Inhibition of distant caspase homologues by natural caspase inhibitors

Caspases play an important role in the ability of animal cells to kill them selves by apoptosis. Caspase activity is regulated in vivo by members of th ree distinct protease inhibitor families, two of which, baculovirus p35 and members of the inhibitor of apoptosis (IAP) family, are thought to be casp ase specific. However, caspases are members of the clan of cysteine proteas es designated CD, which also includes animal and plant legumains, and the b acterial proteases clostripain, gingipain-R and gingipain-K. Since these pr oteases have been proposed to have a common mechanism and evolutionary orig in, we hypothesized that the caspase inhibitors may also regulate these oth er proteases. We tested this hypothesis by examining the effect of the natu ral caspase inhibitors on other members of protease clan CID. The IAP famil y proteins were found to have only a slight inhibitory effect on gingipain- R. The cowpox viral cytokine-response modifier A (CrmA) serpin had no effec t on any of the proteases tested but a single point mutation of CrmA (Asp L ys) resulted in strong inhibition of gingipain-K. More substantial, with re spect to the hypothesis, was the strong inhibition of gingipain-K by wild-t ype p35. The site in p35, required for inhibition of aingipain-K, was mappe d to Lys(94). seven residues C-terminal to the caspase inhibitory site. Our data indicate that the virally encoded caspase inhibitors have adopted a m echanism that allows them to regulate disparate members of clan CD protease s.