ANTHRAX TOXIN LETHAL FACTOR CONTAINS A ZINC METALLOPROTEASE CONSENSUSSEQUENCE WHICH IS REQUIRED FOR LETHAL TOXIN ACTIVITY

Comparison of the anthrax toxin lethal factor (LF) amino acid sequence with sequences in the Swiss protein database revealed short regions o f similarity with the consensus zinc-binding site, HEXXH, that is char acteristic of metalloproteases. Several protease inhibitors, including bestatin and captopril, prevented intoxication of macrophages by leth al toxin. LF was fully inactivated by site-directed mutagenesis that s ubstituted Ala for either of the residues (H-686 and H-690) implicated in zinc binding. Similarly, LF was inactivated by substitution of Cys for E-687, which is thought to be an essential part of the catalytic site. In contrast, replacement of E-720 and E-721 with Ala had no effe ct on LF activity. LF bound Zn-65 both in solution and on protein blot s. The Zn-65 binding was reduced for several of the LF mutants. These data suggest that anthrax toxin LF is a zinc metallopeptidase, the cat alytic function of which is responsible for the lethal activity observ ed in cultured cells and in animals.