Structural studies on recombinant T=3 capsids of Sesbania mosaic virus coat protein mutants

Abstract

When expressed in Escherichia coli, the recombinant coat protein (rCP) of Sesbania mosaic virus (SeMV) was shown to self-assemble into T=3 capsids encapsidating CP mRNA and 23S rRNA derived from the host. Expression of CP-P53A, in which a conserved proline at position 53 in the β-annulus was substituted by alanine (CP-P53A), also produced similar capsids. Purified rCP and CP-P53A particles were crystallized and X-ray crystal structures of their mutant capsids were determined to resolutions of 3.6 and 4.1 Å, respectively. As in the native viral CP, the CPs in these recombinant capsids adopt the jelly-roll β-sandwich fold. The amino-terminal residues of the C subunits alone are ordered and form the β-annulus structure at the quasi-sixfold axes. A characteristic bend in the β-annulus remains unaffected in CP-P53A. The quasi-threefold interfaces of the capsids harbour calcium ions coordinated by ligands from the adjacent threefold-related subunits in a geometry that is analogous to that observed in the native capsid. Taken together with studies on deletion and substitution mutants of SeMV CP, these results suggest the possibility that the β-annulus and nucleic acid-mediated interactions may be less important for the assembly of sobemoviruses than previously envisaged.