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

Abstract 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 Aα-chain of fibrinogen with a high efficiency, and the Bβ- and γ-chains (Aα > Bβ > γ) with a lower efficiency. The proteolytic activity peaks at pH 7.8 is 45 °C. The activity is completely inhibited by serine protease inhibitors DFP (I 50  = 5.8 × 10 −4  M) and PMSF (I 50  = 5.5 × 10 −2  M), and almost completely by TLCK (I 50  = 7.7 × 10 −1 M). But aprotinin, elastinal, SBTI, benzamidine, PCMB, EDTA, EGTA, iodoacetate, E64, and β-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).