Hsp85 conformational change within the heat shock temperature range

Area of Science:

• Molecular Biology
• Biochemistry
• Protein Folding

Background:

• Heat shock proteins (HSPs) are crucial for cellular response to stress.
• Mammalian heat shock protein 85 (hsp85) is known to aggregate with detergents upon heating.
• Understanding hsp85 conformational dynamics is key to cellular stress response mechanisms.

Purpose of the Study:

• To investigate the conformational changes of hsp85 within physiological and heat shock temperatures.
• To determine the temperature range and characteristics of these conformational transitions.
• To assess the influence of detergents on hsp85 conformation during heating.

Main Methods:

• Intrinsic fluorescence spectroscopy to monitor changes in protein structure.
• Tryptic proteolysis susceptibility assays to evaluate protein unfolding and exposure of sites.
• Heating experiments conducted in the presence and absence of n-octyl-O-glucoside detergent.

Main Results:

• Heating hsp85 from 15°C to 50°C caused decreased fluorescence intensity and a 2.5 nm red shift in emission spectra.
• A major conformational transition, indicated by spectral red shift, occurred between 38°C-45°C in the absence of detergent.
• Proteolysis susceptibility increased sharply between 40°C-45°C, indicating partial unfolding and exposure of sites.

Conclusions:

• hsp85 undergoes a significant conformational change, consistent with partial unfolding, between 40°C and 50°C.
• This thermal transition occurs within the temperature range that induces heat shock protein synthesis.
• The conformational change exposes new sites, influencing interactions with the aqueous environment and detergent binding.