Cryo-EM density of a reaction intermediate in the ACLY active site. (a)
Cryo-EM density (2.5 σ) corresponding to modeled phosphor-(3S)-citryl-CoA
in the ASH domain of the ACLY-E599Q–ATP–citrate–CoA-D2
structure (PDB- 6UUW/EMD-20902) reported in Wei et
al.2. (b)
Cryo-EM density (3.5 σ) corresponding to the reaction intermediate in the
ASH domain after non-uniform refinement together with particle symmetry
expansion, particle subtraction and local refinement in Cryosparc
(PDB-7LIW/EMD-23387) 4,5. While the cryo-EM density can be best
modeled as CoA + phospho-citrate (top, PDB-7LIW), at the current resolution we
cannot exclude the additional presence of phosphate + (3S)-citryl-CoA (middle),
and phospho-(3S)-citryl-CoA (bottom). The figure was made in Pymol. Important
contacting residues, and catalytic E599, with their corresponding cryo-EM
densities are shown for reference.