THE HISTONE ACETYLTRANSFERASE ACTIVITY OF HUMAN GCN5 AND PCAF IS STABILIZED BY COENZYMES

Here we report that PCAF and human GCN5, two related type A histone ac etyltransferases, are unstable enzymes that under the commonly used as say conditions are rapidly and irreversibly inactivated, Ire addition, we report that free histone H1, although not acetylated in vivo, is a preferred and convenient in vitro substrate for the study of PCAF, hu man GCN5, and possibly other type A histone acetyltransferases, Using either histone H1 or histone H3 as substrates, we find that preincubat ion with either acetyl-CoA or CoA stabilizes the acetyltransferase act ivities of PCAF, human GCN5 and an enzymatically active PCAF deletion mutant containing the C-terminal half of the protein. The stabilizatio n requires the continuous presence of coenzyme, suggesting that the ac etyltransferase-coenzyme complexes are stable, while the isolated apoe nzymes are not, Human GCN5 and the N-terminal deletion mutant of PCAF are stabilized equally well by preincubation with either CoA or acetyl -CoA, while intact PCAF is better stabilized by acetyl-CoA than by CoA , Intact PCAF, but not the N-terminal truncation mutant or human GCN5, is auto-acetylated. These findings raise the possibility that the int racellular concentrations of the coenzymes affect the stability and th erefore the nuclear activity of these acetyltransferases.