Isolation and characterization of a novel proteinase inhibitor from the snake serum of Taiwan habu (Trimeresurus mucrosquamatus)

A proteinase inhibitor (designated as TMD was isolated and purified from th e snake serum of Taiwan habu (Trimeresurus mucrosquamatus) by using success ive chromatographies which included Sephadex G-100, DEAE-Sephacel chromatog raphies, and C-4 reverse-phase HPLC. The purified inhibitor was shown to be a homogeneous protein with a molecular mass of about 47 or 36 kDa in the p resence or absence of a reducing agent, beta-mercaptoethanol. The inhibitor decreases in molecular mass by about 23% with N-Linked neuraminidase treat ment, suggesting that it is a glycoprotein. Further enzymatic analyses indi cated that this inhibitor possesses strong inhibitory activities toward thr ee zinc-dependent metalloproteinases and not fibrinogenolytic serine protea ses previously isolated from the venom of the same snake species with an IC 50 of about 0.2-1.1 mu M. Its IC50 value was approximately three orders of magnitude more effective than those of the tripeptide inhibitors we previou sly purified from the crude venom of the same snake Biochem. Biophys. Res. Commun. 248, 562-568 (1998)). The purified inhibitor showed stronger inhibi tory action against caseinolytic activities of crude venoms from closely re lated species of Taiwan habu than those from unrelated species. N-terminal sequence analysis showed that its sequence is distinctly different from seq uences of those serum inhibitors reported for other snake species in the li terature. Based on inhibition susceptibility and primary structures of vari ous snake protease inhibitors, it is suggested that this novel inhibitor is olated from the serum of Taiwan habu may be a unique self-defense protein f actor mainly for protection against envenomation from snakes of the same ge nus. (C) 1999 Academic Press.