CAREER: Chemical Tools to Probe Histone H4 Modifications in the Nucleosome Core

Ohio State University Research Foundation -Do Not Use, Columbus OH

Investigators

Abstract

This award is funded under the American Recovery and Reinvestment Act of 2009 (Public Law 111-5) The nucleosome is the fundamental unit of chromatin, formed of 147 base pairs of DNA wrapped around a histone octamer protein core. Post-translational modifications in this structured histone core are likely to play vital roles in assembly, maintenance, and regulation of chromatin structures. However, the difficulty of preparing well-defined samples has hampered the study of how these chemical modifications translate into physical structure and biological function. This project builds up a toolkit using peptide and protein chemistry to introduce modifications into protein-protein and protein-DNA interfaces in the nucleosome core, with a focus on histone H4 modifications that are likely to impact nucleosome assembly and DNA repair. Chemical techniques will be developed for the total synthesis of histone proteins by native chemical ligation on the solid phase, which will allow the facile assembly of histones bearing precise combinations of modifications throughout the protein sequence. Histone synthesis and semisynthesis will be used to prepare histone H4 with combinations of acetylation, methylation, or phosphorylation modifications at DNA-interaction motifs across the nucleosome lateral surface and at the tetramer-dimer interface in the histone octamer. An array of biochemical and biophysical methods will be used to determine how these modifications affect nucleosome and chromatin structure and function. The broader impacts of this research include improved methods for total protein synthesis that will not only open up new areas in the study of chromatin modifications, but will also be applicable to any small protein. The research project provides highly interdisciplinary training and mentoring for undergraduate and graduate students across four departmental boundaries and will be leveraged in the classroom through development of a curricular unit in chromatin biology. The project will support development of a web-based community to support undergraduate research. Early participants in this community will be undergraduate researchers on-campus and summer research students recruited primarily from historically economically and ethnically diverse institutions; these students can remain a part of the support network after returning to their home institution. Finally, the project supports an integrated plan for 1) increasing undergraduate research participation 2) training in advanced research methods 3) career planning and mentoring 4) research ethics training and 5) dissemination of undergraduate research results to a wider community through organization of undergraduate research forums.