Allosteric mechanisms of post-translational modification regulation in human cyclic GMP-AMP synthase catalytic...

Cyclic GMP-AMP synthase (cGAS) activation is regulated by post-translational modifications (PTMs), yet the structural basis by which these modifications tune DNA binding, nucleotide engagement, and dimer-dependent activation remains unclear. Here, we investigated human cGAS catalytic-core complexes bound to dsDNA and ATP, comparing wild type with S305-phosphorylated and K384-, K394-, and K414-acetylated monomeric systems, followed by 2:2 cGAS-DNA dimeric assemblies focused on K394 acetylation. Monomeric simulations showed that PTMs regulate cGAS in a site-specific manner rather than through broad destabilization of the complex. S305 phosphorylation and K414 acetylation weakened ATP engagement by perturbing the catalytic-pocket environment, with K414 showing an allosteric effect through helix α7 rearrangement. K384 acetylation mainly altered the Zn-finger/DNA-contacting region, whereas K394 acetylation produced comparatively modest effects in the monomeric state. Because cGAS activity requires DNA-induced dimerization, we further analyzed WT_WT, WT_acK394, and acK394_acK394 dimers containing ATP, Mg²⁺, and Zn²⁺ over 1000 ns. The WT_WT dimer maintained the most stable DNA association and ATP-pocket organization, whereas WT_acK394 showed intermediate perturbation and acK394_acK394 displayed the strongest reduction in DNA-protein contacts, hydrogen bonding, and ATP-pocket stability. Together, these results indicate that PTMs remodel cGAS through residue-specific effects on catalytic and DNA-binding hotspots, with K394 acetylation exerting a stronger destabilizing influence in the dimeric assembly than in the monomer. This monomer-to-dimer analysis provides mechanistic insight into how PTM-dependent structural remodeling may tune cGAS activation and innate immune signaling.