Effects of oxygen on kynurenine-3-monooxygenase activity

Abstract

Kynurenine-3-monooxygenase (KM), the third enzyme in the kynurenine (KYN) pathway from tryptophan to quinolinic acid (QA), is a monooxygenase requiring oxygen, NADPH and FAD for the catalytic oxidation of L-kynurenine to 3-hydroxykynurenine and water. KM is innately low in the brain and similar in activity to indoleamine oxidase, the rate-limiting pathway enzyme. Accumulation in the CNS of QA, a known excitotoxin, is proposed to cause convulsions in several pathologies. Thus, we theorized that hyperbaric oxygen (HBO) induced convulsions arise from increased QA via oxygen K_m effects on this pathway [Brown OR, Draczynska-Lusiak. Oxygen activation and inactivation of quinolinate-producing and iron-requiring 3-hydroxyanthranilic acid oxidase: a role in hyperbaric oxygen-induced convulsions? Redox Report 1995; 1: 383–385]. To complement prior studies on the effects of oxygen on pathway enzymes, in this paper we report the effects of oxygen on KM. Brain and liver KM enzyme are not known to be identical, and some systemically-produced KYN pathway intermediates can permeate the brain and might stimulate the brain pathway. Thus, KM from both brain and liver was assayed at various oxygen substrate concentrations to evaluate, in vitro, the potential effects of increases in oxygen, as would occur in mammals breathing therapeutic and convulsive HBO. In crude tissue extracts, KM was not activated during incubation in HBO up to 6 atm. The effects of oxygen as substrate on brain and liver KM activity was nearly identical: activity was nil at zero oxygen with an apparent oxygen K_m of 20–22 µM. Maximum KM activity occurred at about 1000 µM oxygen and decreased slightly to plateau from 2000 to 8000 µM oxygen. This compares to approximately 30–40 µM oxygen typically reported for brain tissue of humans or rats breathing air, and an unknown but surely much lower value (perhaps below 1 µM) intracellularly at the site of KM. Thus HBO, as used therapeutically and at convulsive pressures, likely stimulates flux through the KM-catalyzed step of the KYN pathway in liver and in brain and could increase brain QA, by K_m effects on brain KM, or via increased KM pathway intermediates produced systemically (in liver) and transported into the brain.