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Presentation Time: 1:55-2:15
Home University: UNC-Chapel Hill
Research Mentor: Brian D. Strahl, Biochemistry & Biophysics
Program: SMART
Research Title: Effect of Ada2 SANT Mutations on H3 and H4 Histone Tail Binding

Post-translational histone modifications (e.g., acetylation) are important mechanisms that contribute to gene transcription and expression. Misregulation of gene expression can lead to cancer or be devastating to embryonic development. The SAGA complex (Spt-Ada-Gcn5-acetyltransferase) is a yeast transcriptional co-activator that regulates acetylation of histones. Within the SAGA complex, the Ada2 protein is a transcriptional activator that is required for binding of H3 and H4 histone tails in yeast. Ada2 contains two distinct structural domains: ZZ and SANT. These domains work together to bind H3 and H4 histones. The purpose of this project was to better understand how each domain functions and binds histones independently. To investigate this, point-mutations were made in regions of the SANT domain suspected to be involved in binding the histone tails. These regions of the SANT domain were determined via sequence alignment to identify highly conserved residues, specifically acidic pockets within the SANT domain that work to bind histones. The Ada2 SANT wild type and point mutants were recombinantly expressed, purified, subsequently used in vitro analysis within Escherichia coli and visualized via western blot. Results suggest that triple-point mutations resulted in decreased binding with H3 and H4. Overall, the mutated proteins were unable to fully eliminate histone binding in H3 and H4. In the future, a crystal structure of the protein must be obtained to determine the actual location for histone binding.