Long-lived o-Semiquinone Radical Anion is Formed from N-β-alanyl-5-S-glutathionyl-3,4-dihydroxyphenylalanine (5-S-GAD), an Insect-derived Antibacterial Substance

• Published in: Journal of biochemistry (Tokyo) Vol. 142; no. 1; pp. 41 - 48
• Main Authors: Akiyama, Nobuko, Nakanishi, Ikuo, Ohkubo, Kei, Satoh, Kazue, Tsuchiya, Koichiro, Nishikawa, Takeshi, Fukuzumi, Shunichi, Ikota, Nobuo, Ozawa, Toshihiko, Tsujimoto, Masafumi, Natori, Shunji
• Format: Journal Article
• Language: English
• Published: England Japanese Biochemical Society 01.07.2007; Oxford University Press
• Subjects: 5-S-GAD; Anti-Bacterial Agents - chemistry; Anti-Bacterial Agents - metabolism; Antineoplastic Agents - chemistry; Antineoplastic Agents - metabolism; Antioxidants - chemistry; Antioxidants - metabolism; Benzoquinones - chemistry; Benzoquinones - metabolism; Dihydroxyphenylalanine - analogs & derivatives; Dihydroxyphenylalanine - chemistry; Dihydroxyphenylalanine - metabolism; electron spin resonance; Electron Spin Resonance Spectroscopy; Free Radicals - chemistry; Free Radicals - metabolism; Glutathione - analogs & derivatives; Glutathione - chemistry; Glutathione - metabolism; hydrogen peroxide; Hydrogen Peroxide - metabolism; Hydrogen-Ion Concentration; Models, Molecular; o-semiquinone radical anion; Oxidation-Reduction; superoxide dismutase; Superoxides - chemistry; Superoxides - metabolism; Time Factors; electron spin resonance; 5-S-GAD; semiquinone radical anion; superoxide dismutase; hydrogen peroxide
• ISSN: 0021-924X; 1756-2651
• DOI: 10.1093/jb/mvm101

N-β-Alanyl-5-S-glutathionyl-3,4-dihydroxyphenylalanine (5-S-GAD), an insect-derived antibacterial peptide, generates hydrogen peroxide (H₂O₂) that exerts antitumour activity. We have investigated the precise mechanism of H₂O₂ production from 5-S-GAD by autoxidation aiming to understand its action toward tumour cells. Using the electron spin resonance (ESR) technique, we detected a strong signal due to radical formation from 5-S-GAD. Surprisingly, the ESR signal of the radical derived from 5-S-GAD appeared after incubation for 30 min at 37°C in the buffer at pH 7.4; the signal was persistently detected for 10 h in the absence of catalytic metal ions. The computer simulation of the observed ESR spectrum together with the theoretical calculation of the spin density of the radical species indicates that an o-semiquinone radical anion was formed from 5-S-GAD. We demonstrated that H₂O₂ is produced via the formation of superoxide anion ([Formula: see text]) by the electron-transfer reduction of molecular oxygen by the 5-S-GAD anion, which is in equilibrium with 5-S-GAD in the aqueous solution. The radical formation and the subsequent H₂O₂ production were inhibited by superoxide dismutase (SOD), when the antitumour activity of 5-S-GAD was inhibited by SOD. Thus, the formation of the o-semiquinone radical anion would be necessary for the antitumour activity of 5-S-GAD as an intermediate in the production of cytotoxic H₂O₂.