Analysis of p300/CBP histone acetyltransferase regulation using circular permutation and semisynthesis.
The histone acetyltransferase (HAT) p300/CBP has been shown to undergo autoacetylation on lysines in an apparent regulatory loop that stimulates HAT activity. Here we have developed a strategy to introduce acetyl-Lys at up to six known modification sites in p300/CBP HAT using a combination of circular permutation and expressed protein ligation. We show that these semisynthetic, circularly permuted acetylated proteins retain high affinity for an acetyl-CoA substrate analogue and that HAT activity correlates positively with degree of acetylation. This study provides novel evidence for control of p300/CBP HAT activity by site-specific autoacetylation and outlines a potentially general strategy for using expressed protein ligation and circular permutation to chemically interrogate internal regions of proteins.
Karukurichi
Kannan R
KR
Department of Pharmacology and Molecular Sciences, Johns Hopkins School of Medicine, Baltimore, Maryland 21205, USA.
Wang
Ling
L
Uzasci
Lerna
L
Manlandro
Cara Marie
CM
Wang
Qing
Q
Cole
Philip A
PA
D000105:Acetyl Coenzyme A / Q000378:metabolism; D000107:Acetylation; D000595:Amino Acid Sequence; D008958:Models, Molecular; D008969:Molecular Sequence Data; D050880:p300-CBP Transcription Factors / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism*; D000099224:p300-CBP-Associated Factor
D000105:Acetyl Coenzyme A; D050880:p300-CBP Transcription Factors; D000099224:p300-CBP-Associated Factor