The mechanisms that lead to the activation of procaspases play central roles in the regulation of apoptosis and inflammation. For all procaspases, formation of a dimer is a critical event in processing because the active sites of the enzyme are comprised of loops contributed by both monomers of the dimer. However, a fundamental difference exists in the caspase subfamilies regarding dimerization and maturation, and this difference is a key aspect for regulating apoptosis. Several inflammatory and initiator procaspases exist as inactive monomers in the cell, but the proteins have relatively high enzymatic activity upon dimerization. For those caspases, chain cleavage simply stabilizes the active site. In contrast, the executioner procaspase-3 is a stable dimer, but it has very little enzymatic activity. In this case, maturation is dependent on cleavage by the initiator procaspases. Our long-term goals are to understand how dimerization, activation and enzymatic activity are coupled for procaspases. Overall, we want to determine why procaspase-1 is a monomer, but yet the active sites form properly upon dimerization. In contrast, we want to determine why the stable dimer of procaspase-3 is not active. We hypothesize that the procaspase dimer exists in two primary conformations, only one of which is active, and that the inactive form is most stable in the case of procaspase-3. Furthermore, we suggest that an allosteric site in the dimer interface affects the conformational switch between the two forms and couples dimerization to active site formation. We have shown that three regions of the protein may be important in affecting the oligomeric state and in stabilizing the inactive form of the procaspase. We propose to use biochemical and biophysical assays, protein folding studies, and X-ray crystallographic studies to examine these three regions of the protein. The procaspase activation mechanisms are important biologically because they allow for the tight regulation of caspase activity in cell death, from maintaining the monomeric state of initiator procaspases to stabilizing the inactive form of dimeric effector procaspases. Understanding the caspase activation process provides a key t develop strategies to intervene in their activities in the cell.
Understanding caspase dimerization and activation has the potential to affect therapeutic strategies for a number of autoimmune diseases, heart disease, and cancers. Learning to selectively manipulate the level of apoptosis is an important step in treatment because the dysregulation of apoptosis is a common factor to these diseases.
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|MacKenzie, Sarah H; Schipper, Joshua L; England, Erika J et al. (2013) Lengthening the intersubunit linker of procaspase 3 leads to constitutive activation. Biochemistry 52:6219-31|
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|Walters, Jad; Swartz, Paul; Mattos, Carla et al. (2011) Thermodynamic, enzymatic and structural effects of removing a salt bridge at the base of loop 4 in (pro)caspase-3. Arch Biochem Biophys 508:31-8|
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