title: Expression and characterization of ARSP1 from Eisenia fetida.
abstract: To study the characterization of a protease ARSP1 (apoptosis-related serine protease) of Eisenia fetida, a recombinant ARSP1 was constructed. ARSP1 was produced in E. coli BL21-CodonPlus (DE3)-RIL after IPTG induction and exited in inclusion body. After refolding in vitro, the protein was purified by DEAE-Sepharose F.F. and Sephacryl S-100 chromatography in sequence. ARSP1 showed high sequence identity to other chymotrypsin-like serine proteases and the catalytic triad was His41-Asp90-Ser188. ARSP1 could degrade casein following Michaelis-Menten kinetics with a Vmax of 43.9 U/mg protein and a Km for casein of 0.83 g/l. Studies with inhibitors indicated that ARSP1 was a chymotrypsin-like serine protease. Experiments in vitro demonstrated that ARSP1 could not only hydrolyze fibrinogen and fibrin directly, but also activate plasminogen to plasmin. ARSP1 inhibited thrombin activity and ADP-induced platelet aggregation in a dose-response correlation. These results showed that ARSP1 has thrombolytic activity and also an anti-thrombus function.
authors.affiliation: College of Life Sciences, National Lab of Protein Engineering, Peking University, Beijing, 100871, PR China. arm@pku.du.cn
authors.forename: Leiliang
authors.initials: L
authors.lastname: Zhang
authors.forename: Jing
authors.initials: J
authors.lastname: Wang
authors.forename: Meimin
authors.initials: M
authors.lastname: Yu
authors.forename: Binggen
authors.initials: B
authors.lastname: Ru
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis / Q000235:genetics; D005342:Fibrinolysis / Q000187:drug effects; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology*; D006801:Humans; D008969:Molecular Sequence Data; D009835:Oligochaeta / Q000201:enzymology*; D010975:Platelet Aggregation Inhibitors / Q000737:chemistry / Q000378:metabolism; D012697:Serine Endopeptidases / Q000096:biosynthesis* / Q000737:chemistry* / Q000235:genetics; D013918:Thrombin Time
chemical_list: D010975:Platelet Aggregation Inhibitors; D012697:Serine Endopeptidases