Reactivity of a Zn(Cys)2(His)2 Zinc Finger with Singlet Oxygen: Oxidation Directed toward Cysteines but not Histidines
Résumé
Singlet oxygen ((1) O2 ) is an important reactive oxygen species in biology that has deleterious effects. Proteins constitute the main target of (1) O2 in cells. Several organisms are able to mount a transcriptional defense against (1) O2 . ChrR and MBS are two proteins with Zn(Cys)2 (His)2 zinc finger sites that are involved in the regulation of the defense against (1) O2 . In this article, we investigate the reactivity of ZnCPF, a Zn(Cys)2 (His)2 classical betabetaalpha zinc finger, with (1) O2 . We show that ZnCPF interacts with (1) O2 mainly by physical quenching using a combination of (1) O2 luminescence quenching and kinetic competition experiments. The chemical reaction, which accounts for 5% of the interaction, leads to oxidation of cysteines but not histidines. Primary photooxidation products, identified by HPLC and mass spectrometry, are sulfinate (75±5%) and disulfides (25±5%). The peptides that have a single cysteine thiolate oxidized into a sulfinate are still able to bind one equivalent Zn(2+) but with a dramatic reduction of the binding constant compared to ZnCPF despite the preservation of the betabetaalpha fold, as shown by NMR and CD titrations. Finally, ZnCPF is compared to ZnLTC , a treble clef Zn(Cys)4 zinc finger, to gain further insight into the behavior of zinc fingers toward (1) O2 .