Figure 4
Structural Changes of the Allosteric Site and the Proposed Inhibition Mechanism. (A) The acyl-intermediate model of MuPEP1-MuComC, in which the thioester bond is formed between Cys17 of PEP and the cleavage site Gly of MuComC (blue), was obtained by molecular dynamics simulation18. The positional difference for each C α atom between the free MuPEP1 (PDB ID code 3K8U) and the acyl-intermediate model was calculated and shown. Residues that showed significant differences (≥1 Å) are indicated as spheres, of which the diameters define relative difference distances. Arrowheads indicate the binding site of Compound 2. The N-terminal (1–62) and the C-terminal (63–150) subdomains are colored in light and deep tones, respectively. Arg93, which shows a large structural shift, is located at the entrance to the narrow cleft of the Gly–Gly binding site. The structural changes around Arg93 upon formation of the acyl-intermediate might be propagated and amplified to cause large structural changes around the remote Compound 2-binding site. (B) Superimposition of the overall structures of the tMuPEP1-Compoud 2 complex (gray), the free MuPEP1 (blue), and the acyl-intermediate model of MuPEP1-MuComC18 (pink). MuComC and Compound 2 are shown as stick and sphere models, respectively. The distinction between light and deep tones is the same as in (A). (C) The reaction scheme of MuPEP1 and mechanism for non-competitive inhibition. E, enzyme; S, substrate; P, product; I, inhibitor; K m, Michaelis constant; k cat, turnover number; and K i, inhibition constant. After the substrate binding step, the reaction proceeds via a transition state (ES‡), in which the space of a pocket on the protein surface is squeezed (broken arrows). The inhibitor binds to this pocket and inhibits the enzyme-catalyzed reaction by preventing structural changes. In the non-competitive inhibition model, K i ′ is supposed to be the same as K i.
