Lysine deacetylases (KDACs) catalyze the hydrolysis of e-N-acetyl lysine residues in proteins to generate an unmodified lysine residue and acetate. KDACs play a critical role in transcriptional regulation, cell cycle progression, and developmental events. Perturbations in this process have been linked to diseases, including cancer and inflammation. While thousands of acetylated proteins have been identified, little information is known concerning the specificity profiles of KDACs. Our aim is to identify the essential elements involved in molecular recognition of substrate proteins by KDACs. Assays performed using standard commercial reagents and acetylated peptides as substrates suggest that KDACs and their variants have unique specificity profiles. To better understand this in a biologically relevant context, we have engineered model proteins which more closely resemble natural KDAC substrates. Each is a fixed scaffold that contains a single acetylated lysine in a defined variable sequence and secondary structure. Binding and activity for KDAC-substrate pairs are used to determine KDAC substrate preference and specificity. Preliminary data with a range of these model substrates suggests substantial differences from those previously reported with commercial assays, and kinetic parameters that are more likely to be biologically relevant. We have combined these results with structural data, molecular modeling, and mutagenesis of KDACs to gain insights into the mechanism of KDAC substrate recognition.

Substrate recognition in lysine deacetylases

248th National Meeting (2014)

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