Structural Basis\nfor Alternative Self-Assembly Pathways\nLeading to Different Human Immunodeficiency Virus Capsid-Like Nanoparticles

The\nmechanisms that underlie the spontaneous and faithful\nassembly\nof virus particles are guiding the design of self-assembling protein-based\nnanostructures for biomedical or nanotechnological uses. In this study,\nthe human immunodeficiency virus (HIV-1) capsid was used as a model\nto investigate what molecular feature(s) may determine whether a protein\nnanoparticle with the intended architecture, instead of an aberrant\nparticle, will be self-assembled <i>in vitro</i>. Attempts\nof using the HIV-1 capsid protein CA for achieving <i>in vitro</i> the self-assembly of cone-shaped nanoparticles that contain CA hexamers\nand pentamers, similar to authentic viral capsids, had typically yielded\nhexamer-only tubular particles. We hypothesized that a reduction in\nthe stability of a transient major assembly intermediate, a trimer\nof CA dimers (ToD), will increase the propensity of CA to assemble <i>in vitro</i> into cone-shaped particles instead of tubes. Certain\namino acid substitutions at CA-CA interfaces strongly favored <i>in vitro</i> the assembly of cone-shaped nanoparticles that\nresembled authentic HIV-1 capsids. All-atom MD simulations indicated\nthat ToDs formed by CA mutants with increased propensity for assembly\ninto cone-shaped particles are destabilized relative to ToDs formed\nby wt CA or by another mutant that assembles into tubes. The results\nalso indicated that ToD destabilization is mediated by conformational\ndistortion of different CA–CA interfaces, which removes some\ninterprotein interactions within the ToD. A model is proposed to rationalize\nthe linkage between reduced ToD stability and increased propensity\nfor the formation of CA pentamers during particle growth <i>in\nvitro</i>, favoring the assembly of cone-shaped HIV-1 capsid-like\nnanoparticles.