Crystal Structure of an Invertebrate Caspase

Abstract

Caspases play an essential role in the execution of apoptosis. These cysteine proteases are highly conserved among metazoans and are translated as inactive zymogens, which are activated by proteolytic cleavages to generate the large and small subunits and remove the N-terminal prodomain. The 2.3 Å resolution crystal structure of active Sf-caspase-1, the principal effector caspase of the insect Spodoptera frugiperda, is presented here. The structure represents the first nonhuman caspase to be resolved. The structure of the cleaved and active protease was determined with the tetrapeptide inhibitor N-acetyl-Asp-Glu-Val-Asp-chloromethylketone covalently bonded to the active site cysteine. As expected, the overall fold of Sf-caspase-1 is exceedingly similar to that of the five active caspases from humans solved to date. The overall structure and active site arrangement of Sf-caspase-1 is most comparable with that of the human effector caspases, with which it shares highest sequence homology. The most prominent structural difference with Sf-caspase-1 is the position of the N-terminal region of the large subunit. Unlike the N terminus of human caspases, the N terminus of Sfcaspase-1 originates from the active site side where it interacts with active site loop L2 and then extends to the backside of the heterodimer. This unusual structural arrangement raises the possibility that the N-terminal prodomain plays a regulatory role during effector caspase activation or enzyme activity in insects.Apoptosis is a cellular pathway that eliminates damaged, potentially dangerous, superfluous, or unwanted cells in metazoan organisms. This programmed cell death pathway is a naturally occurring physiological process that is vital to normal organismal development and tissue homeostasis. Abnormalities in the regulation of apoptosis can trigger many diseases including cancer, neurodegenerative disorders, autoimmune disorders, and ischemic injury. The apoptotic pathway is highly conserved in the Metazoa kingdom and can be triggered by both intracellular and extracellular stimuli. Apoptosis is executed through the activity of the caspases that are aspartyl-specific proteases (1-6). Because of their central role in apoptosis, these cysteine proteases are attractive targets for therapeutic intervention.Synthesized as dormant single-chain zymogens, the caspases are activated by a hierarchical series of proteolytic cleavages. The apical initiator caspases typically contain a large N-terminal prodomain that interacts with cellular factors that initiate apoptosis. These initiator caspases oligomerize through their prodomains that also associate with other proteins and promote enzyme activation in the absence of proteolytic processing (7). The activated initiator caspases subsequently cleave the proform of downstream effector caspases, which usually possess a short N-terminal prodomain. The active effector caspases proteolytically cleave cellular substrates that produce the morphological events associated with apoptosis, including cytoplasmic shr...

Baculovirus Caspase Inhibitors P49 and P35 Block Virus-Induced Apoptosis Downstream of Effector Caspase DrICE Activation in Drosophila melanogaster Cells

“…Thus, our studies indicate that activation of DrICE follows a path that is common to invertebrate effector caspases (Fig. 10), including that of the closely related effector caspase Sf-caspase-1 from Spodoptera frugiperda (10,26,27,43,56).…”

Porphyromonas gingivalis Virulence Factor Gingipain RgpB Shows a Unique Zymogenic Mechanism for Cysteine Peptidases

“…Zymogens of family C14, in turn, include the structurally studied mammalian caspases 1 (58); 3 (59); 7 (60,61), and 8 (62,63) and the insect Drosophila caspase-9 ortholog DRONC (64) and Spodoptera frugiperda caspase-1 ortholog (Ref. 65 and PDB 2NN3). Caspases are oligomeric functional enzymes, and activation cleavage entails major rearrangement of the loops flanking the active-site cleft from an incompetent to a competent conformation.…”

Molecular Cloning and Characterization of the First Caspase in the Striped Stem Borer, Chilo suppressalis

“…Two cleavage sites were found to be located between the prodomain and large subunit at Asp 23 , and between the large and small subunits at Asp 194 . Moreover, Cs caspase-1 contains QACQG pentapeptide active-site motif, which is also found in lepidopteran caspase-1 (Figure 1A) [5,14–17]. …”

“…To investigate the potential structure-function relationship of Cs caspase-1, we generated its homology model with Phyre using Spodoptera frugiperda caspase-1 (PDB ID: 2NN3) as a template [14]. The structure of the Cs caspase-1 model was very similar to that of caspase-1 of S. frugiperda (confidence, 100% statistically and identity, 87% based on sequence alignment) (Figure 1B).…”

“…66 caspase sequences representing 27 lepidopteran insect species can be classified into at least five groups by phylogenetic analyses [13]. However, completed studies on the genome, structure and functions of caspases in insect were found infrequently, especially the relationship with thermotolerance [5,14–17]. …”