Structural Basis of HDAC Substrate Specificity and Inhibition

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Project Summary/Abstract Reversible protein lysine acetylation is a fundamental posttranslational modification observed in histone and non-histone proteins. Lysine acetylation can alter protein-protein and protein-DNA interactions, protein stability, and enzyme activation/deactivation. Among the major regulators of lysine acetylation is the histone deacetylase (HDAC) family. Of the 18 known human HDACs, 11 are metal-dependent hydrolases related to the acetylpolyamine amidohydrolases (APAHs). The HDACs contribute to the regulation of gene expression and many other critical cellular processes. Notably, abnormal lysine acetylation is observed in multiple human disorders, including cancer; thus HDACs are a validated drug target. Despite their critical biological functions and clinical roles as drug targets, little is known about the molecular basis for HDAC substrate specificity and inhibition. This is particularly the case for HDACs 10 and 11, which are the least well characterized of the metal-dependent HDACs. Our preliminary studies coupled with phylogenetic comparisons suggest that HDACs 10 and 11 may function as dual acetyllysine and acetylpolyamine deacetylases with unique substrate binding site architectures. However, how HDAC10 and 11 could accommodate small polyamine substrates as well as large protein substrates containing sissile acetyllysine moieties is unclear. In addition, while classic HDAC inhibitors such as SAHA are known to inhibit HDACs 10 and 11, the molecular basis for this inhibition is unknown as no HDAC10-inhibitor or HDAC11-inibitor complex structures are available. In fact, no structure of HDAC11 is available, despite the fact that HDAC11 represents a unique class of HDAC due to its limited sequence identity with other HDACs. We propose to study structure-function relationships for HDACs 10 and 11 to establish a molecular foundation for understanding substrate recognition, catalysis, and inhibition. Due to a lack of structural and mechanistic studies focusing on HDACs 10 and 11, we are currently unequipped to design HDAC isozyme-specific inhibitors. I propose to study the molecular mechanisms of HDAC substrate recognition and inhibition by (1) exploring the structural basis of HDAC10 substrate specificity; (2) defining the structural basis of HDAC10 inhibition; and (3) determining structure-function relationships for HDAC11. As mentioned above, aberrant lysine acetylation is a hallmark of certain cancers and other human disorders; therefore HDACs are critical drug targets. Currently, four broad-specificity HDAC inhibitors are FDA-approved for cancer chemotherapy, but isozyme-specific HDAC inhibitors are mostly unavailable. Our studies aim to better understand the structure and function of poorly characterized HDACs with the goal of facilitating the design of specific HDAC inhibitors for use in human disorders, particularly cancer.