Molecular modeling the reaction mechanism of serine-carboxyl peptidases

We performed molecular modeling on the mechanism of serine-carboxyl peptidases, a novel class of enzymes active at acidic pH and distinguished by the conserved triad of amino acid residues Ser-Glu-Asp. Catalytic cleavage of a hexapeptide fragment of the oxidized B-chain of insulin by the Pseudomonas sedolisin, a member of the serine-carboxyl peptidases family, was simulated. Following motifs of the crystal structure of the sedolisin-inhibitor complex (PDB accession code 1NLU ) we designed the model enzyme-substrate (ES) complex and performed quantum mechanical-molecular mechanical calculations of the energy profile along a reaction route up to the acylenzyme (EA) complex through the tetrahedral intermediate (TI). The energies and forces were computed by using the PBE0 exchange-correlation functional and the basis set 6-31+G** in the quantum part and the AMBER force field parameters in the molecular mechanical part. Analysis of the ES, TI, and AE structures as well as of the corresponding transition states allows us to scrutinize the chemical transformations catalyzed by sedolisin. According to the results of simulations, the reaction mechanism of serine-carboxyl peptidases should be viewed as a special case of carboxyl (aspartic) proteases, with the nucleophilic water molecule being replaced by the Ser residue. The catalytic triad Ser-Glu-Asp in sedolisin functions differently compared to the well-known triad Ser-His-Asp of serine proteases, despite the structural similarity of sedolisin and the serine proteases member, subtilisin.