Transcriptional coactivator protein p300. Kinetic characterization of its histone acetyltransferase activity

Paul R. Thompson, Hisanori Kurooka, Yoshihiro Nakatani, Philip A. Cole

Research output: Contribution to journalArticle

Abstract

The p300/cAMP response element-binding protein-binding protein (CBP) family members include human p300 and cAMP response element-binding protein-binding protein, which are both important transcriptional coactivators and histone acetyltransferases. Although the role of these enzymes in transcriptional regulation has been extensively documented, the molecular mechanisms of p300 and CBP histone acetyltransferase catalysis are poorly understood. Herein, we describe the first detailed kinetic characterization of p300 using full-length purified recombinant enzyme. These studies have employed peptide substrates to systematically examine the substrate specificity requirements and the kinetic mechanism of this enzyme. The importance of nearby positively charged residues in lysine targeting was demonstrated. The strict structural requirement of the lysine side chain was shown. The catalytic mechanism of p300 was shown to follow a ping-pong kinetic pathway and viscosity experiments revealed that product release and/or a conformational change were likely rate-limiting in catalysis. Detailed analysis of the p300 selective inhibitor Lys-CoA showed that it exhibited slow, tight-binding kinetics.

Original languageEnglish (US)
Pages (from-to)33721-33729
Number of pages9
JournalJournal of Biological Chemistry
Volume276
Issue number36
DOIs
StatePublished - Sep 7 2001

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Histone Acetyltransferases
Cyclic AMP Response Element-Binding Protein
Kinetics
Catalysis
Protein Binding
Lysine
Carrier Proteins
Proteins
Enzymes
Substrates
Coenzyme A
Substrate Specificity
Viscosity
Peptides
Experiments

ASJC Scopus subject areas

  • Biochemistry

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Transcriptional coactivator protein p300. Kinetic characterization of its histone acetyltransferase activity. / Thompson, Paul R.; Kurooka, Hisanori; Nakatani, Yoshihiro; Cole, Philip A.

In: Journal of Biological Chemistry, Vol. 276, No. 36, 07.09.2001, p. 33721-33729.

Research output: Contribution to journalArticle

Thompson, Paul R. ; Kurooka, Hisanori ; Nakatani, Yoshihiro ; Cole, Philip A. / Transcriptional coactivator protein p300. Kinetic characterization of its histone acetyltransferase activity. In: Journal of Biological Chemistry. 2001 ; Vol. 276, No. 36. pp. 33721-33729.
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