Abstract
The metal-dependent histone deacetylases (HDACs) are critical regulatory enzymes that modulate myriad cellular processes. Implicated in cancer, neurodegenerative diseases, and other clinical disorders, various HDAC isozymes serve as validated drug targets. However, structural similarities among the HDAC isozymes challenge efforts in targeting a single isozyme for therapeutic intervention with an inhibitor. X-ray crystallography remains the premiere technique for studying the chemistry of isozyme-selective inhibition. While crystal structures of many HDAC-inhibitor complexes have been determined, especially with the class I isozyme HDAC8, the study of complexes with large inhibitors is complicated by flexible regions of the protein structure that can hinder crystallization. Here, we outline an approach for the identification of regions in HDAC8 that may hinder crystallization. We also describe protocols for the design and preparation of a truncated HDAC8 construct, HDAC8374, that enabled the successful crystallization and structure determination of the HDAC8-Trapoxin A complex at 1.24Å resolution.