1999 Volume 125 Issue 1 Pages 64-69
A non-hemorrhagic proteinase, brevilysin L6 (L6), has been purified to homogeneity from Agkistrodon halys brevicaudus venom by gel filtration and DEAE-Toyopearl 650M chromatography. It is an acidic protein with an isoelectric point of 4.8, and its molecular mass was estimated to be 21.5 kDa by SDS-PAGE. The optimum pH of L6 was about 9. EDTA and o-phenanthroline inhibited the proteolytic activity, suggesting that L6 is a metalloprotease. Cysteine also inhibited the activity of L6, but glutathione did not. The protein was stable in the pH range of 5-8.5 and below 40°C. Calcium ions had no effect on the proteolytic activity of L6 or on its thermal stability. The enzyme preferentially cleaved X-Leu, X-Phe, X-Val, and X-His bonds. L6 showed weak α-fibrinogenase activity. The complete amino acid sequence of L6 was also determined by manual Edman degradation. L6 is a nonglycosylated single-chain polypeptide consisting of 203 residues with an N-terminal pyroglutamic acid and a calculated molecular weight of 22, 713 Da. Its entire sequence is highly homologous to those of other metalloproteases from various snake venoms. A zinc-binding motif, HEXXHXXGXXH, is located at residues 143-153 in the sequence of L6.
