Fig. 4: The crucial role of the activation loop of cGAS for PCIs to inhibit phase separation.
From: De novo design of protein condensation inhibitors by targeting an allosteric site of cGAS
a Sequence alignment of residues around the activation loop of cGAS from Homo sapiens (human; Uniprot Q8N884), Rhesus macaque (Rhesus monkey; Uniprot A0A0F7J4C8), Mus musculus (mouse; Uniprot Q8C6L5), Gallus gallus (chicken; Uniprot A0A8V0X930), Xenopus tropicalis (xenopus; Uniprot A0A6I8R967), and Danio rerio (zebrafish; Uniprot F1QCP4). Residue numbers above the sequences are from h-cGAS. Blue triangles highlight the residues of the activation loop that interact directly with DNA, according to the crystal structure with PDB ID of 6CT9. The black underlines indicate the highly conserved residues of the activation loop. The alignment was visualized with the ENDscript server. b Key residues involved in the interactions with XL-3156. c EC50 values determined by the DNA condensation assays for WT h-cGASCD and various mutants incubated with different compounds (50 μM). The residues at the activation loop (V218) and orthosteric site (S435 and Y436) that interact with XL-3156 were mutated. The Δloop refers to the substitution of YYEHVK with SG. Data are presented as mean ± SD from three independent experiments. d Distance variation between the Cα atoms of S213 at the activation loop and K414 at the helix (red) represents conformational states of the activation loop in DNA- (cyan) and ligand-bound (green) forms. Violin plots display the Cα-Cα distance distributions between S213 and K414 of h-cGASCD in an apo state and in complex with XL-3156, and G150 calculated based on 1-μs MD simulation trajectories. Violin plot elements: white dot (median), bounds of box (25th-75th percentiles, IQR = Interquartile Range), whiskers (1.5 × IQR from box edges). Data from 1 μs production phase trajectories (n = 500,000 frames/group, 2 ps sampling interval). Source data is available with this manuscript as a Source Data file.
