Isolation and properties of two forms of thrombin inhibitor from the nymphs of the camel tick Hyalomma dromedarii (Acari: Ixodidae).
Two forms of the nymphal thrombin inhibitors (NTI) 3.2 kDa and 14.9 kDa were purified by chromatography on CM-cellulose. Sephacryl S-300 and Sephadex G-50 columns and designated NTI- 1 and NTI-2 respectively. The NTI-2 turned out to be homogenous monomeric protein in both native-PAGE and denatured SDS-PAGE with M(r) value of 14.9 kDa approximately and its pI value ranged from 7.2 to 7.5. The NTI-1 and NTI-2 displayed anticoagulant activity since they prolonged both the activated partial thromboplastin time (APTT) and the prothrombin time (PT) of the camel plasma in a concentration-dependent manner. The potency of NTI-I toward thrombin was 5-fold higher than that toward FXa, while NTI-2 was 3-fold active toward FXa than thrombin. However, both of them did not inhibit any of the other examined proteases. The types of inhibition of thrombin by NTI-1 and NTI-2 were non-competitive and competitive with inhibition constants (Ki) values of 11.7 microM and 211 nM respectively. One binding site was deduced on thrombin for each inhibitor.
Ibrahim
M A
MA
Molecular Biology Dept., National Research Centre, Dokki, Cairo, Egypt. ibrahimm70@hotmail.com
Ghazy
A H
AH
Maharem
T
T
Khalil
M
M
D000818:Animals; D001777:Blood Coagulation; D002162:Camelus / Q000469:parasitology*; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D004791:Enzyme Inhibitors / Q000032:analysis*; D026863:Ixodidae / Q000737:chemistry*; D007700:Kinetics; D009758:Nymph; D013917:Thrombin / Q000037:antagonists & inhibitors*; D017186:Titrimetry
D004791:Enzyme Inhibitors; D013917:Thrombin