The proposed research explores molecular approaches for the study and regulation of aberrant metalloenzyme activity in human disease, focusing on the structural and chemical biology of the arginases and the histone deacetylases (HDACs). The arginases utilize a binuclear Mn(II)-Mn(II) cluster for catalysis, whereas the HDACs utilize a single Zn(II) or Fe(II) ion for catalysis. Unexpectedly, these two enzyme families are evolutionarily related and share a common three-dimensional fold despite insignificant amino acid sequence identity and divergent metal ion stoichiometry and selectivity. Increased arginase activity is implicated in cardiovascular disease, asthma, cancer, and parasitic infections, and increased HDAC activity is found in cancer. Both metalloenzymes are validated targets for structure-based drug design. Furthermore, decreased HDAC8 activity is found in Cornelia de Lange Syndrome (CdLS), a congenital birth defect that occurs in one out of 10,000 births. Accordingly, HDAC8 mutants responsible for decreased activity are potential therapeutic targets for molecular activators that can restore normal biological function. To advance our understanding of structure-function relationships in the arginase-deacetylase fold, and to enable innovative molecular approaches for new disease therapies, we will pursue the following lines of investigation: (1) We will fully characterize the reaction kinetics and determine X-ray crystal structures of HDAC8 mutants identified in CdLS. These studies will provide the first molecular view of compromised HDAC8 catalysis underlying the birth defect. Additionally, we will evaluate the ability of molecular activators to restore normal catalytic function in these mutants, and we will determine X-ray crystal structures of HDAC8-activator complexes to delineate their mode of action. (2) We will determine X-ray crystal structures of mutationally inactivated HDAC8 complexed with peptide and/or protein substrates to understand the structural basis of substrate recognition and catalysis. These structures may reveal how some CdLS HDAC8 mutants perturb the enzyme-substrate interface. (3) Finally, we will establish structure-function relationships for parasitic arginases to guide the design of species-specific arginase inhibitors, and we will explore the ability of selected inhibitors to block polyamine biosynthesis in parasites We will also characterize the arginase-like metalloenzyme from Trypanosoma cruzi, which catalyzes an unusual reaction of histidine catabolism.

Public Health Relevance

This research program explores molecular approaches for the study and regulation of aberrant metalloenzyme activity in human disease, focusing on the structural and chemical biology of arginases and histone deacetylases (HDACs). Increased arginase activity is implicated in cardiovascular disease, asthma, cancer, and parasitic infections;the structure-based design of arginase inhibitors will lead to new therapeutic strategies for treating these diseases. Certain mutations in HDAC8 decrease enzyme activity and cause Cornelia de Lange Syndrome, a congenital birth defect;understanding structure-function relationships in HDAC8 mutants will reveal the molecular basis of the birth defect and guide the development of activators that can rescue catalysis.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
2R01GM049758-20A1
Application #
8691743
Study Section
Macromolecular Structure and Function A Study Section (MSFA)
Program Officer
Smith, Ward
Project Start
1994-05-01
Project End
2018-03-31
Budget Start
2014-04-01
Budget End
2015-03-31
Support Year
20
Fiscal Year
2014
Total Cost
$320,701
Indirect Cost
$111,621
Name
University of Pennsylvania
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
Gantt, Sister M Lucy; Decroos, Christophe; Lee, Matthew S et al. (2016) General Base-General Acid Catalysis in Human Histone Deacetylase 8. Biochemistry 55:820-32
Porter, Nicholas J; Christianson, Nicolas H; Decroos, Christophe et al. (2016) Structural and Functional Influence of the Glycine-Rich Loop G(302)GGGY on the Catalytic Tyrosine of Histone Deacetylase 8. Biochemistry 55:6718-6729
Hai, Yang; Christianson, David W (2016) Histone deacetylase 6 structure and molecular basis of catalysis and inhibition. Nat Chem Biol 12:741-7
Hai, Yang; Christianson, David W (2016) Crystal structures of Leishmania mexicana arginase complexed with α,α-disubstituted boronic amino-acid inhibitors. Acta Crystallogr F Struct Biol Commun 72:300-6
Decroos, Christophe; Christianson, David W (2015) Design, Synthesis, and Evaluation of Polyamine Deacetylase Inhibitors, and High-Resolution Crystal Structures of Their Complexes with Acetylpolyamine Amidohydrolase. Biochemistry 54:4692-703
Hai, Yang; Kerkhoven, Eduard J; Barrett, Michael P et al. (2015) Crystal structure of an arginase-like protein from Trypanosoma brucei that evolved without a binuclear manganese cluster. Biochemistry 54:458-71
Decroos, Christophe; Christianson, Nicolas H; Gullett, Laura E et al. (2015) Biochemical and structural characterization of HDAC8 mutants associated with Cornelia de Lange syndrome spectrum disorders. Biochemistry 54:6501-13
Decroos, Christophe; Clausen, Dane J; Haines, Brandon E et al. (2015) Variable active site loop conformations accommodate the binding of macrocyclic largazole analogues to HDAC8. Biochemistry 54:2126-35
Hai, Yang; Edwards, Jennifer E; Van Zandt, Michael C et al. (2014) Crystal structure of Schistosoma mansoni arginase, a potential drug target for the treatment of schistosomiasis. Biochemistry 53:4671-84
Hai, Yang; Dugery, Reilly Jane; Healy, David et al. (2013) Formiminoglutamase from Trypanosoma cruzi is an arginase-like manganese metalloenzyme. Biochemistry 52:9294-309

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