Mutations affecting the internal equilibrium of the reaction catalyzed by 6-aminohexanoate-dimer hydrolase

Abstract

The enzyme 6-aminohexanoate-dimer hydrolase catalyzes amide synthesis. The yield of this reverse reaction in 90% t-butyl alcohol was found to vary drastically when enzyme mutants with substitutions of several amino acids located at the entrance of the catalytic cleft were used. Movement of the loop region and the flip-flop of Tyr170 generate a local hydrophobic environment at the catalytic center of the enzyme. Here, we propose that the shift of the internal equilibrium between the enzyme–substrate complex and enzyme–product complex by the ‘water-excluding effect’ alters the rate of the forward and reverse reactions. Moreover, we suggest that the local hydrophobic environment potentially provides a reaction center suitable for efficient amide synthesis. Database: PDB code 3VWL: Hyb-24DNY-S187. PDB code 3VWM: Hyb-24DNY-A187. PDB code 3VWN: Hyb-24DNY-G187. PDB code 3A65: Hyb-24DN-A112/Ahx complex. PDB code 3A66: Hyb-24DNY-A112/Ahx complex. PDB code 3VWP: Hyb-24DNY-S187A112/Ahx complex. PDB code 3VWQ: Hyb-24DNY-A187A112/Ahx complex. PDB code 3VWR: Hyb-24DNY-G187A112/Ahx complex.