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 transcriptional activation of eukaryotic target genes. The recent identification 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, little 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 GCN5 family of HATs. Our results suggest that the HAT reaction involves the formation of a ternary complex (histones, acetyl-CoA, and enzyme) where the epsilon-amino group of histone lysine residues directly attacks the bound acetyl-CoA. The acetylation reaction requires deprotonation of the epsilon-amino group prior to nucleophilic attack. Employing site-directed mutagenesis, chemical modification, steady-state, and pH-dependent rate analysis, it is demonstrated that glutamic acid 173 is an essential catalytic residue, acting as a general base catalyst by deprotonating the histone substrate.
Tanner
K G
KG
Department of Biochemistry and Molecular Biology, Oregon Health Sciences University, Portland, Oregon 97201-3098, USA.
Trievel
R C
RC
Kuo
M H
MH
Howard
R M
RM
Berger
S L
SL
Allis
C D
CD
Marmorstein
R
R
Denu
J M
JM
D000105:Acetyl Coenzyme A / Q000378:metabolism; D000123:Acetyltransferases / Q000235:genetics*; D002384:Catalysis; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D005656:Fungal Proteins / Q000378:metabolism; D018698:Glutamic Acid / Q000378:metabolism*; D051548:Histone Acetyltransferases; D006657:Histones / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008956:Models, Chemical; D011494:Protein Kinases / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D015534:Trans-Activators / Q000737:chemistry / Q000378:metabolism*; D014157:Transcription Factors; D050880:p300-CBP Transcription Factors; D000099224:p300-CBP-Associated Factor
D000105:Acetyl Coenzyme A; D000123:Acetyltransferases; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D018698:Glutamic Acid; D051548:Histone Acetyltransferases; D006657:Histones; D011494:Protein Kinases; D029701:Saccharomyces cerevisiae Proteins; D015534:Trans-Activators; D014157:Transcription Factors; D050880:p300-CBP Transcription Factors; D000099224:p300-CBP-Associated Factor; C501392:GCN5 protein, S cerevisiae; C501179:KAT2A protein, human