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Kinetic analysis of acetylation-dependent Pb1 bromodomain-histone interactions

Authors:
Kupitz, C.
Chandrasekaran, R.
Thompson, M.
Published by: Biophysical Chemistry
Date: 7 2008
Volume, Number, Pages: 136 1 7-12

Accession Number: ISI:000257008000002

Abstract:

Stopped-flow fluorescence anisotropy was used to determine the kinetic parameters that define acetylation-dependent bromodomain-histone interactions. Bromodomains are acetyllysine binding motifs found in many chromatin associated proteins. Individual bromodomains were derived from the polybrorno-1 protein, which is a subunit of the PBAF chromatin-remodeling complex that has six tandem bromodomains in the amino-terminal region. The average k(on) and k(off) values for the formation of high-affinity complexes are 275 M-1 s(-1) and 0.41 x 10(-3) s(-1), respectively. The average k(on) and k(off) values for the formation of low-affinity complexes are 119 M-1 s(-1) and 1.42 x 10(-3) s(-1), respectively. Analysis of the on- and off-rates yields acetylation site-dependent equilibrium dissociation constants averaging 1.4 and 12.9 mu M for high- and low-affinity complexes, respectively. This work represents the first examination of kinetic mechanisms of acetylation-dependent bromodomain-histone interactions. (C) 2008 Elsevier B.V. All rights reserved.