Molecular cloning and expression of Green pit viper (Trimeresurus albolabris) serine protease

Abstract

Viper is common snake family for venomous snakebites in Thailand. The venom destroys maining blood coagulation. Green pit viper (Trimeresurus albolabris) is the most common in Bangkok and boundary that have problem in medical and security. T albolabris venums are many kind proteins that have potential in blood coagulation. Serine protease have effect to thrombin clotting, plasminogen activator, fibrinogenolysis, anticoagulant agent and corrected anticoagulant. The study results of nucleic acid and amino acid from T albolerine primary cDNA library. Serine protease digest fibrinogen (albofibrase) that dase not full-length sequence. Using 5 ́-RACE method, the full length sequece of albofibrase was derived. Lbofibrase from Pichia pastoris was purified using affinity chromatography. The yield of albofibrase was 0.658 mg/liter of culture medium with molecular weight of approximately 30 kDa. Albofibrase could activate plasminogen (plasminogen activator albofibrase). Albofibrase have been thrombin clotting time at 0.058 unit thrombin. Albofibrase could digest fibrinogen at Aα-chain (fibrinogenolysis) and 180 minutes time incubation. Albofibrase could inhibit blood coagulation (anti-coagulant agent), so prolong of prothrmbin time (PT) and activated partial thromboplastin time (aPTT). Albofibrase could corrected time to normal PT and aPTT by green pit viper serum (corrected anti-coagulant). Suggesting that defibrination syndrome in patients is a combination of these enzymatic effects. Albofibrase is a novel serine protease has potentials to be therapeutic anti-thrombotic agent, fibrinogenolysis agent and anti-coagulant agent in the future.