Thermal denaturation of nucleosomal core particles

Area of Science:

• Biochemistry
• Molecular Biology
• Structural Biology

Background:

• Core particles are fundamental units of chromatin, composed of DNA and histone proteins.
• Understanding their thermal stability is crucial for elucidating DNA packaging and accessibility within the nucleus.

Purpose of the Study:

• To investigate the thermal denaturation process of chicken erythrocyte chromatin core particles.
• To characterize the distinct contributions of DNA and histone structures to thermal stability.

Main Methods:

• Differential scanning calorimetry (DSC) to measure heat capacity changes.
• UV-Vis spectroscopy to monitor base unstacking via absorbance changes.
• Circular dichroism (CD) spectroscopy to assess DNA and histone secondary structure alterations.

Main Results:

• Thermal denaturation of core particles occurred in two phases, with inflexions at 60°C and 74°C, distinct from isolated DNA denaturation at 44°C.
• CD spectroscopy revealed biphasic changes in DNA secondary structure preceding base unstacking.
• Histone secondary structure alterations were observed only in the second denaturation phase (melting at 71°C).
• Increasing ionic strength from 1 mM to 10 mM resulted in nearly monophasic melting curves without inducing conformational changes at room temperature.

Conclusions:

• The biphasic melting of core particles suggests sequential denaturation events: initial DNA denaturation followed by histone-DNA complex breakdown.
• The findings highlight the cooperative nature of histone-DNA interactions in maintaining chromatin structure.
• Ionic strength plays a significant role in stabilizing the core particle structure.