Unraveling the role of exit tunnel residues of histone deacetylase‐8 in catalysis

Histone Deacetylases (HDACs) are an important class of enzymes which are involved in epigenetic regulations and they have been identified as high priority targets for treatment of cancers. The structural data reveals that apart from the active site pocket, HDAC‐8 harbors an internal cavity through which the acetate product dissociates from the enzyme. To probe the role of the amino acid residues at the “exit port” of the above cavity, we performed site directed mutagenesis of Y18, Y20 and H42 to Ala. Although these mutations did not influence the binding affinity of the substrate, they drastically impaired the catalytic rate of HDAC‐ 8. In case of Y18A mutation, the magnitude of activation by our recently identified activator (TM‐2‐51) was far more pronounced than the wild‐type enzyme. Our circular dichroism studies indicated a change in the secondary structure of the enzyme in case of Y18A mutation (but not in Y20A and H42A mutations) which was manifested in an increased thermal stability of the mutant. Our experimental data reveals that there is a long range communication between the exit tunnel residues and the active site pocket of HDAC‐8, and such communication presumably regulates the overall catalytic feature of the enzyme Research supported by NIH grants CA113746 and CA132034 to DKS and NIH COBRE grant NCRR‐P20‐RR15566 to GC.