A novel fibrinolytic serine protease from the polychaete Nereis (Neanthes) virens (Sars): Purification and characterization

A novel fibrinolytic serine protease has been identified and purified to homogeneity from the coelomic fluid of polychaete Nereis (Neanthes) virens (Sars), and named N-V protease. N-V protease is a 29 kDa single chain protein with an isoelectric point of pH 4.5. It hydrolyzes A alpha-chain of fibrinogen with a high efficiency, and the B beta- and gamma-chains (A alpha > B beta > gamma) with a lower efficiency. The proteolytic activity peaks at pH 7.8 is 45 degrees C. The activity is completely inhibited by serine protease inhibitors DFP (I-50 = 5.8 x 10(-4) M) and PMSF (I-50 = 5.5 x 10(-2) M), and almost completely by TLCK (I-50 = 7.7 x 10(-1) M). But aprotinin, elastinal, SBTI, benzamidine, PCMB, EDTA, EGTA, iodoacetate, E64, and beta-mercaptoethanol have no effect on the protease activity. Therefore, N-V protease is identified as a serine protease. The primary amino acid sequence of N-V protease was determined by mass spectrometry (N-V protease, No. P83433). According to the MALDI-TOF MS analysis, there is no existing protein in the NCBI Non-redundant Protein Sequence Database that matches the N-V protease sequence. Therefore, N-V protease is a novel and special protein in N. virens. Furthermore, we have successfully established an expression cDNA library from the whole body of N. virens (data not shown). (c) 2006 Elsevier Masson SAS. All rights reserved.