Ultrahigh resolution drug design. II. Atomic resolution structures of human aldose reductase holoenzyme complexed with fidarestat and minalrestat: Implications for the binding of cyclic imide inhibitors

Abstract

The X-ray structures of human aldose reductase holoenzyme in complex with the inhibitors Fidarestat (SNK-860) and Minalrestat (WAY-509) were determined at atomic resolutions of 0.92 A and 1.1 A, respectively. The hydantoin and succinimide moieties of the inhibitors interacted with the conserved anion-binding site located between the nicotinamide ring of the coenzyme and active site residues Tyr48, His110, and Trp111. Minalrestat's hydrophobic isoquinoline ring was bound in an adjacent pocket lined by residues Trp20, Phe122, and Trp219, with the bromo-fluorobenzyl group inside the "specificity" pocket. The interactions between Minalrestat's bromo-fluorobenzyl group and the enzyme include the stacking against the side-chain of Trp111 as well as hydrogen bonding distances with residues Leu300 and Thr113. The carbamoyl group in Fidarestat formed a hydrogen bond with the main-chain nitrogen atom of Leu300. The atomic resolution refinement allowed the positioning of hydrogen atoms and accurate determination of bond lengths of the inhibitors, coenzyme NADP+ and active-site residue His110. The 1'-position nitrogen atom in the hydantoin and succinimide moieties of Fidarestat and Minalrestat, respectively, form a hydrogen bond with the Nepsilon2 atom of His 110. For Fidarestat, the electron density indicated two possible positions for the H-atom in this bond. Furthermore, both native and anomalous difference maps indicated the replacement of a water molecule linked to His110 by a Cl-ion. These observations suggest a mechanism in which Fidarestat is bound protonated and becomes negatively charged by donating the proton to His110, which may have important implications on drug design.

Aldose Reductase Inhibitors as Potential Therapeutic Drugs of Diabetic Complications

“…These ligand-dependent conformations indicate a remarkable induced fit or flexibility of the active site. Nevertheless, at least three distinct binding pockets in the active site can be proposed as shown in Figure 2 according to a number of studies on crystal structures of AR by X-ray crystallography and mutagenesis [30,[33][34][35][36][37][38]. The first is usually occupied by the anion head of ligand and thus named "anion binding pocket".…”

“…The second is a hydrophobic pocket, known as specificity pocket, and lined by the residues Leu300, Cys298, Cys303, Trp111, Cys303, and Phe122 [30]. The specificity pocket displays a high degree of flexibility and the residues lining this pocket are not conserved in other aldo-keto reductases such as aldehyde reductase, The third is another hydrophobic pocket formed by the residues Trp20, Trp111, Phe122, and Trp219 [34]. Aldehyde reductase (EC 1.1.1.2) , another member of the aldo-keto reductase superfamily, is mentioned here because of its close similarities to AR which may be associated with the specificity of ARIs.…”

Binding of Fidarestat Stereoisomers with Aldose Reductase

“…A full geometry optimization was carried out at the Hartree-Fock/6-31G* level using GAMESS [43]. Optimization was started from standard CHARMM [39][40][41] values for the internal coordinates and the crystal structure geometry for each of the isomers of fidarestat [29][30][31][32][33]. Geometrical convergence was obtained after 43 cycles of optimization with a final gradient of 0.214×10 -3 Hartree/Bohr.…”

“…To keep the temperature at 300 K, the system was coupled to a heat bath with a time constant of 0.1 ps. Structures of (2S,4S)-Fidarestat and (2R,4S)-Fidarestat were taken from the crystal structures [31][32][33], in which fidarestat setereoisomers are complexed to ALR2. The complex was solvated in a periodic truncated octahedron using a modified TIP3P water model [45].…”

“…In this study, we report the calculation of the relative free energies of binding to ALR2 of fidarestat and its (2R,4S) stereoisomer using molecular dynamics (MD) simulations in an explicit aqueous environment. X-ray structural and mass spectrometric studies of binding of these and other ligands to the same template have been reported [32][33][34]. In our calculations, we have employed the thermodynamic integration (TI) approach [35] utilizing either Mulliken [36] or electrostatic potential fitted (EPF) [37][38] charges under CHARMM/MMFF [39][40][41][42] force fields for the perturbed part of the substrate.…”