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Study of the Redox Properties of Naphthazarin (5,8-dihydroxy-1,4-naphthoquinone) and Its Glutathionyl Conjugate in Biological Reactions: One- And Two-Electron Enzymatic Reduction

Study of the Redox Properties of Naphthazarin (5,8-dihydroxy-1,4-naphthoquinone) and Its Glutathionyl Conjugate in Biological Reactions: One- And Two-Electron Enzymatic Reduction

K Ollinger, J Llopis, E Cadenas

Arch Biochem Biophys. 1989 Dec;275(2):514-30.

PMID: 2512857

Abstract:

Naphthazarin (5,8-dihydroxy-1,4-naphthoquinone), the basic unit of several tetracyclic antitumor antibiotics, and its glutathione conjugate were reduced by the one- and two-electron transfer flavoproteins NADPH-cytochrome P450 reductase and DT-diaphorase to their semi- and hydroquinone forms, respectively. Kinetic studies performed on purified DT-diaphorase showed the following results: KNADPHm = 68 microM, KQuinonem = 0.92 microM, and Vmax 1300 nmol X min-1 X microgram enzyme-1. Similar studies performed on purified NADPH-cytochrome P450 reductase indicated a lower KNADPHm (10.5 microM) and higher KQuinonem (2.3 microM). The Vmax values were 20-fold lower (46 nmol X min-1 X micrograms enzyme-1) than those observed with DT-diaphorase. DT-diaphorase reduced the naphthazarin-glutathione conjugate with an efficiency 5-fold lower than that observed with the parent quinone. The nucleophilic addition of GSH to naphthazarin proceeded with GSH consumption at rates slower than those observed with 1,4-naphthoquinone and its monohydroxy derivative, 5-hydroxy-1,4-naphthoquinone. The initial rate of GSH consumption during these reactions did not vary whether the assay was carried out under anaerobic or aerobic conditions. Autoxidation accompanied the DT-diaphorase and NADPH-cytochrome P450 reductase catalysis of naphthazarin and its glutathionyl adduct as well as the 1,4-reductive addition of GSH to naphthazarin. Superoxide dismutase at catalytic concentrations (nM range) enhanced slightly (1.1- to 1.6-fold) the autoxidation following the enzymatic catalysis of naphthazarin. Autoxidation during the GSH reductive addition to 1,4-naphthoquinones decreased with increasing number of -OH substituents, 1,4-naphthoquinone greater than 5-hydroxy-1,4-naphthoquinone greater than 5,8-dihydroxy-1,4-naphthoquinone, thus revealing that the contribution of redox transitions other than autoxidation, e.g., cross-oxidation, to the decay of the primary product of nucleophilic addition increases with increasing number of -OH substituents. Superoxide dismutase enhanced substantially the autoxidation of glutathionyl-naphthohydroquinone adducts, thereby affecting only slightly the total GSH consumed and GSSG formed during the reaction. The present results are discussed in terms of the relative contribution of one- and two-electron transfer flavoproteins to the bioreductive activation of naphthazarin and its glutathionyl conjugate as well as the importance of autoxidation reactions in the mechanism(s) of quinone cytotoxicity.

Chemicals Related in the Paper:

Catalog Number	Product Name	Structure	CAS Number	Price
AP475387	5,8-Dihydroxy-1,4-naphthoquinone		475-38-7	Price

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