Influence of in situ Limited Proteolysis of Potato Virus X on Change in the Structure of Virions According to the Small-Angle X-Ray Scattering Data and Tritium Labeling

Capsids of the potexvirus family virions are characterized by the presence on the surface of virions of partially disordered N-terminal protein fragments of various lengths. The present study is devoted to studying the effect of <i>in situ</i> removal of the N-terminal domain of coat protein (CP) on structural organization and physicochemical properties of the potato virus X (PVX) virions. The work considers PVX virions containing an intact Ps-form CP, as well as virions including an <i>in situ</i> degraded Pf-form devoid of 19/21 amino acid residues from the N-end (PVXΔN). Synchrotron small-angle X-ray scattering (SAXS), transmission electron microscopy (TEM), tritium bombardment, and several other physicochemical methods were used in the study. Analysis of the images obtained using TEM revealed similarities in the architecture of filamentous PVX and PVXΔN virions. SAXS results demonstrated differences in organization of the capsid of PVX and PVXΔN virions: the latter was characterized by the reduced size of the ordered regions, indicating partial disruption of the structure of the viral protein framework. In addition, based on the SAXS scattering curves, parameters of the spiral packing of virions in solution were calculated, and structural modeling of particles was performed. Modeling results also indicate changes in the structure of the capsid due to removal of the ΔN-peptide. Using information about the secondary structure of the PVX model (PDB ID: 6R7G) and data from our previous studies on tritium labeling of the surface sites of PVX and PVXΔN virions, comparative analysis of the label incorporation profiles into elements of the protein secondary structure was conducted. This approach made it possible to predict localization of the ΔN-peptide above the amino acid residues of neighboring helical subunits (122-129 and 143-153) and demonstrate stabilizing role of this peptide on the overall structure of the virion. Increase in the level of labelling in the C-terminal region after removal of the ΔN-peptide also indicates decrease in the compactness of the virion. In general, the gained knowledge could be useful when using virus-like nanoparticles in biotechnology.