High-resolution structures of scytalone dehydratase-inhibitor complexes crystallized at physiological pH

Abstract

Scytalone dehydratase is a molecular target of inhibitor design efforts aimed at preventing the fungal disease caused by Magnaporthe grisea. A method for cocrystallization of enzyme with inhibitors at neutral pH has produced several crystal structures of enzyme-inhibitor complexes at resolutions ranging from 1.5 to 2.2 Å . Four high resolution structures of different enzyme-inhibitor complexes are described. In contrast to the original X-ray structure of the enzyme, the four new structures have well-defined electron density for the loop region comprising residues 115-119 and a different conformation between residues 154 and 160. The structure of the enzyme complex with an aminoquinazoline inhibitor showed that the inhibitor is in a position to form a hydrogen bond with the amide of the Asn131 side chain and with two water molecules in a fashion similar to the salicylamide inhibitor in the original structure, thus confirming design principles. The aminoquinazoline structure also allows for a more confident assignment of donors and acceptors in the hydrogen bonding network.

Halogen…π Interactions as Important Contributors to Binding Affinity in Medicinal Chemistry

“…This protein-ligand interaction motif is also experimentally observed in other crystal structures of complexes of the same protein (PDB files 4STD and 7STD) with bromine and chlorine involved in this favourable contact to the π -electron system of the Gly165 carbonyl moiety. Interestingly, these contacts in the scytalone dehydratase complexes represent the only contact with main-chain atoms in this and related inhibitor complexes [92].…”

“…In Figure 8.4e the X-ray structure of scytalone dehydratase with a very potent inhibitor is displayed (PDB 6STD, resolution 1.8Å) [92]. The ligand-binding mode of the cyclopropane-carboxamide-based inhibitor exhibiting a K i value of 2.3 nM also shows an orthogonal interaction of a bromine atom at a phenyl ring with the plane of the neighbouring Gly165 carbonyl moiety (distances Br .…”

Roles of Substrate Distortion and Intramolecular Hydrogen Bonding in Enzymatic Catalysis by Scytalone Dehydratase

“…H110 is hydrogen-bonded to the substrate's C3 hydroxyl group and may be considered to function in selecting the axial conformation of the leaving group. F53, the most mobile active-site residue as determined from comparisons of X-ray structures of several SD inhibitor complexes ( Figure 2) (18,28), may be considered to function by pushing down the C3 position of the substrate so that it attains the productive geometry. In order for the influences of H110 and F53 to have an effect on the orientation of the C3 group, there must be a pivot to direct their leverage.…”

“…Here these influences are investigated with respect to the catalytic mechanism of scytalone dehydratase. Scytalone dehydratase (SD, EC 4.2.1.94), 1 the single dehydratase operating within the fungal melanin biosynthesis pathway of Magnaporthe grisea, is an efficient catalyst of the dehydration of its physiological substrates, scytalone and vermelone ( Figure 1) (11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(21). SD (22)(23)(24)(25)(26)(27)(28)(29)(30)(31) and 3HNR (1,3,8-trihydroxynaphthalene reductase) (32)(33)(34)(35)(36)(37)(38)(39), one of two naphthol reductases in the pathway (40,41), are the biochemical targets of fungicides in development and practice for protecting rice plants from blast.…”

“…Adding to their inherent value in inhibitor design, X-ray structures of the enzymeinhibitor complexes are useful in modeling substrates in the respective active sites (11, 14, 16-18, 35, 38, 41). Owing, in part, to its having a 3-5-fold larger binding pocket resulting in many more protein contacts with inhibitors (18,38), the potent inhibitors of SD function (K i ∼ 10 pM) have 1000-fold greater affinity than the potent 3HNR inhibitors (K i ∼ 10 nM). SD is the target of more recently developed fungicides that have lower application rates for disease control than the older fungicides that target 3HNR.…”