Crystal structure of the external aldimine of Citrobacter freundii methionine γ-lyase with glycine provides insight in mechanisms of two stages of physiological reaction and isotope exchange of α- and β-protons of competitive inhibitors

Pre-steady-state Kinetic and Structural Analysis of Interaction of Methionine γ-Lyase from Citrobacter freundii with Inhibitors

“…In the structure, there are tilts of the PLP ring around the C5-C2 axis and Tyr-113 ring to the solvent in comparison with their positions in the holoenzyme (25°and 19°, respectively). Similar tilts of the cofactor (17°) and Tyr-113 (28°) rings was demonstrated in a three-dimensional structure of external aldimine of the enzyme with glycine (36). The N⑀ atom of Lys-210 occupies two slightly different positions close to the ketimine nitrogen atom (ϳ3.5 Å).…”

“…This interpretation of the absorbance changes during binding of glycine is in agreement with the occurrence of fast C␣-proton exchange. According to the crystal structure of the external aldimine of MGL with glycine (36) and the known role of active site Lys residue as abstracting pro-R-proton in external aldimines of many PLP-dependent enzymes (E⅐Gly) 1 , the conformation fits with that having an orthogonal pro-R-proton.…”

“…The exocyclic oxygen makes hydrogen bonds with NH1 and NH2 atoms of Arg-374. This residue is involved in a binding of carboxylic groups in the external aldimine of MGL with glycine (36) and in Michaelis complexes of the enzyme with some substrates and inhibitors (20). The position of the bound cycloserine is additionally stabilized by stacking with side chains of Tyr-58 from the neighboring subunit and with Tyr-113 of its own protein chain.…”

Gene cloning, characterization, and cytotoxic activity of methionine γ‐lyase from Clostridium novyi

“…Alignment of known bacterial MGL sequences demonstrated that C118 is highly conserved . Crystal structures of C. freundii MGL and P. putida holoenzymes and C. freundii MGL complexes with amino acids mimicking intermediates of the physiological reaction revealed that this residue is part of the active site. In C. freundii, among the seven cysteine residues present in the amino acid sequence, two are highly reactive to modification by DTNB.…”

“…The content of homologous amino acid residues is high (74%–87%). Residues which form, according to X‐ray data, the active site of MGL from C. freundii and from C. sporogenes are marked with circles. Based on these structures, K214 in C. novyi is the cofactor‐binding residue forming a Schiff base.…”