Immature retroviral particles are assembled by self-association of the structural polyprotein precursor Gag. During maturation the Gag polyprotein is proteolytically cleaved yielding mature structural proteins: matrix (MA), capsid (CA) and nucleocapsid (NC) that re-assemble into a mature viral particle. Proteolytic cleavage causes the N-terminus of CA to fold back to form a ?-hairpin, anchored by an internal salt bridge between the N-terminal proline and inner aspartate. Using an in vitro assembly system of capsid-nucleocapsid protein (CANC), we studied formation of virus-like particles (VLP) of a Gammaretrovirus, the xenotropic murine leukemia virus (MLV)-related virus (XMRV). We show here that unlike for other retroviruses, XMRV CA and CANC do not assemble tubular particles characteristic of mature assembly. The prevention of the ?-hairpin formation, either by the deletion of the N-terminal proline or ten initial amino acids enabled the assembly of ?ProCANC or ?10CANC into immature-like, spherical particles. Detailed 3D structural analysis of these particles revealed that below a disordered N-terminal CA layer the C-terminus of CA assembles a typical immature lattice, which is linked by rod-like densities with the RNP.
