Nine distinct genes in human encode CHD polypeptides containing a pair of tandem chromodomains and a SNF2-type ATPase/Helicase domain. CHD proteins are implicated in chromatin remodelling for epigenetic regulation of gene expression. Recent studies have implicated CHD proteins in diverse roles such as facilitating transcription by RNA Polymerase II, assisting transcription co-repressor complexes, and regulating differentiation. They also have been implicated in diseases such as dermatomyositis, hodgkin's lymphoma, and neuroblastoma (linked with CHD3, CHD4, and CHD5), as well as CHARGE syndrome and idiopathic scoliosis (linked with CHD7). A structure based sequence alignment in the chromodomains of CHD proteins has suggested that the family can be divided into three groups. The variance between chromodomains appears to promote distinct epigenetic signalling for specific localization of each CHD protein. Therefore, we are interested in elucidating the structure and function of previously uncharacterized human CHD chromodomains. The best characterized example is the human CHD1 pair of tandem chromodomains that fold into a cooperative unit and bind a histone H3 tail with lysine methylation at residue 4 (H3K4 methylation).
Our first aim i s to determine chromodomain target specificity of representatives from the two uncharacterized groups, two and three. To do this we will express recombinant protein and utilize fluorescence-based binding assays to determine the dissociation constant of binding to a peptides resembling histone tail modifications. Isothermal titration calorimetry will also be used to determine the thermodynamic parameters of binding.
In aim two we will map the surfaces of interaction through structure determination with the use of nuclear magnetic resonance spectroscopy and x-ray crystallography. Our long term goal is to characterize the chromodomains of this family by determining their roles in epigenetic signalling. Recent studies link epigenetic misregulations with important human diseases including cancer, diabetes, and developmental disorders. To better understand epigenetic regulation, we will use structural biology and biochemistry methods to delineate the signalling mechanisms in a family of human proteins.
