CAREER: Folding of the Core Histones: Insights into Nucleosome Folding

Washington State University, Pullman WA

Investigators

Abstract

Gloss 9983831 The biophysical techniques of equilibrium and stopped-flow CD and fluorescence (FL) spectroscopies will be used to characterize the stability and folding reactions of the histone proteins of the core nucleosome and the nucleosome itself. The equilibrium and kinetic folding responses of variant forms of the core histones will determined with unmodified recombinant histones, truncated histones that lack the highly charged N-terminal tails or contain site-directed mutations that eliminate one or more basic residues from the tails, histones with specific acetylation patterns on the N-terminal tails, and histones containing mutational substitutions that mimic the effects of the ATP-dependent nucleosome remodeling complexes, whose activity is essential for gene regulation. The stability and folding of these histones variants will be examined in isolation and in complex with specific DNA fragments, including in the intact nucleosome. This research will permit a deeper understanding of the folding reactions of oligomeric proteins in general and provide insights into the assembly and stability of the nucleosome. Of particular importance is the biophysical characterization of the effects of the regulatory alterations brought about by histone acetylation and ATP-dependent chromatin remodelling complexes on histone and nucleosome structure and stability. Detailed biochemical and biophysical analyses of the nucleosome and its component histones, in the presence and absence of these effectors, are essential to understanding the nature of these alterations and the role they play in DNA packaging and gene regulation. The educational plan includes formal lectures on biophysics and protein folding in existing departmental courses, development of a techniques-oriented course on CD and fluorescence spectroscopies and their practical applications to biochemical experiments, including, but not limited to, protein folding, mentoring of graduate and undergraduate students in the PI's laboratory and in the department with an emphasis on multi-disciplinary approaches to biological problems.