Slow folding and assembly of a procaspase-3 interface variant

Area of Science:

• Biochemistry and Molecular Biology
• Cellular Biology
• Protein Folding and Assembly

Background:

• Caspases are key regulators of apoptosis, existing as inactive zymogens (procaspases).
• Initiator procaspases dimerize for activation, while effector procaspases like procaspase-3 are stable dimers requiring processing.
• The dimer interfaces differ between initiator and effector procaspases, with their role in oligomerization being unclear.

Purpose of the Study:

• To investigate the role of the dimer interface in the folding and activation of procaspase-3.
• To determine if the dimer interface is crucial for procaspase-3 oligomerization and assembly.

Main Methods:

• Equilibrium and kinetic folding studies were conducted on a procaspase-3 interface variant (C163S,V266H).
• Analysis of refolding and unfolding kinetics at different pH values.
• Investigated the impact of introducing histidine residues into the dimer interface.

Main Results:

• Procaspase-3 folding exhibits hysteresis, indicating a kinetically controlled process.
• Refolding involves monomeric intermediates that can become kinetically trapped and aggregate.
• Unfolding reveals structured monomeric intermediates, with slow dimerization kinetics due to inefficient orientation of interface residues.

Conclusions:

• The dimer interface is critical for procaspase-3 folding and assembly.
• Introducing histidine into the interface acts as a negative design element, preventing dimerization and activation.
• Caspase assembly is regulated at the interface by controlling the rate of dimerization.