title: Single-electron reduction of quinone and nitroaromatic xenobiotics by recombinant rat neuronal nitric oxide synthase.
abstract: We examined the kinetics of single-electron reduction of a large number of structurally diverse quinones and nitroaromatic compounds, including a number of antitumour and antiparasitic drugs, and nitroaromatic explosives by recombinant rat neuronal nitric oxide synthase (nNOS, EC 1.14.13.39), aiming to characterize the role of nNOS in the oxidative stress-type cytotoxicity of the above compounds. The steady-state second-order rate constants (kcat/Km) of reduction of the quinones and nitroaromatics varied from 10² M⁻¹s⁻¹ to 10⁶ M⁻¹s⁻¹, and increased with an increase in their single-electron reduction potentials (E¹₇). The presence of Ca²⁺/calmodulin enhanced the reactivity of nNOS. These reactions were consistent with an 'outer sphere' electron-transfer mechanism, considering the FMNH∙/FMNH₂ couple of nNOS as the most reactive reduced enzyme form. An analysis of the reactions of nNOS within the 'outer sphere' electron-transfer mechanism gave the approximate values of the distance of electron transfer, 0.39-0.47 nm, which are consistent with the crystal structure of the reductase domain of nNOS. On the other hand, at low oxygen concentrations ([O₂] = 40-50 μM), nNOS performs a net two-electron reduction of quinones and nitroaromatics. This implies that NOS may in part be responsible for the bioreductive alkylation by two-electron reduced forms of antitumour aziridinyl-substituted quinones under a modest hypoxia.
authors.affiliation: Institute of Biochemistry of Vilnius University, Vilnius, Lithuania.
authors.forename: Žilvinas
authors.initials: Ž
authors.lastname: Anusevičius
authors.forename: Henrikas
authors.initials: H
authors.lastname: Nivinskas
authors.forename: Jonas
authors.initials: J
authors.lastname: Šarlauskas
authors.forename: Marie-Agnes
authors.initials: MA
authors.lastname: Sari
authors.forename: Jean-Luc
authors.initials: JL
authors.lastname: Boucher
authors.forename: Narimantas
authors.initials: N
authors.lastname: Čėnas
mesh_terms: D000818:Animals; D001388:Aziridines / Q000378:metabolism*; D016227:Benzoquinones / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D002147:Calmodulin / Q000378:metabolism; D004579:Electron Transport; D007700:Kinetics; D052248:Nitric Oxide Synthase Type I / Q000378:metabolism*; D009574:Nitro Compounds / Q000378:metabolism; D010084:Oxidation-Reduction; D021281:Quantitative Structure-Activity Relationship; D011809:Quinones / Q000378:metabolism*; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D015262:Xenobiotics / Q000378:metabolism*
chemical_list: D001388:Aziridines; D016227:Benzoquinones; D002147:Calmodulin; D009574:Nitro Compounds; D011809:Quinones; D011994:Recombinant Proteins; D015262:Xenobiotics; D052248:Nitric Oxide Synthase Type I; D002118:Calcium