Catalytic mechanism and function of invariant glutamic acid 173 from the histone acetyltransferase GCN5 transcriptional coactivator

Within chromatin, reversible acetylation of core histones is critical for t ranscriptional activation of eukaryotic target genes. The recent identifica tion of intrinsic histone acetyltransferase (HAT) catalytic activity from a number of transcriptional co-activators (including yeast GCN5, p300/CBP, P /CAF, and TAFII250), has underscored the importance of protein acetylation in transcriptional control. The GCN5 family is the prototype for a diverse group of at least four distinct human HATs families. Although there is now a clear link between in vivo HAT catalytic activity and gene activation, li ttle is known about the molecular mechanisms of histone acetylation. Herein , we report the first detailed biochemical study that probes the catalytic mechanism and the function of invariant glutamic acid 173 within the GCNB f amily of HATs, Our results suggest that the HAT reaction involves the forma tion of a ternary complex (histones, acetyl-CoA and enzyme) where the E-ami no group of histone lysine residues directly attacks the bound acetyl-CoA, The acetylation reaction requires deprotonation of the E-amino group prior to nucleophilic attack. Employing site-directed mutagenesis, chemical modif ication, steady-state, and pH-dependent rate analysis, it is demonstrated t hat glutamic acid 173 is an essential catalytic residue, acting as a genera l base catalyst by deprotonating the histone substrate.