Factors determining the relative stability of anionic tetrahedral complexes in serine protease catalysis and inhibition

Quantum mechanical ab initio (RHF/6-31+ G*//RHF/3-21G) calculations were us ed to simulate the formation of the tetrahedral complex intermediate (TC) i n serine protease active site by substrates and transition-state analog inh ibitors. The enzyme active site was simulated by an assembly of the amino a cids participating in catalysis, whereas the substrates and inhibitors were simulated by small ligands, acetamide ((1) over bar) and trifluoroacetone ((1) over bar), respectively. For the first time, the principal factors det ermining the relative stability of the TC in serine proteases are arranged according to their energy contributions, These include (a) formation of the new covalent bond between Ser195 O-gamma and the electrophilic center of a ligand; (b) stabilization of the oxyanion in the oxyanion hole; (c) basic catalysis by His57; and (d) hydrogen bond between Asp102 carboxylate and N- delta of the protonated His57 We have directly calculated the gas-phase rel ative free energy of formation of TCAS ((1) over bar) and TCAS ((1) over ba r), the value of Delta Delta G(g)[TCAS((2) over bar,(1) over bar)]. It is D elta E-cov, the relative energy of the new covalent bond between the enzyme and the ligand formed in a TC that determines the experimentally observed large difference in the stability of TCs formed by substrates and TS-analog inhibitors of serine proteases, me demonstrated that the relative stabilit y of TCs formed by a series of mono- and dipeptide amides and TFKs, derived from experimental kinetic data, can be rather well approximated by the sum of the theoretically calculated! value of Delta Delta G(g)[TCAS((2) over b ar,(1) over bar)] and the difference in hydration free energies of isolated Ligands. (C) 2000 WiIey-Liss, Inc.