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Catalysis by the Non-Heme Iron(II) Histone Demethylase PHF8 Involves Iron Center Rearrangement and Conformational Modulation of Substrate Orientation
[Image: see text] PHF8 (KDM7B) is a human non-heme 2-oxoglutarate (2OG) JmjC domain oxygenase that catalyzes the demethylation of the di/mono-N(ε)-methylated K9 residue of histone H3. Altered PHF8 activity is linked to genetic diseases and cancer; thus, it is an interesting target for epigenetic mod...
Autores principales: | , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
American Chemical
Society
2019
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Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6970271/ https://www.ncbi.nlm.nih.gov/pubmed/31976154 http://dx.doi.org/10.1021/acscatal.9b04907 |
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author | Chaturvedi, Shobhit S. Ramanan, Rajeev Lehnert, Nicolai Schofield, Christopher J. Karabencheva-Christova, Tatyana G. Christov, Christo Z. |
author_facet | Chaturvedi, Shobhit S. Ramanan, Rajeev Lehnert, Nicolai Schofield, Christopher J. Karabencheva-Christova, Tatyana G. Christov, Christo Z. |
author_sort | Chaturvedi, Shobhit S. |
collection | PubMed |
description | [Image: see text] PHF8 (KDM7B) is a human non-heme 2-oxoglutarate (2OG) JmjC domain oxygenase that catalyzes the demethylation of the di/mono-N(ε)-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 N(ε)-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 2OG 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 N(ε)-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 N(ε)-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. |
format | Online Article Text |
id | pubmed-6970271 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2019 |
publisher | American Chemical
Society |
record_format | MEDLINE/PubMed |
spelling | pubmed-69702712020-01-21 Catalysis by the Non-Heme Iron(II) Histone Demethylase PHF8 Involves Iron Center Rearrangement and Conformational Modulation of Substrate Orientation Chaturvedi, Shobhit S. Ramanan, Rajeev Lehnert, Nicolai Schofield, Christopher J. Karabencheva-Christova, Tatyana G. Christov, Christo Z. ACS Catal [Image: see text] PHF8 (KDM7B) is a human non-heme 2-oxoglutarate (2OG) JmjC domain oxygenase that catalyzes the demethylation of the di/mono-N(ε)-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 N(ε)-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 2OG 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 N(ε)-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 N(ε)-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. American Chemical Society 2019-12-11 2020-01-17 /pmc/articles/PMC6970271/ /pubmed/31976154 http://dx.doi.org/10.1021/acscatal.9b04907 Text en Copyright © 2019 American Chemical Society This is an open access article published under a Creative Commons Attribution (CC-BY) License (http://pubs.acs.org/page/policy/authorchoice_ccby_termsofuse.html) , which permits unrestricted use, distribution and reproduction in any medium, provided the author and source are cited. |
spellingShingle | Chaturvedi, Shobhit S. Ramanan, Rajeev Lehnert, Nicolai Schofield, Christopher J. Karabencheva-Christova, Tatyana G. Christov, Christo Z. Catalysis by the Non-Heme Iron(II) Histone Demethylase PHF8 Involves Iron Center Rearrangement and Conformational Modulation of Substrate Orientation |
title | Catalysis by the
Non-Heme Iron(II) Histone Demethylase
PHF8 Involves Iron Center Rearrangement and Conformational Modulation
of Substrate Orientation |
title_full | Catalysis by the
Non-Heme Iron(II) Histone Demethylase
PHF8 Involves Iron Center Rearrangement and Conformational Modulation
of Substrate Orientation |
title_fullStr | Catalysis by the
Non-Heme Iron(II) Histone Demethylase
PHF8 Involves Iron Center Rearrangement and Conformational Modulation
of Substrate Orientation |
title_full_unstemmed | Catalysis by the
Non-Heme Iron(II) Histone Demethylase
PHF8 Involves Iron Center Rearrangement and Conformational Modulation
of Substrate Orientation |
title_short | Catalysis by the
Non-Heme Iron(II) Histone Demethylase
PHF8 Involves Iron Center Rearrangement and Conformational Modulation
of Substrate Orientation |
title_sort | catalysis by the
non-heme iron(ii) histone demethylase
phf8 involves iron center rearrangement and conformational modulation
of substrate orientation |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6970271/ https://www.ncbi.nlm.nih.gov/pubmed/31976154 http://dx.doi.org/10.1021/acscatal.9b04907 |
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