Fig. 3: Structures and assays of F102 variants.
From: A highly dynamic mononuclear non-heme iron enzyme for the two-step isonitrile biosynthesis
A Structure 6 shown in a 1.0 σ 2mFo-DFc composite omit map. Water molecules fill the polar part of the substrate-binding site. Two rotamer conformations of F102W keeps the alkyl chain binding site largely hydrophobic. Water molecules are shown as red spheres. Mg ions are shown as green spheres. Black dashed lines represent hydrogen bonds or ligations to metals. B Structure 7 in a 1.0 σ 2mFo-DFc composite omit map. When CADA is bound (shown in orange sticks), F102W side chain is only in one conformation and forms an additional hydrogen bond with CADA carboxylate (shown in magenta dashed line). Vanadyl ion is shown in sticks with V in gray and the oxo in red. C Structure 8 in a 1.0 σ 2mFo-DFc composite omit map. F102Y side chain is locked in one conformation. The substrate-binding site is occupied by a water molecule network that extends to the protein surface. D Structure 9 in a 1.0 σ 2mFo-DFc composite omit map. CADA binding makes the substrate-binding site hydrophobic but T203 alternate conformations could still allow water access from the protein surface. E Structure 10 in a 1.0 σ 2mFo-DFc composite omit map. An extended water network can be observed. F Structure 11 in a 1.0 σ 2mFo-DFc composite omit map. Electron density for CADA cannot be clearly observed in mFo-DFc difference map (+2.8 σ in green mesh). G Activity assays conducted with a 10-min incubation period and using a non-saturating concentration of CADA (1 mM) show relative product formations of three F102 variants in comparison to the WT enzyme. For each assay, data and error bars represent average and standard deviation from three independently performed experiments, respectively. Source data are provided as a Source Data file.
