Assessing the Binding Capabilities of Bromodomain-Containing Protein 9

Sarah Hopson (Advisor- Dr. Martin Thompson)

Doctoral Student, Department of Chemistry,Michigan Technological University

Monday, March 2, 2015-9:00 am- Admin 404

Abstract

Post-translational modifications of histones, such as the acetylation of lysines, play an importantrole in regulating transcription. Histone tails have a large proportion of positively-charged residues, which create electrostatic interactions with the negatively-charged DNA backbone. Lysine acetylation is thought to weaken these interactions, because it neutralizes lysine’s positively-charged side chain.

Proteins recognize the acetylated lysines using bromodomains; bromodomains are acetylated lysine “readers” and play a critical role in modulation of gene expression. Of the 46 bromodomain-containing proteins in the human proteome, 15 function as transcriptional regulators and 8 function as chromatin remodelers. Nearly all of the other bromodomain proteins influence transcription in some manner (histone acetyltransferase, transcription repressor, transcription initiation, etc.). Due to their significant influence on transcription, mutations of bromodomains are often linked with cancers.

Bromodomain-containing protein 9 (BRD9) has not yet been studied. The aim of this proposed research is to determine the specificity and affinity of BRD9 toward acetyl-lysine sites on the tails of the four core histone proteins.

A high-throughput examination of possible histone interactions with the bromodomain of BRD9 will be conducted using a modified SPOT array. The peptides demonstrating the strongest interactions with the bromodomain will be synthesized using standard Fmoc peptide synthesis. A quantitative examination of the binding affinities of these peptides to the bromodomain, the bromodomain and DUF3512 (domain of unknown function), and the full length BRD9 will be conducted using isothermal titration calorimetry. The results will be compared to determine how the surrounding amino acid sequences affect the bromodomain’s binding capabilities.


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