Fig. 4: Structures reveal determinant of difference in Zn²⁺ binding by AlaRS-Ed and ThrRS-Ed.

a Structural superimposition of AfuAlaRS cis-Ed (PDB: 2ZTG) in grey and EcThrRS-Ed (PDB: 1TKE) in blue showing good overlap of the structures with r.m.s.d. of ~ 2.4 Å over 135 Cα. b Zoomed in view of the catalytic pocket showing HXXXH and CXGXH motifs displaying proper steric organisation of the coordination sphere in AlaRS-Ed for coordinating Zn²⁺, but not in ThrRS-Ed, as reflected by the difference in inter-ligand distances. The distorted steric arrangement of the ligand residues arises from the distinct positioning of the loop spanning CXGXH in ThrRS-Ed is due to the Y173, which in case of AlaRS-Ed is an aliphatic amino acids V/I/L/T. c Sequence logo showing conservation of Y173 in ThrRS-Ed in organisms across bacteria, archaea and eukarya, whereas in case of AlaRS-Ed the corresponding residue position is occupied by aliphatic side chain amino acids V/I/L/T (Supplementary Fig. 7a). d Backbone trace view of superposition of all the structures of AlaRS-Ed and ThrRS-Ed available in PDB database with resolution ≥2.5 Å, showing distinct positioning of the loop region spanning CXGXH motif in both the proteins. e Structural superposition of SaThrRS-Ed holo (PDB: 1NYR) in green, EcThrRS-Ed (PDB: 1TKE) in blue and AfuAlaRS cis-Ed (PDB: 2ZTG) in grey, showing rotation of the side chain of Y172 in SaThrRS-Ed holo ~130° away from the position of Y173 in EcThrRS-Ed. f Zoomed in view of CXGXH loop region showing the distinct orientation of the main-chain carbonyl O of the conserved Gly in ThrRS-Ed apo, ThrRS-Ed holo and AlaRS-Ed. g Mean Phi and Psi torsion angles of conserved Gly (CXGXH) in ThrRS-Ed apo, ThrRS-Ed holo and AlaRS-Ed. h Ramachandran plot of conserved Gly (CXGXH) in all the structures of ThrRS-Ed apo, ThrRS-Ed holo and AlaRS-Ed.