MOLECULAR-CLONING AND SEQUENCE-ANALYSIS OF THE CDNA FOR ANCROD, A THROMBIN-LIKE ENZYME FROM THE VENOM OF CALLOSELASMA-RHODOSTOMA

The 1.54 kb cDNA for ancrod, a thrombin-like enzyme, was cloned from a lambdaZAP cDNA library derived from the venom glands of Calloselasma (Agkistrodon) rhodostoma. The cDNA sequence reveals that ancrod is syn thesized as a pre-zymogen of 258 amino acids, including a putative sec retory peptide of 18 amino acids and a proposed zymogen peptide of 6 a mino-acid residues. The animo-acid sequence of the predicted active fo rm of the enzyme exhibits a high degree of sequence similarity to thos e of mammalian serine proteases (trypsin and pancreatic kallikrein) an d other thrombin-like enzymes (batroxobin and flavoxobin). Key amino-a cid residues (His43, Asp88, Ser182 and Asp176) that are thought to be involved in the substrate cleavage and in the substrate-binding reacti on are conserved. Ancrod contains 13 cysteine residues. Based on align ment with the amino-acid sequences of trypsin and batroxobin, six disu lphide bridges can be predicted to be present in the ancrod protein. T he existence of a free cysteine, which changes the common sequence sur rounding the Ser182 active site from Gly-Asp-Ser-Gly-Gly-Pro to Cys-As p-Ser-Gly-Gly-Pro, is unusual for a serine protease.