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. 2023 Jul 11;13(14):9817–9828. doi: 10.1021/acscatal.3c01294

Figure 6.

Figure 6

GntC X-ray crystal structure and site-directed mutagenesis identify important amino acid residues for catalysis. (A) GntC crystal structure with bound PLP-1 substrate (yellow) crystallized as a dimer of homodimers at 2.04 Å resolution (PDB ID: 8FFU, only one homodimer depicted for simplicity). The active site is at the interface of the homodimer between the green and teal monomers, consistent with other PLP-dependent enzyme structures. (B) GntC active site with Fo-Fc electron density map for bound PLP-1 (blue mesh, σ = 1.0) imposed on the ligand (left) and amino acid residues proposed to participate in substrate recognition and catalysis (right), including E9, S25, T84, and K219 (green monomer), and N52* and H250* (teal monomer). (C) Simplified active site comparison of GntC (green/teal) and capreomycidine cyclase OrfR residues (gray, PDB ID: 4M2M) with bound PLP-1 (yellow) and PLP-l-arg (gray) substrates, respectively. (D) In vitro enzyme assays for 2 production with soluble GntC mutants in comparison to wild-type (WT) GntC.