Vibrio vulnificus, an etiologic agent of wound infections and septicemia in humans, elaborates a metalloprotease which is known to be an important virulence factor of the Vibrio. The proteolytic activity of V. vulnificus metalloprotease (VVP) toward casein and elastin was inhibited by α₂-macroglobulin (α₂ M) at the molar ratio of 1:1, although partial activity was maintained. Permeability-enhancing and hemorrhagic activities were also inhibited, but the peptidase activity toward Z-Gly-Phe-NH₂ was not reduced, even by an excess amount of α₂ M. VVP formed a complex with α₂ M through cleavage of the bait regions of all four α₂ M subunits and elicitation of conformational change of the α₂M molecule, which resulted in entrapment of VVP in the α₂ M molecule. The peptidase activity of α₂ M-VVP complex was inhibited by low-molecular-weight inhibitors such as phosphoramidon, but IgG antibody against VVP failed to neutralize its peptidase activity. Of human plasma proteins, α₂ M was the only inhibitor for VVP. These findings indicate that VVP produced during V. vulnificus infection is inactivated by plasma α₂ M that leaks from the vascular system.