In order to establish a base of structural knowledge concerning the structure, function, and regulation of the enigmatic family of manganese metalloenzymes, we have selected the metallohydrolase rat liver arginase as the paradigm for protein engineering and rational ligand design experiments. This enzyme is extraordinary in that it contains a binuclear, spin-coupled manganese cluster in its active site which is implicated in the chemistry of arginine hydrolysis. Surprisingly, this metal cluster is also implicated in the chemistry of hydrogen peroxide disproportionation (i.e., the """"""""catalase"""""""" reaction). In addition to yielding the first structure of a mammalian urea cycle enzyme (where cytosolic arginase catalyzes the hydrolysis of arginine into ornithine plus urea), structural studies of arginase will complement studies of its role in macrophage biology. The activities of nitric oxide synthase and arginase in the macrophage are reciprocally coordinated in order to modulate NO-dependent cytotoxicity. In this role, arginase is implicated as an inhibitor of liver transplant rejection as well as a tumoricidal agent. We have achieved the goals outlined in the original grant proposal: we have crystallized and determined the three-dimensional structure of native rat liver arginase at 2.1 A resolution. Additionally, we have crystallized recombinant wild-type arginase and several single-point variants expressed in E.coli (a T7 expression system allows for the production of large amounts of wild-type and variant arginases); structure determinations and refinements of these variant structures are currently in progress. We have synthesized and assayed hydroxamate-based inhibitors of arginase, and we have synthesized a boronic acid-based inhibitor which may bind to the enzyme as a transition state analogue. Herein, we request support to continue X-ray crystallographic studies of arginase, its site-specific variants and its inhibitor complexes. Additionally, using arginase as the prototype, we will continue the development of a structure-based rationale for the design and synthesis and potent metalloenzyme inhibitors that target bimetallic clusters.
| Porter, Nicholas J; Wagner, Florence F; Christianson, David W (2018) Entropy as a Driver of Selectivity for Inhibitor Binding to Histone Deacetylase 6. Biochemistry 57:3916-3924 |
| Bhatia, Sanil; Krieger, Viktoria; Groll, Michael et al. (2018) Discovery of the First-in-Class Dual Histone Deacetylase-Proteasome Inhibitor. J Med Chem 61:10299-10309 |
| Porter, Nicholas J; Osko, Jeremy D; Diedrich, Daniela et al. (2018) Histone Deacetylase 6-Selective Inhibitors and the Influence of Capping Groups on Hydroxamate-Zinc Denticity. J Med Chem 61:8054-8060 |
| Mackwitz, Marcel K W; Hamacher, Alexandra; Osko, Jeremy D et al. (2018) Multicomponent Synthesis and Binding Mode of Imidazo[1,2- a]pyridine-Capped Selective HDAC6 Inhibitors. Org Lett 20:3255-3258 |
| Shinsky, Stephen A; Christianson, David W (2018) Polyamine Deacetylase Structure and Catalysis: Prokaryotic Acetylpolyamine Amidohydrolase and Eukaryotic HDAC10. Biochemistry 57:3105-3114 |
| Porter, Nicholas J; Mahendran, Adaickapillai; Breslow, Ronald et al. (2017) Unusual zinc-binding mode of HDAC6-selective hydroxamate inhibitors. Proc Natl Acad Sci U S A 114:13459-13464 |
| Hai, Yang; Shinsky, Stephen A; Porter, Nicholas J et al. (2017) Histone deacetylase 10 structure and molecular function as a polyamine deacetylase. Nat Commun 8:15368 |
| Bitler, Benjamin G; Wu, Shuai; Park, Pyoung Hwa et al. (2017) ARID1A-mutated ovarian cancers depend on HDAC6 activity. Nat Cell Biol 19:962-973 |
| Porter, Nicholas J; Christianson, David W (2017) Binding of the Microbial Cyclic Tetrapeptide Trapoxin A to the Class I Histone Deacetylase HDAC8. ACS Chem Biol 12:2281-2286 |
| 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 |
Showing the most recent 10 out of 53 publications
