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Histone Deacetylase 8: Characterization of Physiological Divalent Metal Catalysis
Michael R Nechay1, Nathan M Gallup1, Amanda Morgenstern2
1Department of Chemistry and Biochemistry, University of California, Los Angeles , Los Angeles, California 90095, United States.
Abstract:
Histone deacetylases (HDACs) are responsible for the removal of acetyl groups from histones, resulting in gene silencing. Overexpression of HDACs is associated with cancer, and their inhibitors are of particular interest as chemotherapeutics. However, HDACs remain a target of mechanistic debate. HDAC class 8 is the most studied HDAC, and of particular importance due to its human oncological relevance. HDAC8 has traditionally been considered to be a Zn-dependent enzyme. However, recent experimental assays have challenged this assumption and shown that HDAC8 is catalytically active with a variety of different metals, and that it may be a Fe-dependent enzyme in vivo. We studied two opposing mechanisms utilizing a series of divalent metal ions in physiological abundance (Zn(2+), Fe(2+), Co(2+), Mn(2+), Ni(2+), and Mg(2+)). Extensive sampling of the entire protein with different bound metals was done with the mixed quantum-classical QM/DMD method. Density functional theory (DFT) on an unusually large cluster model was used to describe the active site and reaction mechanism. We have found that the reaction profile of HDAC8 is similar among all metals tested, and follows one of the previously published mechanisms, but the rate-determining step is different from the one previously claimed. We further provide a scheme for estimating the metal binding affinities to the protein. We use the quantum theory of atoms in molecules (QTAIM) to understand the different binding affinities for each metal in HDAC8 as well as the ability of each metal to bind and properly orient the substrate for deacetylation. The combination of this data with the catalytic rate constants is required to reproduce the experimentally observed trend in metal-depending performance. We predict Co(2+) and Zn(2+) to be the most active metals in HDAC8, followed by Fe(2+), and Mn(2+) and Mg(2+) to be the least active.
Insights
Histone deacetylase 8 (HDAC8) activity varies with different metals. This study reveals Co(2+) and Zn(2+) are most active, challenging previous assumptions about HDAC8
Area of Science:
- Biochemistry
- Molecular Biology
- Computational Chemistry
Background:
- Histone deacetylases (HDACs) regulate gene expression by removing acetyl groups from histones, and their dysregulation is linked to cancer.
- HDAC class 8 (HDAC8) is a key target in oncology, traditionally viewed as zinc-dependent, but recent findings suggest alternative metal dependencies.
- Understanding the catalytic mechanism and metal ion interactions of HDAC8 is crucial for developing targeted cancer therapies.
Purpose of the Study:
- To investigate the role of various divalent metal ions in HDAC8 catalytic activity and mechanism.
- To elucidate the binding affinities and substrate orientation capabilities of different metals within the HDAC8 active site.
- To reconcile experimental observations with computational predictions regarding HDAC8 metal dependency.
Main Methods:
- Mixed quantum-classical QM/DMD simulations were employed for extensive sampling of HDAC8 with various bound metal ions.
- Density functional theory (DFT) using a large cluster model was applied to analyze the active site and reaction mechanism.
- Quantum theory of atoms in molecules (QTAIM) was utilized to assess metal-protein binding affinities and substrate interactions.
Main Results:
- The reaction profile of HDAC8 was found to be similar across tested metals (Zn(2+), Fe(2+), Co(2+), Mn(2+), Ni(2+), Mg(2+)), but the rate-determining step differs from prior claims.
- A method for estimating metal binding affinities to HDAC8 was developed, correlating with experimental trends.
- Computational analysis revealed distinct binding affinities and substrate orientations for each metal ion, influencing catalytic performance.
Conclusions:
- HDAC8 exhibits catalytic activity with multiple metal ions, with Co(2+) and Zn(2+) predicted as the most active.
- The study provides a framework for predicting metal ion efficacy in HDAC8 based on binding affinities and catalytic rates.
- Findings challenge the traditional view of HDAC8 as solely zinc-dependent and offer new insights for therapeutic strategies targeting HDACs in cancer.
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