ISOLATION AND CHARACTERIZATION OF AN ANTICOAGULANT FROM THE SALIVARY-GLANDS OF THE TICK, ORNITHODOROS-SAVIGNYI (ACARI, ARGASIDAE)

An inhibitor of activated coagulation factor X (fXa) was isolated from salivary gland extracts prepared from Ornithodoros savignyi using a t wo-step procedure, involving reversed-phase high-performance liquid ch romatography (RP-HPLC) and diethylaminoethyl (DEAE) ion-exchange chrom atography. From its behaviour during DEAF chromatography it could be d educed that it possesses an acidic pi (similar to 4.6). Capillary zone electrophoresis (CZE) of the purified inhibitor showed it to be homog eneous. The molecular mass was determined as 12 kDa using capillary ge l electrophoresis (CGE) and as 7183.4 using laser desorption mass spec trometry (LDMS). The N-terminal amino acid sequence (residues 1-12) wa s determined and found to share a 66% identity with tick anticoagulant peptide (TAP). The O. savignyi peptide is a slow, tight-binding inhib itor of fXa (K-i=0.83+/-0.10 nM). The interaction of the first-inhibit or was found to be competitive and dependent on ionic strength. Prelim inary investigations show that the inhibitor may be specific for fXa.