Structural Studies of Two Related Metallohydrolases: Human Histone Deacetylase 8 and Malarial Arginase
type: CreativeWork
description: Metal-dependent histone deacetylases (HDACs) catalyze the deacetylation of lysine residues in histones and other proteins in eukaryotic cells. Isozyme HDAC8 is perhaps the archetypical member of the class I HDAC family and serves as a paradigm for studying structure-function relationships. We report the structures of HDAC8 complexes in a new crystal form. Comparison of unliganded and liganded structures illustrates ligand-induced conformational changes in the L2-loop that likely accompany substrate binding and catalysis. These structures, along with four D101 variants, support the proposal that D101 is critical for the function of the L2 loop. Additionally, the structure of H143A HDAC8 complexed with an intact substrate confirms the importance of D101 for substrate binding and reveals how Y306 and the active site zinc ion together bind and activate the scissile amide linkage of acetyl-L-lysine. The metal-dependent HDACs adopt an α/β protein fold first identified in rat liver arginase. Despite insignificant overall amino acid sequence identity, these enzymes share a strictly conserved metal-binding site with divergent metal specificity and stoichiometry. HDAC8, originally thought to be a Zn2+-metallohydrolase, exhibits increased activity with Co2+ and Fe2+ based on kcat/KM (Gantt, S. L., Gattis, S. G. & Fierke, C. A. (2006). Biochemistry 45, 6170-6178). Here, we report the first X-ray crystal structures of metallosubstituted HDAC8: Co2+-HDAC8, D101L Co2+-HDAC8, and D101L Fe2+-HDAC8 in complex with the inhibitor M344. The arginase enzyme from Plasmodium falciparum contains a low complexity region insert of ~75 residues within the L2-loop, confirming this protein region to be highly evolvable in proteins which adopt the arginase fold. Kinetics analyses indicate this insertion to be nonessential for catalytic activity. An N-terminally tagged construct shows KM and kcat values similar to the mammalian arginase enzymes and has been used for crystallization. We have solved…
abstract: Metal-dependent histone deacetylases (HDACs) catalyze the deacetylation of lysine residues in histones and other proteins in eukaryotic cells. Isozyme HDAC8 is perhaps the archetypical member of the class I HDAC family and serves as a paradigm for studying structure-function relationships. We report the structures of HDAC8 complexes in a new crystal form. Comparison of unliganded and liganded structures illustrates ligand-induced conformational changes in the L2-loop that likely accompany substrate binding and catalysis. These structures, along with four D101 variants, support the proposal that D101 is critical for the function of the L2 loop. Additionally, the structure of H143A HDAC8 complexed with an intact substrate confirms the importance of D101 for substrate binding and reveals how Y306 and the active site zinc ion together bind and activate the scissile amide linkage of acetyl-L-lysine. The metal-dependent HDACs adopt an α/β protein fold first identified in rat liver arginase. Despite insignificant overall amino acid sequence identity, these enzymes share a strictly conserved metal-binding site with divergent metal specificity and stoichiometry. HDAC8, originally thought to be a Zn2+-metallohydrolase, exhibits increased activity with Co2+ and Fe2+ based on kcat/KM (Gantt, S. L., Gattis, S. G. & Fierke, C. A. (2006). Biochemistry 45, 6170-6178). Here, we report the first X-ray crystal structures of metallosubstituted HDAC8: Co2+-HDAC8, D101L Co2+-HDAC8, and D101L Fe2+-HDAC8 in complex with the inhibitor M344. The arginase enzyme from Plasmodium falciparum contains a low complexity region insert of ~75 residues within the L2-loop, confirming this protein region to be highly evolvable in proteins which adopt the arginase fold. Kinetics analyses indicate this insertion to be nonessential for catalytic activity. An N-terminally tagged construct shows KM and kcat values similar to the mammalian arginase enzymes and has been used for crystallization. We have solved…
author: Dowling, Daniel P.
date published: 2009-12-04
date modified: 2026-07-24T03:45:49Z
provider: University of Pennsylvania
publisher: Networked Digital Library of Theses and Dissertations
type: WebPage
source organization: University of Pennsylvania