Assays for the Determination of Structure and Dynamics of the Interaction of the Chromodomain with Histone Peptides

Abstract

The histone code hypothesis predicts that multiple histone modifications, acting in a combinatorial or sequential fashion on one or multiple histone tails, specify unique downstream functions. To study the specificities and affinities of the interactions of chromodomains with histone tails, several biophysical methods, including nuclear magnetic resonance (NMR) spectroscopy, X-ray crystallography, fluorescence spectroscopy, and isothermal titration calorimetry (ITC), are successfully applied—all are described in the chapter. NMR spectroscopy is best used to screen peptides for interaction with a specific protein module and determine the extent of a minimally structured complex. NMR or X-ray crystallography can subsequently be used to solve the structure of a complex formed between a protein module and a target peptide in order to gain the details of interaction at atomic resolution. Binding affinities and thermodynamics of interaction are best determined by ITC and/or fluorescence spectroscopy. All of these applications are used successfully to study the interactions of chromodomain and bromodomain chromatin binding modules with histone tails.