Theoretical investigation of the catalytic mechanism of the protein arginine deiminase 4 enzyme

摘要

The mechanism of the transcriptional coregulator protein arginine deiminase 4 (PAD4) in catalyzing the calcium-dependent conversion of specific arginine residues to citrulline and ammonia, was studied in the framework of density functional theory using the hybrid B3LYP exchange correlation potential and a medium-large basis set. Active site of enzyme was modeled by four key catalytic residues: Asp350, His471, Asp473 and Cys645. A guanidinium-containing derivative was considered as substrate model. Results of the present investigation support the experimental hypothesis concerning the occurrence of a tetrahedral intermediate having ammonium as leaving group, during the catalytic process. Besides, in the transition state leading to this intermediate the nucleophilic activity of the thiolate anion of Cys645 and the ability of the imidazolinium cation of His471 in giving the proton to the substrate, appear mutually enhanced. The rate determining step of the whole process occurring in gas phase was recognized in the ammonia release step. Solvation effects on the catalytic mechanism determine significant modifications as far as the energetics is concerned. The data obtained in the protein medium indicate that the kinetics of the process could be controlled by the nucleophilic attack of cysteine sulfur to the substrate carbon atom concerted with the proton shift from imidazolium ion to one of guanidinium amino groups.