title: Purification of serine protease from polychaeta, Lumbrineris nipponica, and assessment of its fibrinolytic activity.
abstract: Ischemic stroke and cardiovascular disease can occur from blockage of blood vessels by fibrin clots formed naturally in the body. Therapeutic drugs of anticoagulant or thrombolytic agents have been studied; however, various problems have been reported such as side effects and low efficacy. Thus, development of new candidates that are more effective and safe is necessary. The objective of this study is to evaluate fibrinolytic activity, anti-coagulation, and characterization of serine protease purified from Lumbrineris nipponica, polychaeta, for new thrombolytic agents. In the present study, we isolated and identified a new fibrinolytic serine protease from L. nipponica. The N-terminal sequence of the identified serine protease was EAMMDLADQLEQSLN, which is not homologous with any known serine protease. The size of the purified serine protease was 28 kDa, and the protein purification yield was 12.7%. The optimal enzyme activity was observed at 50°C and pH 2.0. A fibrin plate assay confirmed that indirect fibrinolytic activity of the purified serine protease was higher than that of urokinase-PA, whereas direct fibrinolytic activity, which causes bleeding side effects, was relatively low. The serine protease did not induce any cytotoxicity toward the endothelial cell line. In addition, anticoagulant activity was verified by an in vivo DVT animal model system. These results suggest that serine protease purified from L. nipponica has the potential to be an alternative fibrinolytic agent for the treatment of thrombosis and use in various biomedical applications.
authors.affiliation: Department of Biological Engineering, Inha University, Incheon, South Korea.
authors.forename: Seung Ju
authors.initials: SJ
authors.lastname: Yeon
authors.affiliation: Department of Biological Engineering, Inha University, Incheon, South Korea.
authors.forename: Goo Yong
authors.initials: GY
authors.lastname: Chung
authors.affiliation: Department of Biological Engineering, Inha University, Incheon, South Korea.
authors.forename: Jae Sang
authors.initials: JS
authors.lastname: Hong
authors.affiliation: Department of Biological Engineering, Inha University, Incheon, South Korea. kjinha@hotmail.com.|ST Pharm Co., Ltd., Gyeonggi-do, South Korea. kjinha@hotmail.com.
authors.forename: Jin Ha
authors.initials: JH
authors.lastname: Hwang
authors.affiliation: Department of Biological Engineering, Inha University, Incheon, South Korea. hsshin@inha.ac.kr.
authors.forename: Hwa Sung
authors.initials: HS
authors.lastname: Shin
mesh_terms: D000595:Amino Acid Sequence / Q000235:genetics; D000818:Animals; D002545:Brain Ischemia / Q000188:drug therapy*; D005337:Fibrin / Q000737:chemistry / Q000235:genetics; D005343:Fibrinolytic Agents / Q000737:chemistry / Q000302:isolation & purification* / Q000627:therapeutic use; D011077:Polychaeta / Q000201:enzymology; D057057:Serine Proteases / Q000737:chemistry / Q000302:isolation & purification* / Q000627:therapeutic use; D020521:Stroke / Q000188:drug therapy*
chemical_list: D005343:Fibrinolytic Agents; D005337:Fibrin; D057057:Serine Proteases
keywords: Anticoagulant activity; Bi-functional thrombolytic activity; Fibrinolytic activity; Lumbrineris nipponica; Serine protease