PDBsum entry 1e3v

Isomerase

PDB id: 1e3v

Contents

Protein chains

128 a.a. *

Ligands

DXC ×2

Waters ×312

* Residue conservation analysis

References listed in PDB file

Key reference

Title

Detection of large pka perturbations of an inhibitor and a catalytic group at an enzyme active site, A mechanistic basis for catalytic power of many enzymes.

Authors

N.C.Ha, M.S.Kim, W.Lee, K.Y.Choi, B.H.Oh.

Ref.

J Biol Chem, 2000, 275, 41100-41106. [DOI no: 10.1074/jbc.M007561200]

PubMed id

11007792

Abstract

Delta(5)-3-Ketosteroid isomerase catalyzes cleavage and formation of a C-H bond at a diffusion-controlled limit. By determining the crystal structures of the enzyme in complex with each of three different inhibitors and by nuclear magnetic resonance (NMR) spectroscopic investigation, we evidenced the ionization of a hydroxyl group (pK(a) approximately 16.5) of an inhibitor, which forms a low barrier hydrogen bond (LBHB) with a catalytic residue Tyr(14) (pK(a) approximately 11.5), and the protonation of the catalytic residue Asp(38) with pK(a) of approximately 4.5 at pH 6.7 in the interaction with a carboxylate group of an inhibitor. The perturbation of the pK(a) values in both cases arises from the formation of favorable interactions between inhibitors and catalytic residues. The results indicate that the pK(a) difference between catalytic residue and substrate can be significantly reduced in the active site environment as a result of the formation of energetically favorable interactions during the course of enzyme reactions. The reduction in the pK(a) difference should facilitate the abstraction of a proton and thereby eliminate a large fraction of activation energy in general acid/base enzyme reactions. The pK(a) perturbation provides a mechanistic ground for the fast reactivity of many enzymes and for the understanding of how some enzymes are able to extract a proton from a C-H group with a pK(a) value as high as approximately 30.

Figure 1.
Fig. 1. Dimeric structure, enzymatic reaction mechanism, and competitive inhibitors of PI. a, the dimeric structure of KSI in complex with equilenin is shown with the three critical active site residues and equilenin in a ball-and-stick model. b, the enzyme mechanism of KSI proceeding through a dienolic intermediate is shown with the catalytic residues. The H-bond between the Tyr14 OH and the oxyanion of the intermediate is indicated in a conventional way to denote a LBHB formation, the proton being in the middle of the two heavy atoms. c, the three competitive inhibitors used in this study are shown along with their markedly different pK[a] values.

Figure 3.
Fig. 3. Interaction of DC with PI. a, stereoview of the bound inhibitor and the three catalytic residues is shown along with the 2F[o] F[c] electron density map at 2.0 Å resolution contoured at 1.0 . The H-bonds and their distances are indicated. b, four possible ionization states in the interaction of DC with the catalytic residues. Whereas the first binding mode results in three charged H-bonds, the rest of the binding modes result in one charged and two neutral H-bonds.

The above figures are reprinted by permission from the ASBMB: J Biol Chem (2000, 275, 41100-41106) copyright 2000.