Abstract
Human linker histones (H1s) play an important role in chromatin compaction. The effect each H1 variant has on higher-order chromatin structure and nuclear processes, such as transcription, is not yet fully understood. Linker histones consist of a central, structured globular domain flanked by an N-terminal (NTD) and C-terminal domain (CTD). The intrinsic unstructured nature of the terminal domains may allow interaction with a wide range of binding partners or DNA architectures. Linker histone variant H1x is the most recently classified linker histone variant with terminal domains that are distinct from that of other H1 variants. The globular domain structure of human H1x was previously determined using nuclear magnetic resonance (NMR) spectroscopy. However, no information is available with regards to the structure of the NTD, or the effect of the NTD on chromatin structure. This study aimed to investigate these issues. The secondary structure of the H1x NTD was probed by NMR spectroscopy. A truncated version of H1x lacking the CTD (NGH1x) was employed, as the CTD is prone to degradation, adds significant bulk to the protein (approximately half the mass of the protein) and is inherently unstructured. Two a-helices (aN1 at residues A28 – L30; aN2 at residues S33 – K40) formed in the NTD in the presence of perchlorate ions, mimicking DNA. The NTD helices were distinct from other H1 NTDs, suggesting a unique DNA-binding mode in chromatin. The effect of the NTD in the context of the chromatosome was further investigated by molecular dynamic (MD) simulations. The NTD leads to the stabilization of an off-dyad binding mode of the globular domain relative to the nucleosome and altered the histone-DNA interaction pattern in the chromatosome.
Ph.D. (Biochemistry)