Antioxidative Function and Substrate Specificity of NAD(P)H- dependent Alkenal/one Oxidoreductase

Abstract

There are several known routes for the metabolic detoxication of ␣,␤-unsaturated aldehydes and ketones, including conjugation to glutathione and reduction and oxidation of the aldehyde to an alcohol and a carboxylic acid, respectively. In this study, we describe a fourth class of detoxication that involves the reduction of the ␣,␤-carbon‫؍‬carbon double bond to a single bond. This reaction is catalyzed by NAD(P)H-dependent alkenal/ one oxidoreductase (AO), an enzyme heretofore known as leukotriene B 4 12-hydroxydehydrogenase, 15-oxoprostaglandin 13-reductase, and dithiolethione-inducible gene-1. AO is shown to effectively reduce cytotoxic lipid peroxidation products such as 4-hydroxy-2-nonenal (HNE) (k cat ). Furthermore, transfection of human embryonic kidney cells with a rat liver AO expression vector protected these cells from challenge with HNE. The concentration of HNE at which 50% of the cells were killed after 24 h increased from ϳ15 M in control cells to ϳ70 M in AO-transfected cells. Overexpression of AO also completely abolished protein alkylation by HNE at all concentrations tested (up to 30 M). Thus, we describe a novel antioxidative activity of a previously characterized bioactive lipid-metabolizing enzyme that could prove to be therapeutically or prophylactically useful due to its high catalytic rate and inducibility.

Analysis of the Cytosolic Proteome in a Cell Culture Model of Familial Amyotrophic Lateral Sclerosis Reveals Alterations to the Proteasome, Antioxidant Defenses, and Nitric Oxide Synthetic Pathways

“…LTB 4 12HD may have a wider role in antioxidant defense as well as in lipid messenger metabolism. A recent study (54) demonstrated that LTB 4 12HD was effective at reducing a wide variety of cytotoxic ␣,␤-unsaturated aldehydes and ketones including products of lipid peroxidation such as 4-hydroxy-2-nonenal (HNE). HNE has been shown to be elevated in the cerebrospinal fluid of SALS cases (55) and SALS spinal cord (56,57).…”

“…HNE has been shown to be elevated in the cerebrospinal fluid of SALS cases (55) and SALS spinal cord (56,57). A potential chemoprotective role for the enzyme was suggested by its ability to confer resistance of LTB 4 12HD-transfected cell lines to HNE-induced apoptosis (54). The association of a significant amount of LTB 4 12HD with membranes (54) and the nuclear associated staining of motor neurons in this study supports the notion that the enzyme may help to protect membranes from oxidative damage.…”

The Catalytic and Kinetic Mechanisms of NADPH-dependent Alkenal/one Oxidoreductase

“…1 is an enzyme that reduces the carbon-carbon double bond of a variety of ␣,␤-unsaturated aldehydes and ketones (1). It is coregulated in the rat with a variety of phase 2/antioxidative enzymes, including NAD(P)H:quinone reductase, glutathione S-transferases, and UDP-glucuronosyltransferases, through the Keap1/Nrf2 signaling pathway (2).…”

“…Because saturated carbonyls lack this reactive moiety, they are often far less toxic. Thus, hydrogenation of the ␣,␤-double bond by AOR conceptually results in detoxication (1,3).…”

“…These reactive molecules probably mediate many of the detrimental effects of oxidative stress. 4HNE is extremely cytotoxic, an abundant product of lipid peroxidation, and an excellent substrate of AOR (1). Cells engineered to overexpress AOR are resistant to 4HNE-mediated cell death and protein adduct formation (1).…”