Molecular-Level Insight into the Differential Oxidase and Oxygenase Reactivities of de Novo Due Ferri Proteins
详细信息 查看全文
- 作者：Rae Ana Snyder ; Susan E. Butch ; Amanda J. Reig ; William F. DeGrado ; Edward I. Solomon
- 刊名：Journal of the American Chemical Society
- 出版年：2015
- 出版时间：July 29, 2015
- 年：2015
- 卷：137
- 期：29
- 页码：9302-9314
- 全文大小：809K
- ISSN：1520-5126
文摘
Using the single-chain <i>due ferrii> (DFsc) peptide scaffold, the differential oxidase and oxygenase reactivities of two 4A鈫?G variants, one with two histidines at the diiron center (G4DFsc) and the other with three histidines (3His-G4DFsc(Mut3)), are explored. By controlling the reaction conditions, the active form responsible for 4-aminophenol (4-AP) oxidase activity in both G4DFsc and 3His-G4DFsc(Mut3) is determined to be the substrate-bound biferrous site. Using circular dichroism (CD), magnetic CD (MCD), and variable-temperature, variable-field (VTVH) MCD spectroscopies, 4-AP is found to bind directly to the biferrous sites of the DF proteins. In G4DFsc, 4-AP increases the coordination of the biferrous site, while in 3His-G4DFsc(Mut3), the coordination number remains the same and the substrate likely replaces the additional bound histidine. This substrate binding enables a two-electron process where 4-AP is oxidized to benzoquinone imine and O2 is reduced to H2O2. In contrast, only the biferrous 3His variant is found to be active in the oxygenation of <i>pi>-anisidine to 4-nitroso-methoxybenzene. From CD, MCD, and VTVH MCD, <i>pi>-anisidine addition is found to minimally perturb the biferrous centers of both G4DFsc and 3His-G4DFsc(Mut3), indicating that this substrate binds near the biferrous site. In 3His-G4DFsc(Mut3), the coordinative saturation of one iron leads to the two-electron reduction of O2 at the second iron to generate an end-on hydroperoxo-Fe(III) active oxygenating species.