Preliminary attempt to follow the enthalpy of an enzymatic reaction by ab initio computations: Catalytic action of papain

Abstract

AbstractA new theoretical approach to study the enthalpy variations occurring during an enzymatic reaction is presented. The structural modifications of the enzyme–substrate complex along the reaction path are distinguished as macro‐ and microdeformations. Macrodeformations, which concern primarily the approach of the substrate to the enzyme and the release of the reaction products and arise from nonbonded interactions, are treated with an empirical method for computing the energy of a macromolecule. Microdeformations, which are local displacements driven by variations of the electronic structure and its energy and involve only a limited portion of the complex, are treated with the ab initio SCF‐LCAO‐MO method. The reaction path is idealized as a sequence of major steps: at each step, first the empirical program REFINE is used to calculate the geometry of the system for that step, then the energy of an appropriate subsystem is computed ab initio with the program IBMOL, using the geometry provided by REFINE and applying small concerted atomic displacements. Thus along the entire reaction path one can obtain an energy profile computed with the ab initio method and compatible with the structure of the whole complex. This approach was applied here to the first steps of the reaction of proteolysis catalyzed by papain. The formation of an ion pair ImH+ …S− between the side chains of residues His‐159 and Cys‐25 was examined in detail. The results show that the instability of the ion pair decreases by ˜ 11.5 kcal/mol when the interactions with residues Asn‐175 and Ala‐ 160 are taken into account; the instability is further decreased by ˜2 kcal/mol after a partial geometry optimization. The energies of the noncovalent enzyme–substrate complex and of the tetrahedral intermediate were computed, considering N‐methyl acetamide (NMA) as model substrate and representing papain with the residues Cys‐25, His‐159, Gln‐19, and Ala‐160. The interaction energy of the noncovalent complex is ‐3.8 kcal/mol, compared to the value of +7.4 kcal/mol for the CH3S− ‐NMA complex. The tetrahedral intermediate is found to be less stable than the noncovalent complex by 27 and 38.5 kcal/mol, respectively, for the papain–NMA and the CH3S− ‐NMA systems. While these rather large energy differences are possibly due to the incorrect geometry of the tetrahedral intermediate and optimization of the structure is required, it appears that the interactions with the various protein residues represent a very important stabilization factor, which lowers the onthalpy variations during the reaction.