Targeting the intersubunit cavity of Plasmodium falciparum glutathione reductase by a novel natural inhibitor: computational and experimental evidence.

Abstract Glutathione reductase (GR), a homodimeric FAD-dependent disulfide reductase, is essential for redox homeostasis of the malaria parasite Plasmodium falciparum and has been proposed as an antimalarial drug target. In this study we performed a virtual screening against Pf GR, using the structures of about 170,000 natural compounds. Analysis of the two top-scoring molecules, TTB and EPB, indicated that these ligands are likely to interact with the homodimer intersubunit cavity of Pf GR with high binding energy scores of −9.67 and −9.60 kcal/mol, respectively. Both compounds had a lower affinity for human GR due to differences in structure and electrostatic properties. In order to assess the putative interactions in motion, molecular dynamics simulations were carried out for 30 ns, resulting in TTB being more dynamically and structurally favored than EPB. A closely related compound MDPI 21618 was tested on recombinant Pf GR and hGR, resulting in IC 50 values of 11.3 ± 2.5 μM and 10.2 ± 1.7 μM, respectively. Kinetic characterization of MDPI 21618 on Pf GR revealed a mixed-type inhibition with respect to glutathione disulfide ( K i  = 9.7 ± 2.3 μM) and an uncompetitive inhibition with respect to NADPH. Furthermore, MDPI 21618 was found to inhibit the growth of the chloroquine-sensitive P. falciparum strain 3D7 with an IC 50 of 3.2 ± 1.9 μM and the chloroquine-resistant Dd2 strain with an IC 50 of 3.2 + 1.6 μM. In drug combination assays with chloroquine, artemisinin, or mefloquine MDPI 21618 showed an antagonistic action, which might suggest partially overlapping routes of action. This study further substantiates research on Pf GR as a potential antimalarial drug target.