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The three-dimensional structure of caspase-8: an initiator enzyme in apoptosis
H Blanchard1, L Kodandapani, P R Mittl
1Biochemisches Institut Universität Zürich Winterthurer Strasse 190, CH-8057, Zürich, Switzerland.
Background:
In the initial stages of Fas-mediated apoptosis the cysteine protease caspase-8 is recruited to the cell receptor as a zymogen (procaspase-8) and is incorporated into the death-signalling complex. Procaspase-8 is subsequently activated leading to a cascade of proteolytic events, one of them being the activation of caspase-3, and ultimately resulting in cell destruction. Variations in the substrate specificity of different caspases have been reported.
Results:
We report here the crystal structure of a complex of the activated human caspase-8 (proteolytic domain) with the irreversible peptidic inhibitor Z-Glu-Val-Asp-dichloromethylketone at 2.8 A resolution. This is the first structure of a representative of the long prodomain initiator caspases and of the group III substrate specificity class. The overall protein architecture resembles the caspase-1 and caspase-3 folds, but shows distinct structural differences in regions forming the active site. In particular, differences observed in subsites S(3), S(4) and the loops involved in inhibitor interactions explain the preference of caspase-8 for substrates with the sequence (Leu/Val)-Glu-X-Asp.
Conclusions:
The structural differences could be correlated with the observed substrate specificities of caspase-1, caspase-3 and caspase-8, as determined from kinetic experiments. This information will help us to understand the role of the various caspases in the propagation of the apoptotic signal. The information gained from this investigation should be useful for the design of specific inhibitors.
Insights
The crystal structure of activated human caspase-8 reveals distinct active site differences compared to caspase-1 and caspase-3. These structural variations explain caspase-8
Area of Science:
- Biochemistry
- Structural Biology
- Molecular Biology
Background:
- Fas-mediated apoptosis involves caspase-8 recruitment and activation.
- Activated caspase-8 initiates a proteolytic cascade leading to cell death.
- Variations in caspase substrate specificity are known.
Purpose of the Study:
- Determine the crystal structure of activated human caspase-8.
- Elucidate the structural basis for caspase-8 substrate specificity.
- Provide insights for designing specific caspase inhibitors.
Main Methods:
- X-ray crystallography of activated human caspase-8 complexed with Z-Glu-Val-Asp-dichloromethylketone.
- Analysis of structural differences in active site regions compared to other caspases.
Main Results:
- Reported the crystal structure of activated human caspase-8 at 2.8 A resolution.
- Identified distinct structural differences in active site subsites (S3, S4) and inhibitor interaction loops.
- Explained caspase-8's preference for (Leu/Val)-Glu-X-Asp substrates.
Conclusions:
- Structural differences correlate with observed substrate specificities of caspase-1, -3, and -8.
- Understanding these differences aids in comprehending the apoptotic signal propagation.
- The findings are valuable for the rational design of specific caspase inhibitors.