Inhibition of mutalysin II, a metalloproteinase from bushmaster snake venom by human alpha 2-macroglobulin and rabbit immunoglobulin

Mutalysin II is a 22.5-kDa zinc endopeptidase isolated from Lachesis muta m uta snake venom. In order to determine whether the inhibitors human alpha2- macroglobulin (alpha2-M) and rabbit antibody to mutalysin II share a common mechanism, we have investigated the inhibition of mutalysin II by these tw o different glycoproteins. The proteolytic activity of mutalysin II with di methylcasein as substrate was completely inhibited by human alpha2-M and by a purified rabbit antibody to mutalysin II. The protection of fibrin(ogen) digestion by alpha2-M was slightly better than the protection offered by t he antibody. In addition, the purified antibody reacted only with the metal loproteinase in bushmaster venom, as demonstrated by immunodiffusion. SDS-P AGE analysis of reduced samples showed that the interaction of mutalysin II with alpha2-M resulted in the formation of high molecular complex (similar to 180000) and M-r 90000 fragments generated by the venom enzyme. Also, fr agments at 85 and 23 kDa were detected under non-reducing conditions after incubation of rabbit immunoglobulin with enzyme. Proteolysis of dimethylcas ein as substrate revealed that the stoichiometry of inhibition was 1.0 mol of human alpha2-M. and 1.5 mol of rabbit IgG antimutalysin II per mole of e nzyme. Furthermore, dimethylcasein hydrolysis indicated that several viperi d snake venoms, including Bothrops atrox, B. alternatus and Trimeresurus fl avoviridis cross-reacted with the specific rabbit antibody to varying degre es. (C) 2001 Elsevier Science Inc. All rights reserved.