Ajoene Is an Inhibitor and Subversive Substrate of Human Glutathione Reductase and Trypanosoma cruzi Trypanothione Reductase: Crystallographic, Kinetic, and Spectroscopic Studies

Ajoene ((E,Z)-4,5,9-trithiadodeca-1,6,11-triene 9-oxide), a garlic-derived natural compound, is a covalent inhibitor as well as a substrate of human glutathione reductase (GR) and Trypanosoma cruzi trypanothione reductase (TR). The 2.1-A resolution crystal structure of GR inhibited by (E)-ajoene revealed a mixed disulfide between the active site Cys58 and the CH2CH−CH2−SO−CH2−CHCH−S moiety of ajoene. The modified enzyme has a markedly increased oxidase activity when compared to free GR. GR reduces (Z)-ajoene with a kcat/Km of 6.8 × 103 M-1 s-1 yielding 4,5,9-trithiadodeca-1,6,11-triene (deoxyajoene) and 4,8,9,13-tetrathiahexadeca-1,6,10,15-tetraene as stable reaction products. The reaction leads also to the formation of single-electron reduced products and concomitantly superoxide anion radicals as shown by coupling the reaction to the reduction of cytochrome c. The interactions between the flavoenzymes and ajoene are expected to increase the oxidative stress of the respective cell. The antiparasitic and ...