Advances in Label-Free Screening Approaches for Studying Histone Acetyltransferases

Histone acetyltransferases (HATs) catalyze the transfer of an acetyl group from an acetyl-coenzyme A donor molecule to specific lysine residues within proteins. The acetylation state of proteins, particularly histones, is known to modulate their intermolecular binding properties and control various cellular processes, most notably transcriptional activation. In addition, deregulation of HAT activity has been linked to the development of a number of cancers; therefore, compounds that affect these enzymes have strong potential as therapeutic agents. The research presented here demonstrates three label-free HAT screening approaches, all based on the fast and direct measurement of one or more substrate-product pairs by high-throughput mass spectrometry techniques. The first approach involves monitoring all possible acetylation states of a peptide concurrently to measure HAT activity. The second approach measures acetylation reactions, on both peptides and whole protein substrates, via direct detection of the acetyl-coenzyme A cosubstrate and coenzyme A coproduct. Lastly, the authors demonstrate the ability to monitor directly the acetylation state of whole histone proteins in the same high-throughput manner using time-of-flight mass spectrometry. The generation of compound-mediated inhibition data using each of these techniques establishes mass spectrometry as a versatile, label-free, and biologically relevant screening approach to this challenging target class. (Journal of Biomolecular Screening. 2011:1186-1195)