Investigation into the Properties of Telomeric Mono-nucleosomes

Abstract

One of the hallmark of eukaryotic cells is the presence of telomeres at the ends of their linear chromosomes. These sequences are highly conserved across different eukaryotic organisms, and in vertebrates, exist as repeats of (TTAGGG)n which would eventually end with a G-rich overhang [1]. Telomeres are key in regulating genome stability, and their mis-regulation has been implicated in cancer and senescence [2]. Similar to non-telomeric regions of the genome, telomeres are wrapped around core histones, which, in turn, form the Nucleosome Core Particle (NCP). These NCPs further compacts and forms chromatin [3]. Core histones are multimeric proteins composed of 8 subunits (two of each: H2A, H2B, H3 and H4); and usually has a 147-bp DNA wrapped around this disc-shaped octamer. The structure and biophysics of telomere chromatin are not very well characterized [1]. This project aims to reconstitute telomeric DNA with human histone octamer in vitro, and use these reconstituted NCPs for structural and biophysical characterizations. In addition, large scale expression and purification of the telomeric DNA construct and the components of the histone octamer will be performed to produce samples for biophysical and structural characterizations. This poster reports preliminary work on the large scale expression and purification of human histone H2B (hH2B) and 147-bp telomeric DNA. [1st Award]