Cofactor Analogues as Active Site Probes in Lysine Acetyltransferases

Lysine
acetyltransferases (KATs, also termed histone acetyltransferases,
HATs) catalyze the acetylation of substrate lysine residues by employing
the cofactor acetyl-coenzyme A (AcCoA), thereby providing a dynamic
control mechanism of protein function. Because of their major involvement
in cell development and homeostasis, small-molecule modulators of
KAT activity are urgently needed to assess their therapeutic potential
and for probing their underlying biology. Recent advances in the field
suggest that targeting the cofactor binding site represents a promising
strategy for identifying potent and selective ligands. Here, we present
the synthesis of two functional cofactor-based chemical probes and
their usage as mechanistic tools in a broadly applicable assay platform.
A fluorescence polarization (FP)-based binding assay was combined
with biolayer interferometry competition analysis and a FP competition
activity immunoassay to enable easy, reliable, and profound evaluation
of ligands that target the KAT cofactor binding site.