for the spectra shown in Fig. 3. (Top left) 10% PEG 3K, 20mg/ml BMV; (top right) 10% PEG 8K, 10mg/ml BMV; (bottom left) 5% PEG 20K, 10mg/ml BMV; and (bottom right) 5% PEG 20K, 20mg/ml BMV. Biophysical Journal 2004 87, 2737-2748DOI: (10.1529/biophysj.104.042036) Copyright © 2004 The Biophysical Society Terms and Conditions

Figure 5 Evolution of the intensity of peaks #3 fitted by a Gaussian bell-shaped curve, for PEGs 20K and 8K 5% and 10% (w/v) in 10mg/ml BMV samples. Lag times necessary for microcrystals to appear are indicated by arrows. Biophysical Journal 2004 87, 2737-2748DOI: (10.1529/biophysj.104.042036) Copyright © 2004 The Biophysical Society Terms and Conditions