A hierarchical binding mechanism governs the SARS-CoV-2 N protein-G3BP1 interaction: a computational insight

Abstract

The sequestration of host G3BP1/2 by the SARS-CoV-2 nucleocapsid (N) protein is vital for viral antagonism of the cellular stress response. While N-terminal residues 14–17 are critical for this interaction, the atomistic determinants defining viral variant affinities remain ambiguous. Integrating umbrella sampling and molecular dynamics simulations, we elucidate a hierarchical binding mechanism wherein a primary electrostatic anchor (Asp22) is locked by a secondary hydrophobic interface (Phe17). Our calculations reveal that the F17A mutation exerts a significantly more detrimental effect on binding than the Omicron P13L mutation or the direct removal of the electrostatic anchor (D22A). Crucially, F17A induces an allosteric collapse: the loss of the hydrophobic contact triggers backbone rearrangements that displace the critical Asp22 anchor. In contrast, D22A selectively abrogates the electrostatic interaction while preserving the hydrophobic core, resulting in a less severe affinity loss. These findings identify Phe17 as a structural linchpin allosterically controlling the primary electrostatic interaction, providing a detailed rationale for the mutational landscape of this viral–host interface.