Proteolytic and Clot Lysis Activity Screening of Crude Proteases Extracted from Tissues and Bacterial Isolates of Holothuria Scabra

Abstract

Abstract Cardiovascular diseases (CVDs) are still the leading mortality causes in the last decades. The ailments are multifactorial characterized by excessive clot (thrombus) formation in the blood vessels. Thrombus could be degraded through thrombolysis mechanism by plasmin activated by various fibrinolysis agents including, urokinase, nattokinase, or streptokinase. However, the use of these agents is restricted by relatively high cost, short half-life, allergic reaction, and bleeding effects. The search for more economical and safer thrombolytic (clot lysis) agents are essential to address the underlying problem in CVD therapy. Among Holothurians, H. scabra has been known to have the highest protein content making it ideal substrate for protease enzymes including fibrinolytic types with clot lysis properties. However, isolation of a proteases with antithrombotic activities either from tissue or from bacteria of H. scabra has not been reported. This study aimed to screen proteolytic and clot lysis activities of crude protease extracts from tissue and bacteria isolated from fermented intestine of H. scabra. Crude protease of tissue of H. scabra was extracted by cold centrifugation, which activity was measured using UV-spectrophotometer. Crude protease of proteolytic bacteria selected by clear zones on skim milk agar (SMA) medium was isolated from nutrient broth (NB). Extracts showing proteolytic activity were subjected to gravimetry-based clot lysis test. As results, crude proteases isolated directly from the H. scabra’s tissue showed low proteolytic activities, thus were no proceed to clot lysis activity test. Crude protease extracted bacteria could show both proteolytic and clot lysis activities. In conclusion, based on this screening study, intestine of H. scabra is a rich source of proteolytic bacteria, some of them could produce crude enzymes showing competitive thrombolysis activities with Nattokinase. Thus, they appeared to have more potentials to be developed as thrombolysis agent than those directly extracted from the organism’s tissue.