X-ray Crystal Structures of Candida albicans Dihydrofolate Reductase:  High Resolution Ternary Complexes in Which the Dihydronicotinamide Moiety of NADPH Is Displaced by an Inhibitor

Abstract

X-ray crystallographic analysis of 5-(4'-substituted phenyl)sulfanyl-2,4-diaminoquinazoline inhibitors in ternary complex with Candida albicans dihydrofolate reductase (DHFR) and NADPH revealed two distinct modes of binding. The two compounds with small 4'-substituents (H and CH3) were found to bind with the phenyl group oriented in the plane of the quinazoline ring system and positioned adjacent to the C-helix. In contrast, the more selective inhibitors with larger 4'-substituents (tert-butyl and N-morpholino) were bound to the enzyme with the phenyl group perpendicular to the quinazoline ring and positioned in the region of the active site that typically binds the dihydronicotinamide moiety of NADPH. The cofactor appeared bound to DHFR but with the disordered dihydronicotinamide swung away from the protein surface and into solution. This unusual inhibitor binding mode may play an important role in the high DHFR selectivity of these compounds and also may provide new ideas for inhibitor design.

Crystal Structure of Bacillus anthracis Dihydrofolate Reductase with the Dihydrophthalazine-Based Trimethoprim Derivative RAB1 Provides a Structural Explanation of Potency and Selectivity

“…Many MTX derivatives have been described that also seem to trend toward extended aliphatic extensions, but these compounds lack structural data for direct threedimensional comparison (36,42,43). Interestingly, some Plasmodium and Candida structures, which are considerably different from the B. anthracis enzyme, contain inhibitors, as well as a fortuitous buffer molecule (MES) in a position roughly equivalent with the linker-dihydrophthalazine portion of RAB1 (PDB ID code 2BLB) (37,48) (Fig. 5F), indicating unoccupied volume within the binding site complexed with those inhibitors.…”

Cofactor Hydrogen Bonding onto the Protein Main Chain Is Conserved in the Short Chain Dehydrogenase/Reductase Family and Contributes to Nicotinamide Orientation

“…For example, among the hydroxysteroid dehydrogenases belonging to the SDR family, 17␤-HSDs are attractive targets for structure-based rational drug design for the prevention and control of sex steroid-dependent cancers and 11␤-HSD1 is another good target for drug design for the therapy of several metabolic syndromes (insulin resistance, hyperlipidemia, arterial hypertension, and obesity). A recent study showed that two highly selective inhibitors can occupy the dihydronicotinamide binding site and thus displace the dihydronicotinamide moiety in dihydrofolate reductase (35). Also, in 3␣,20␤-hydroxysteroid dehydrogenase, a member of the SDR family, the hemisuccinate side chain of the inhibitor, carbenoxolone, makes a hydrogen bond with the hydroxyl group of the conserved residue Tyr 152 and occupies the position of the nicotinamide ring of the cofactor (36).…”

On the use of PIPSA to Guide Target‐Selective Drug Design

“…[5] Although known clinical drugs against DHFR show weak activity against C. albicans, [5] potent, selective inhibitors of C. albicans DHFR have been reported. [6,7] Their selectivity for C. albicans versus human DHFR has been ascribed in part to differences in protein-ligand hydrogen bonding. Such differences can be detected by analysis of the protein electrostatic potentials.…”