Molecular Cloning and Sequence Analysis of cDNAs for Metalloproteinases from Broad-Banded Copperhead Agkistrodon contortrix laticinctus

The cDNA sequences of two related genes coding for metalloproteinases from a venom gland library of Agkistrodon contortrix laticinctus have been determined. The ACLPREH cDNA codes for a 220-amino acid zinc-dependent hemorrhagic metalloproteinase, and the predicted sequence is in agreement with the N-terminal sequence of the previously purified protein, ACLPREF cDNA sequence predicts a 222-amino acid molecule having strong similarity to fibrolase, a fibrinolytic enzyme isolated from A. contortrix contortrix venom that is reported to be devoid of hemorrhagic activity. Both cDNAs present the same pattern of domain organization as other members of the metalloproteinase/disintegrin gene family. They code for short toxins which do not have the disintegrin domain and have three-disulfide bonds in the metalloproteinase domain. Both cDNA sequences have the highly conserved 5′ and 3′ untranslated regions that have been described for the metalloproteinase and phospholipase A2 snake venom gene families. ACLPREH and ACLPREF cDNAs share a higher degree of homology in the untranslated regions and proenzyme domain than in the mature protein domain. The amino acid sequences of hemorrhagic and/or fibrinolytic metalloproteinases demonstrate that a few substitutions may result in different enzymatic activities. Also, some of these toxins have valine instead of isoleucine in the CIM Met-turn consensus sequence. From our data, we can suggest that snake venom fibrinolytic metalloproteinase genes belong to the metalloproteinase/disintegrin gene family. This is the first report of cDNA sequences of small snake venom metalloproteinases having three disulfide bonds in the metalloproteinase domain.