Ricq Emily L, Hooker Jacob M, Haggarty Stephen J
From the Department of Chemistry and Chemical Biology, Harvard University, Cambridge, Massachusetts 02138,; the Athinoula A. Martinos Center for Biomedical Imaging, Department of Radiology, Massachusetts General Hospital, Harvard Medical School, Charlestown, Massachusetts 02129, and; the Chemical Neurobiology Laboratory, Center for Human Genetic Research, Departments of Neurology & Psychiatry, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts 02114.
the Athinoula A. Martinos Center for Biomedical Imaging, Department of Radiology, Massachusetts General Hospital, Harvard Medical School, Charlestown, Massachusetts 02129, and.
J Biol Chem. 2016 Nov 18;291(47):24756-24767. doi: 10.1074/jbc.M116.734426. Epub 2016 Sep 15.
Lysine demethylation of proteins such as histones is catalyzed by several classes of enzymes, including the FAD-dependent amine oxidases KDM1A/B. The KDM1 family is homologous to the mitochondrial monoamine oxidases MAO-A/B and produces hydrogen peroxide in the nucleus as a byproduct of demethylation. Here, we show KDM1A is highly thiol-reactive in vitro and in cellular models. Enzyme activity is potently and reversibly inhibited by the drug disulfiram and by hydrogen peroxide. Hydrogen peroxide produced by KDM1A catalysis reduces thiol labeling and inactivates demethylase activity over time. MALDI-TOF mass spectrometry indicates that hydrogen peroxide blocks labeling of cysteine 600, which we propose forms an intramolecular disulfide with cysteine 618 to negatively regulate the catalytic activity of KDM1A. This activity-dependent regulation is unique among histone-modifying enzymes but consistent with redox sensitivity of epigenetic regulators.
诸如组蛋白等蛋白质的赖氨酸去甲基化由几类酶催化，包括依赖黄素腺嘌呤二核苷酸（FAD）的胺氧化酶KDM1A/B。KDM1家族与线粒体单胺氧化酶MAO-A/B同源，并在细胞核中产生过氧化氢作为去甲基化的副产物。在此，我们表明KDM1A在体外和细胞模型中具有高度的硫醇反应性。酶活性受到药物双硫仑和过氧化氢的强烈且可逆的抑制。由KDM1A催化产生的过氧化氢会随着时间的推移减少硫醇标记并使去甲基化酶活性失活。基质辅助激光解吸电离飞行时间质谱（MALDI-TOF）表明过氧化氢会阻断半胱氨酸600的标记，我们推测该半胱氨酸与半胱氨酸618形成分子内二硫键以负向调节KDM1A的催化活性。这种活性依赖性调节在组蛋白修饰酶中是独特的，但与表观遗传调节因子的氧化还原敏感性一致。
