General Base–General Acid Catalysis in Human Histone Deacetylase 8
Abstract
Histone deacetylases (HDACs) regulate cellular processes such as differentiation and apoptosis and are targeted by anticancer therapeutics in development and in the clinic. HDAC8 is a metal-dependent class I HDAC and is proposed to use a general acid–base catalytic pair in the mechanism of amide bond hydrolysis. Here, we report site-directed mutagenesis and enzymological measurements to elucidate the catalytic mechanism of HDAC8. Specifically, we focus on the catalytic function of Y306 and the histidine-aspartate dyads H142-D176 and H143-D183. Additionally, we report X-ray crystal structures of four representative HDAC8 mutants: D176N, D176N/Y306F, D176A/Y306F, and H142A/Y306F. These structures provide a useful framework for understanding enzymological measurements. We report the pH dependence of kcat/KM for wild-type Co(II)-HDAC8 is bell-shaped with two pKa values of 7.4 and 10.0. The upper pKa reflects the ionization of the metal-bound water molecule and shifts to 9.1 in Zn(II)-HDAC8. The H142A mutant has activity 230-fold lower than that of wild-type HDAC8, but the pKa1 value is not altered. Y306F HDAC8 is 150-fold less active than the wild-type enzyme; crystal structures show that Y306 hydrogen bonds with the zinc-bound substrate carbonyl, poised for transition state stabilization. The H143A and H142A/H143A mutants exhibit activity that is >80000-fold lower than thatmore »
- Authors:
-
- Univ. of Michigan, Ann Arbor, MI (United States); Franciscan Univ. of Steubenville, OH (United States)
- Univ. of Pennsylvania, Philadelphia, PA (United States); Aix Marseille Univ., Centrale Marseille (France)
- Univ. of Pennsylvania, Philadelphia, PA (United States)
- Univ. of Pennsylvania, Philadelphia, PA (United States); Harvard Univ., Cambridge, MA (United States)
- Univ. of Pennsylvania, Philadelphia, PA (United States); Univ. of California, San Francisco, CA (United States)
- Univ. of Michigan, Ann Arbor, MI (United States)
- Publication Date:
- Research Org.:
- Argonne National Laboratory (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
- Sponsoring Org.:
- National Institutes of Health (NIH)
- OSTI Identifier:
- 1238290
- Grant/Contract Number:
- GM40602; GM49758
- Resource Type:
- Journal Article: Accepted Manuscript
- Journal Name:
- Biochemistry
- Additional Journal Information:
- Journal Volume: 55; Journal Issue: 5; Journal ID: ISSN 0006-2960
- Publisher:
- American Chemical Society (ACS)
- Country of Publication:
- United States
- Language:
- ENGLISH
- Subject:
- 59 BASIC BIOLOGICAL SCIENCES; Peptides and proteins; Monomers; Molecules; Catalysts; Noncovalent interactions
Citation Formats
Gantt, Sister M. Lucy, Decroos, Christophe, Lee, Matthew S., Gullett, Laura E., Bowman, Christine M., Christianson, David W., and Fierke, Carol A. General Base–General Acid Catalysis in Human Histone Deacetylase 8. United States: N. p., 2016.
Web. doi:10.1021/acs.biochem.5b01327.
Gantt, Sister M. Lucy, Decroos, Christophe, Lee, Matthew S., Gullett, Laura E., Bowman, Christine M., Christianson, David W., & Fierke, Carol A. General Base–General Acid Catalysis in Human Histone Deacetylase 8. United States. https://doi.org/10.1021/acs.biochem.5b01327
Gantt, Sister M. Lucy, Decroos, Christophe, Lee, Matthew S., Gullett, Laura E., Bowman, Christine M., Christianson, David W., and Fierke, Carol A. 2016.
"General Base–General Acid Catalysis in Human Histone Deacetylase 8". United States. https://doi.org/10.1021/acs.biochem.5b01327. https://www.osti.gov/servlets/purl/1238290.
@article{osti_1238290,
title = {General Base–General Acid Catalysis in Human Histone Deacetylase 8},
author = {Gantt, Sister M. Lucy and Decroos, Christophe and Lee, Matthew S. and Gullett, Laura E. and Bowman, Christine M. and Christianson, David W. and Fierke, Carol A.},
abstractNote = {Histone deacetylases (HDACs) regulate cellular processes such as differentiation and apoptosis and are targeted by anticancer therapeutics in development and in the clinic. HDAC8 is a metal-dependent class I HDAC and is proposed to use a general acid–base catalytic pair in the mechanism of amide bond hydrolysis. Here, we report site-directed mutagenesis and enzymological measurements to elucidate the catalytic mechanism of HDAC8. Specifically, we focus on the catalytic function of Y306 and the histidine-aspartate dyads H142-D176 and H143-D183. Additionally, we report X-ray crystal structures of four representative HDAC8 mutants: D176N, D176N/Y306F, D176A/Y306F, and H142A/Y306F. These structures provide a useful framework for understanding enzymological measurements. We report the pH dependence of kcat/KM for wild-type Co(II)-HDAC8 is bell-shaped with two pKa values of 7.4 and 10.0. The upper pKa reflects the ionization of the metal-bound water molecule and shifts to 9.1 in Zn(II)-HDAC8. The H142A mutant has activity 230-fold lower than that of wild-type HDAC8, but the pKa1 value is not altered. Y306F HDAC8 is 150-fold less active than the wild-type enzyme; crystal structures show that Y306 hydrogen bonds with the zinc-bound substrate carbonyl, poised for transition state stabilization. The H143A and H142A/H143A mutants exhibit activity that is >80000-fold lower than that of wild-type HDAC8; the buried D176N and D176A mutants have significant catalytic effects, with more subtle effects caused by D183N and D183A. These enzymological and structural studies strongly suggest that H143 functions as a single general base–general acid catalyst, while H142 remains positively charged and serves as an electrostatic catalyst for transition state stabilization.},
doi = {10.1021/acs.biochem.5b01327},
url = {https://www.osti.gov/biblio/1238290},
journal = {Biochemistry},
issn = {0006-2960},
number = 5,
volume = 55,
place = {United States},
year = {Mon Jan 25 00:00:00 EST 2016},
month = {Mon Jan 25 00:00:00 EST 2016}
}
Web of Science
Works referenced in this record:
Histone deacetylases and cancer
journal, August 2007
- Glozak, M. A.; Seto, E.
- Oncogene, Vol. 26, Issue 37
Histone deacetylase inhibitors: molecular mechanisms of action
journal, August 2007
- Xu, W. S.; Parmigiani, R. B.; Marks, P. A.
- Oncogene, Vol. 26, Issue 37
HDAC inhibitor-based therapies: Can we interpret the code?
journal, October 2012
- New, Maria; Olzscha, Heidi; La Thangue, Nicholas B.
- Molecular Oncology, Vol. 6, Issue 6
Metabolic reprogramming by class I and II histone deacetylases
journal, January 2013
- Mihaylova, Maria M.; Shaw, Reuben J.
- Trends in Endocrinology & Metabolism, Vol. 24, Issue 1
Acetylation and Methylation of Histones and Their Possible role in the Regulation of rna Synthesis
journal, May 1964
- Allfrey, V. G.; Faulkner, R.; Mirsky, A. E.
- Proceedings of the National Academy of Sciences, Vol. 51, Issue 5
Lysine acetylation and the bromodomain: a new partnership for signaling
journal, January 2004
- Yang, Xiang-Jiao
- BioEssays, Vol. 26, Issue 10
Acetylation and deacetylation of non-histone proteins
journal, December 2005
- Glozak, Michele A.; Sengupta, Nilanjan; Zhang, Xiaohong
- Gene, Vol. 363
The Biochemistry of Sirtuins
journal, June 2006
- Sauve, Anthony A.; Wolberger, Cynthia; Schramm, Vern L.
- Annual Review of Biochemistry, Vol. 75, Issue 1
Molecular Evolution of the Histone Deacetylase Family: Functional Implications of Phylogenetic Analysis
journal, April 2004
- Gregoretti, IvanV; Lee, Yun-Mi; Goodson, Holly V.
- Journal of Molecular Biology, Vol. 338, Issue 1
Catalytic Activity and Inhibition of Human Histone Deacetylase 8 Is Dependent on the Identity of the Active Site Metal Ion †
journal, May 2006
- Gantt, Stephanie L.; Gattis, Samuel G.; Fierke, Carol A.
- Biochemistry, Vol. 45, Issue 19
Vorinostat: a new oral histone deacetylase inhibitor approved for cutaneous T-cell lymphoma
journal, June 2007
- Duvic, Madeleine; Vu, Jenny
- Expert Opinion on Investigational Drugs, Vol. 16, Issue 7
Dimethyl sulfoxide to vorinostat: development of this histone deacetylase inhibitor as an anticancer drug
journal, January 2007
- Marks, Paul A.; Breslow, Ronald
- Nature Biotechnology, Vol. 25, Issue 1
FDA Approval Summary: Vorinostat for Treatment of Advanced Primary Cutaneous T-Cell Lymphoma
journal, October 2007
- Mann, B. S.; Johnson, J. R.; Cohen, M. H.
- The Oncologist, Vol. 12, Issue 10
FDA Approval: Belinostat for the Treatment of Patients with Relapsed or Refractory Peripheral T-cell Lymphoma
journal, March 2015
- Lee, Hyon-Zu; Kwitkowski, Virginia E.; Del Valle, Pedro L.
- Clinical Cancer Research, Vol. 21, Issue 12
Panobinostat for the Treatment of Multiple Myeloma
journal, September 2015
- Laubach, Jacob P.; Moreau, Philippe; San-Miguel, Jesús F.
- Clinical Cancer Research, Vol. 21, Issue 21
Therapeutic application of histone deacetylase inhibitors for central nervous system disorders
journal, October 2008
- Kazantsev, Aleksey G.; Thompson, Leslie M.
- Nature Reviews Drug Discovery, Vol. 7, Issue 10
Epigenetic targeting of histone deacetylase: Therapeutic potential in Parkinson's disease?
journal, October 2013
- Harrison, Ian F.; Dexter, David T.
- Pharmacology & Therapeutics, Vol. 140, Issue 1
Crystal structure of a eukaryotic zinc-dependent histone deacetylase, human HDAC8, complexed with a hydroxamic acid inhibitor
journal, October 2004
- Vannini, A.; Volpari, C.; Filocamo, G.
- Proceedings of the National Academy of Sciences, Vol. 101, Issue 42
Structural Snapshots of Human HDAC8 Provide Insights into the Class I Histone Deacetylases
journal, July 2004
- Somoza, John R.; Skene, Robert J.; Katz, Bradley A.
- Structure, Vol. 12, Issue 7
Substrate binding to histone deacetylases as shown by the crystal structure of the HDAC8–substrate complex
journal, August 2007
- Vannini, Alessandro; Volpari, Cinzia; Gallinari, Paola
- EMBO reports, Vol. 8, Issue 9
Zinc hydrolases: the mechanisms of zinc-dependent deacetylases
journal, January 2005
- Hernick, Marcy; Fierke, Carol A.
- Archives of Biochemistry and Biophysics, Vol. 433, Issue 1
Structural Studies of Human Histone Deacetylase 8 and Its Site-Specific Variants Complexed with Substrate and Inhibitors † , ‡
journal, December 2008
- Dowling, Daniel P.; Gantt, Stephanie L.; Gattis, Samuel G.
- Biochemistry, Vol. 47, Issue 51
Activation and Inhibition of Histone Deacetylase 8 by Monovalent Cations
journal, December 2009
- Gantt, Stephanie L.; Joseph, Caleb G.; Fierke, Carol A.
- Journal of Biological Chemistry, Vol. 285, Issue 9
Structures of Metal-Substituted Human Histone Deacetylase 8 Provide Mechanistic Inferences on Biological Function,
journal, June 2010
- Dowling, Daniel P.; Gattis, Samuel G.; Fierke, Carol A.
- Biochemistry, Vol. 49, Issue 24
Structural Basis of the Antiproliferative Activity of Largazole, a Depsipeptide Inhibitor of the Histone Deacetylases
journal, August 2011
- Cole, Kathryn E.; Dowling, Daniel P.; Boone, Matthew A.
- Journal of the American Chemical Society, Vol. 133, Issue 32
Compromised Structure and Function of HDAC8 Mutants Identified in Cornelia de Lange Syndrome Spectrum Disorders
journal, July 2014
- Decroos, Christophe; Bowman, Christine M.; Moser, Joe-Ann S.
- ACS Chemical Biology, Vol. 9, Issue 9
Variable Active Site Loop Conformations Accommodate the Binding of Macrocyclic Largazole Analogues to HDAC8
journal, March 2015
- Decroos, Christophe; Clausen, Dane J.; Haines, Brandon E.
- Biochemistry, Vol. 54, Issue 12
Structures of a histone deacetylase homologue bound to the TSA and SAHA inhibitors
journal, September 1999
- Finnin, Michael S.; Donigian, Jill R.; Cohen, Alona
- Nature, Vol. 401, Issue 6749
On the importance of orientation in general base catalysis by carboxylate
journal, June 1981
- Gandour, Richard David
- Bioorganic Chemistry, Vol. 10, Issue 2
Stereoelectronic effects at carboxylate: a syn-oriented model for the histidine-aspartate couple in enzymes
journal, August 1988
- Zimmerman, Steven C.; Cramer, Katherine D.
- Journal of the American Chemical Society, Vol. 110, Issue 17
Carboxylate-histidine-zinc interactions in protein structure and function
journal, August 1989
- Christianson, David W.; Alexander, Richard S.
- Journal of the American Chemical Society, Vol. 111, Issue 16
Conformational differences between various myoglobin ligated states as monitored by proton NMR spectroscopy
journal, October 1984
- Bradbury, J. Howard; Carver, John A.
- Biochemistry, Vol. 23, Issue 21
Assignment of proton NMR resonances of histidine and other aromatic residues in met-, cyano-, oxy-, and (carbon monoxy)myoglobins
journal, October 1984
- Carver, John A.; Bradbury, J. Howard
- Biochemistry, Vol. 23, Issue 21
The pKa values of two histidine residues in human haemoglobin, the Bohr effect, and the dipole moments of α-helices
journal, June 1985
- Perutz, Max F.; Gronenborn, Angela M.; Clore, G. Marius
- Journal of Molecular Biology, Vol. 183, Issue 3
Crystal Structure of a Bacterial Class 2 Histone Deacetylase Homologue
journal, November 2005
- Nielsen, Tine Kragh; Hildmann, Christian; Dickmanns, Achim
- Journal of Molecular Biology, Vol. 354, Issue 1
Probing the elusive catalytic activity of vertebrate class IIa histone deacetylases
journal, March 2008
- Jones, Philip; Altamura, Sergio; De Francesco, Raffaele
- Bioorganic & Medicinal Chemistry Letters, Vol. 18, Issue 6
Human HDAC7 Harbors a Class IIa Histone Deacetylase-specific Zinc Binding Motif and Cryptic Deacetylase Activity
journal, April 2008
- Schuetz, Anja; Min, Jinrong; Allali-Hassani, Abdellah
- Journal of Biological Chemistry, Vol. 283, Issue 17
Structural basis of the action of thermolysin and related zinc peptidases
journal, September 1988
- Matthews, Brian W.
- Accounts of Chemical Research, Vol. 21, Issue 9
Carboxypeptidase A
journal, February 1989
- Christianson, David W.; Lipscomb, William N.
- Accounts of Chemical Research, Vol. 22, Issue 2
Noncomplexing Tertiary Amines as “Better” Buffers Covering the Range of pH 3−11. Temperature Dependence of Their Acid Dissociation Constants
journal, August 1999
- Kandegedara, Ashoka; Rorabacher, David B.
- Analytical Chemistry, Vol. 71, Issue 15
Kinetics and Comparative Reactivity of Human Class I and Class IIb Histone Deacetylases
journal, August 2004
- Schultz, Brian E.; Misialek, Shawn; Wu, Jiansheng
- Biochemistry, Vol. 43, Issue 34
[20] Processing of X-ray diffraction data collected in oscillation mode
book, January 1997
- Otwinowski, Zbyszek; Minor, Wladek
- Macromolecular Crystallography Part A, 307-326
PHENIX: a comprehensive Python-based system for macromolecular structure solution
journal, January 2010
- Adams, Paul D.; Afonine, Pavel V.; Bunkóczi, Gábor
- Acta Crystallographica Section D Biological Crystallography, Vol. 66, Issue 2, p. 213-221
Features and development of Coot
journal, March 2010
- Emsley, P.; Lohkamp, B.; Scott, W. G.
- Acta Crystallographica Section D Biological Crystallography, Vol. 66, Issue 4
Pyrrole Hydrogen Ionization of Imidazole Derivatives in Metal Ion Complexes and Carbonic Anhydrase
journal, November 1974
- Martin, R. B.
- Proceedings of the National Academy of Sciences, Vol. 71, Issue 11
Structure and activity of enzymes that remove histone modifications
journal, December 2005
- Holbert, Marc A.; Marmorstein, Ronen
- Current Opinion in Structural Biology, Vol. 15, Issue 6
Structure, mechanism, and inhibition of histone deacetylases and related metalloenzymes
journal, December 2011
- Lombardi, Patrick M.; Cole, Kathryn E.; Dowling, Daniel P.
- Current Opinion in Structural Biology, Vol. 21, Issue 6
Members of the histone deacetylase superfamily differ in substrate specificity towards small synthetic substrates
journal, November 2004
- Riester, Daniel; Wegener, Dennis; Hildmann, Christian
- Biochemical and Biophysical Research Communications, Vol. 324, Issue 3
Substrate and inhibitor specificity of class 1 and class 2 histone deacetylases
journal, June 2006
- Hildmann, Christian; Wegener, Dennis; Riester, Daniel
- Journal of Biotechnology, Vol. 124, Issue 1
UDP-3-O-((R)-3-hydroxymyristoyl)-N-acetylglucosamine Deacetylase Functions through a General Acid-Base Catalyst Pair Mechanism
journal, April 2005
- Hernick, Marcy; Gennadios, Heather A.; Whittington, Douglas A.
- Journal of Biological Chemistry, Vol. 280, Issue 17
Crystal structure of LpxC, a zinc-dependent deacetylase essential for endotoxin biosynthesis
journal, June 2003
- Whittington, Douglas A.; Rusche, Kristin M.; Shin, Hyunshun
- Proceedings of the National Academy of Sciences, Vol. 100, Issue 14
Binding to thermolysin of phenolate-containing inhibitors necessitates a revised mechanism of catalysis
journal, August 1994
- Mock, W. L.; Aksamawati, M.
- Biochemical Journal, Vol. 302, Issue 1
Catalytic Mechanism and Molecular Recognition of E . coli UDP-3- O -( R -3-Hydroxymyristoyl)- N -acetylglucosamine Deacetylase Probed by Mutagenesis †
journal, December 2006
- Hernick, Marcy; Fierke, Carol A.
- Biochemistry, Vol. 45, Issue 51
Differences in transition state stabilization between thermolysin (EC 3.4.24.27) and neprilysin (EC 3.4.24.11)
journal, November 1998
- Marie-Claire, Cynthia; Ruffet, Emmanuel; Tiraboschi, Gilles
- FEBS Letters, Vol. 438, Issue 3
A Correlation of Reaction Rates
journal, January 1955
- Hammond, George S.
- Journal of the American Chemical Society, Vol. 77, Issue 2
Unexpected Deacetylation Mechanism Suggested by a Density Functional Theory QM/MM Study of Histone-Deacetylase-Like Protein
journal, April 2006
- Corminboeuf, Clémence; Hu, Po; Tuckerman, Mark E.
- Journal of the American Chemical Society, Vol. 128, Issue 14
A Proton-Shuttle Reaction Mechanism for Histone Deacetylase 8 and the Catalytic Role of Metal Ions
journal, July 2010
- Wu, Ruibo; Wang, Shenglong; Zhou, Nengjie
- Journal of the American Chemical Society, Vol. 132, Issue 27
Inhibition and Mechanism of HDAC8 Revisited
journal, August 2014
- Chen, Kai; Zhang, Xiaoxiao; Wu, Yun-Dong
- Journal of the American Chemical Society, Vol. 136, Issue 33
Works referencing / citing this record:
Structure–activity relationships of hydroxamate-based histone deacetylase-8 inhibitors: reality behind anticancer drug discovery
journal, December 2017
- Amin, Sk Abdul; Adhikari, Nilanjan; Jha, Tarun
- Future Medicinal Chemistry, Vol. 9, Issue 18
Regulation of Histone Deacetylase 3 by Metal Cations and 10-Hydroxy-2E-DecenoicAcid: Possible Molecular Epigenetic Mechanisms of Queen-Worker Bee Differentiation
posted_content, September 2018
- Polsinelli, Gregory A.; Yu, Hongwei D.
- bioRxiv
HDAC3 and HDAC8 are required for cilia assembly and elongation
journal, July 2019
- Park, Seon-ah; Yoo, Hyunjeong; Seol, Jae Hong
- Biology Open, Vol. 8, Issue 8
Histone deacetylase 6 structure and molecular basis of catalysis and inhibition
journal, July 2016
- Hai, Yang; Christianson, David W.
- Nature Chemical Biology, Vol. 12, Issue 9
Formation of Unstable and very Reactive Chemical Species Catalyzed by Metalloenzymes: A Mechanistic Overview
journal, July 2019
- Fernandes, Henrique S.; Teixeira, Carla S. Silva; Sousa, Sérgio F.
- Molecules, Vol. 24, Issue 13
Factors governing when a metal-bound water is deprotonated in proteins
journal, January 2018
- Grauffel, Cédric; Lim, Carmay
- Physical Chemistry Chemical Physics, Vol. 20, Issue 47
Regulation of histone deacetylase 3 by metal cations and 10-hydroxy-2E-decenoic acid: Possible epigenetic mechanisms of queen-worker bee differentiation
journal, December 2018
- Polsinelli, Gregory A.; Yu, Hongwei D.
- PLOS ONE, Vol. 13, Issue 12
Histone deacetylase 6 structure and molecular basis of catalysis and inhibition
journal, July 2016
- Hai, Yang; Christianson, David W.
- Nature Chemical Biology, Vol. 12, Issue 9
HDAC3 and HDAC8 are required for cilia assembly and elongation
journal, July 2019
- Park, Seon-ah; Yoo, Hyunjeong; Seol, Jae Hong
- Biology Open, Vol. 8, Issue 8
Regulation of histone deacetylase 3 by metal cations and 10-hydroxy-2E-decenoic acid: Possible epigenetic mechanisms of queen-worker bee differentiation
journal, December 2018
- Polsinelli, Gregory A.; Yu, Hongwei D.
- PLOS ONE, Vol. 13, Issue 12
Selective Inhibition of HDAC1 by Macrocyclic Polypeptide for the Treatment of Glioblastoma: A Binding Mechanistic Analysis Based on Molecular Dynamics
journal, March 2020
- Zhang, Yang; Fu, Tingting; Ren, Yuxiang
- Frontiers in Molecular Biosciences, Vol. 7
Formation of Unstable and very Reactive Chemical Species Catalyzed by Metalloenzymes: A Mechanistic Overview
journal, July 2019
- Fernandes, Henrique S.; Teixeira, Carla S. Silva; Sousa, Sérgio F.
- Molecules, Vol. 24, Issue 13