Catalysis by the Non-Heme Iron(II) Histone Demethylase PHF8 Involves Iron Center Rearrangement and Conformational Modulation of Substrate Orientation

被引:69
作者
Chaturvedi, Shobhit S. [1 ]
Ramanan, Rajeev [1 ]
Lehnert, Nicolai [2 ,3 ]
Schofield, Christopher J. [4 ]
Karabencheva-Christova, Tatyana G. [1 ]
Christov, Christo Z. [1 ]
机构
[1] Michigan Technol Univ, Dept Chem, Houghton, MI 49931 USA
[2] Univ Michigan, Dept Chem, Ann Arbor, MI 48109 USA
[3] Univ Michigan, Dept Biophys, Ann Arbor, MI 48109 USA
[4] Univ Oxford, Chem Res Lab, Mansfield Rd, Oxford OX1 5JJ, England
基金
美国国家科学基金会; 英国生物技术与生命科学研究理事会; 英国惠康基金;
关键词
histone lysine demethylation; 2-oxoglutarate; non-heme iron enzyme; QM/MM; metal-center rearrangement; enzyme mechanism; TAURINE/ALPHA-KETOGLUTARATE DIOXYGENASE; ELECTRONIC-STRUCTURE ANALYSIS; LINKED MENTAL-RETARDATION; OXYGEN-ATOM EXCHANGE; H BOND ACTIVATION; MOLECULAR-DYNAMICS; ALPHA-KETOGLUTARATE; CRYSTAL-STRUCTURE; DEPENDENT OXYGENASES; STRUCTURAL INSIGHTS;
D O I
10.1021/acscatal.9b04907
中图分类号
O64 [物理化学(理论化学)、化学物理学];
学科分类号
070304 ; 081704 ;
摘要
PHF8 (KDM7B) is a human non-heme 2-oxoglutarate (20G) JmjC domain oxygenase that catalyzes the demethylation of the di/mono-N-epsilon-methylated K9 residue of histone H3. Altered PHF8 activity is linked to genetic diseases and cancer; thus, it is an interesting target for epigenetic modulation. We describe the use of combined quantum mechanics/molecular mechanics (QM/MM) and molecular dynamics (MD) simulations to explore the mechanism of PHF8, including dioxygen activation, 2OG binding modes, and substrate demethylation steps. A PHF8 crystal structure manifests the 2OG C-1 carboxylate bound to iron in a nonproductive orientation, i.e., trans to His247. A ferryl-oxo intermediate formed by activating dioxygen bound to the vacant site in this complex would be nonproductive, i.e., "off-line" with respect to reaction with NE-methylated K9. We show rearrangement of the "off-line" ferryl-oxo intermediate to a productive "in-line" geometry via a solvent exchange reaction (called "ferryl-flip") is energetically unfavorable. The calculations imply that movement of the 20G C-1 carboxylate prior to dioxygen binding at a five-coordination stage in catalysis proceeds with a low barrier, suggesting that two possible 2OG C-1 carboxylate geometries can coexist at room temperature. We explored alternative mechanisms for hydrogen atom transfer and show that second sphere interactions orient the NE-methylated lysine in a conformation where hydrogen abstraction from a methyl C-H bond is energetically more favorable than hydrogen abstraction from the N-H bond of the protonated NE-methyl group. Using multiple HAT reaction path calculations, we demonstrate the crucial role of conformational flexibility in effective hydrogen transfer. Subsequent hydroxylation occurs through a rebound mechanism, which is energetically preferred compared to desaturation, due to second sphere interactions. The overall mechanistic insights reveal the crucial role of iron-center rearrangement, second sphere interactions, and conformational flexibility in PHF8 catalysis and provide knowledge useful for the design of mechanism-based PHF8 inhibitors.
引用
收藏
页码:1195 / 1209
页数:29
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