Biomedical subjects

Seung R Paik

Publications and source records attributed to Seung R Paik.

At least 19 recordsLinked to original sources

PubMed2006

Calpain-resistant fragment(s) of alpha-synuclein regulates the synuclein-cleaving activity of 20S proteasome.

Alpha-synuclein is a pathological component of Parkinson's disease by participating in Lewy body formation. Imbalance in protein turnover could result in the abnormal protein aggregation responsible for eventual neuronal cell death. This in vitro digestion study showed that both m-calpain and 20S proteasome preferentially hydrolyzed the N-terminal half of alpha-synuclein, which made the hydrophobic NAC and following acidic C-terminal region resistant against the proteolyses. Since the acidic C-terminal region contains the PEST segment-a protein degradation signal enriched with amino acids of proline (P), glutamate (E), serine (S), and threonine (T)-, the PEST segment has not been processed or even required for the proteolyses. Alpha-synuclein would be recognized primarily by m-calpain since the common substrate was processed by m-calpain five times more effectively than 20S proteasome with k(cat)/K(m) of 1.64 x 10(4)M(-1)s(-1) and 0.32 x 10(4) M(-1)s(-1), respectively. The N-terminally truncated protease-resistant C-terminal fragment of alpha-syn61-140 was demonstrated to stimulate the 20S proteasome-mediated breakdown of alpha-synuclein and its mutant forms of Ala53Thr and Ala30Pro. The stimulation for Ala53Thr, however, was noticeably less efficient than those for the other proteins, which might support the previous observation of the prolonged intracellular life span of Ala53Thr by 1.5-fold compared to that of wild-type form. We have hypothesized that the N-terminally truncated C-terminal fragment derived from the abundant alpha-synuclein through intracellular proteolyses could be involved in various physiological or pathological effects which might be related to the formation of abnormal protein aggregation and subsequent neuronal degeneration by influencing the intracellular protein turnover or directly participating in the aggregate formation.

Amino Acid Sequence↗

PubMed2006

Dyrk1A phosphorylates alpha-synuclein and enhances intracellular inclusion formation.

Lewy bodies (LBs) are pathological hallmarks of Parkinson disease (PD) but also occur in Alzheimer disease (AD) and dementia of LBs. Alpha-synuclein, the major component of LBs, is observed in the brain of Down syndrome (DS) patients with AD. Dyrk1A, a dual specificity tyrosine-regulated kinase (Dyrk) family member, is the mammalian ortholog of the Drosophila minibrain (Mnb) gene, essential for normal postembryonic neurogenesis. The Dyrk1A gene resides in the human chromosome 21q22.2 region, which is associated with DS anomalies, including mental retardation. In this study, we examined whether Dyrk1A interacts with alpha-synuclein and subsequently affects intracellular alpha-synuclein inclusion formation in immortalized hippocampal neuronal (H19-7) cells. Dyrk1A selectively binds to alpha-synuclein in transformed and primary neuronal cells. Alpha-synuclein overexpression, followed by basic fibroblast growth factor-induced neuronal differentiation, resulted in cell death. We observed that accompanying cell death was increased alpha-synuclein phosphorylation and intracytoplasmic aggregation. In addition, the transfection of kinase-inactive Dyrk1A or Dyrk1A small interfering RNA blocked alpha-synuclein phosphorylation and aggregate formation. In vitro kinase assay of anti-Dyrk1A immunocomplexes demonstrated that Dyrk1A could phosphorylate alpha-synuclein at Ser-87. Furthermore, aggregates formed by phosphorylated alpha-synuclein have a distinct morphology and are more neurotoxic compared with aggregates composed of unmodified wild type alpha-synuclein. These findings suggest alpha-synuclein inclusion formation regulated by Dyrk1A, potentially affecting neuronal cell viability.

Cell Survival↗

PubMed2006

Ni/NiO core/shell nanoparticles for selective binding and magnetic separation of histidine-tagged proteins.

Ni/NiO core/shell nanoparticles having high affinity with polyhistidine were synthesized by decomposition of a Ni surfactant complex followed by air oxidation. Ni/NiO nanoparticles showed selective and efficient binding to histidine-tagged proteins and easy separation by using a magnet. These provided a more convenient way to efficient purification of histidine-tagged proteins compared with the conventional Ni-NTA complex-bound resins and microbeads.

Histidine↗

PubMed2006

Molecular dynamics simulations of a cyclic-beta-(1-->2) glucan containing an alpha-(1-->6) linkage as a 'molecular alleviator' for the macrocyclic conformational strain.

The conformational preferences of a cyclic osmoregulated periplasmic glucan of Ralstonia solanacearum (OPGR), which is composed of 13 glucose units and linked entirely via beta-(1-->2) linkages excluding one alpha-(1-->6) linkage, were characterized by molecular dynamics simulations. Of the three force fields modified for carbohydrates that were applied to select a suitable one for the cyclic glucan, the carbohydrate solution force field (CSFF) was found to most accurately simulate the cyclic molecule. To determine the conformational characteristics of OPGR, we investigated the glycosidic dihedral angle distribution, fluctuation, and the potential energy of the glucan and constructed hypothetical cyclic (CYS13) and linear (LINEAR) glucans. All beta-(1-->2)-glycosidic linkages of OPGR adopted stable conformations, and the dihedral angles fluctuated in this energy region with some flexibility. However, despite the inherent flexibility of the alpha-(1-->6) linkage, the dihedral angles have no transition and are more rigid than that in a linear glucan. CYS13, which consists of only beta-(1-->2) linkages, is somewhat less flexible than other glycans, and one of its linkages adopts a higher energy conformation. In addition, the root-mean-square fluctuation of this linkage is lower than that of other linkages. Furthermore, the potential energy of glucans increases in the order of LINEAR, OPGR, and CYS13. These results provide evidence of the existence of conformational constraints in the cyclic glucan. The alpha-(1-->6)-glycosidic linkage can relieve this constraint more efficiently than the beta-(1-->2) linkage. The conformation of OPGR can reconcile the tendency for individual glycosidic bonds to adopt energetically favorable conformations with the requirement for closure of the macrocyclic ring by losing the inherent flexibility of the alpha-(1-->6)-glycosidic linkage.

Carbohydrate Conformation↗

PubMed2006

Amino acid sequence motifs and mechanistic features of the membrane translocation of alpha-synuclein.

Many lines of evidence suggest that alpha-synuclein can be secreted from cells and can penetrate into them, although the detailed mechanism is not known. In this study, we investigated the amino acid sequence motifs required for the membrane translocation of alpha-synuclein, and the mechanistic features of the phenomenon. We first showed that not only alpha-synuclein but also beta- and gamma-synucleins penetrated into live cells, indicating that the conserved N-terminal region might be responsible for the membrane translocation. Using a series of deletion mutants, we demonstrated that the 11-amino acid imperfect repeats found in synuclein family members play a critical role in the membrane translocation of these proteins. We further demonstrated that fusion peptides containing the 11-amino acid imperfect repeats of alpha-synuclein can transverse the plasma membrane, and that the membrane translocation efficiency is optimal when the peptide contains two repeat motifs. alpha-Synuclein appeared to be imported rapidly and efficiently into cells, with detectable protein in the cytoplasm within 5 min after exogenous treatment. Interestingly, the import of alpha-synuclein at 4 degrees C was comparable with the import observed at 37 degrees C. Furthermore, membrane translocation of alpha-synuclein was not significantly affected by treatment with inhibitors of endocytosis. These results suggest that the internalization of alpha-synuclein is temperature-insensitive and occurs very rapidly via a mechanism distinct from normal endocytosis.

Amino Acid Motifs↗

PubMed2006

Characterization of surface-confined alpha-synuclein by surface plasmon resonance measurements.

Urea-driven denaturation and renaturation of surface-bound alpha-synuclein are monitored by surface plasmon resonance (SPR) spectroscopy. The differential SPR angle shift (Delta Theta(SPR))(Net) enables us to estimate the Gibbs free energy change (DeltaG(o)) for the denaturation of the supported alpha-synuclein. DeltaG(o) for the denaturation of the supported alpha-synuclein, which is indirectly related to its biological activity can be increased significantly by the mixed self-assembled monolayers of 11-mercaptoundecanoic acid and 1,6-hexanedithiol. These SPR measurements of surface-bound biomolecules suggested herein can be further utilized to design effective biological scaffold for biosensor, biocatalyst, and possible diagnosis.

Coated Materials, Biocompatible↗

PubMed2005

Parkin ubiquitinates and promotes the degradation of RanBP2.

Parkinson disease (PD) is a common neurodegenerative disorder, which involves the deterioration of dopaminergic neurons in the pars compacta of the substantia nigra. The etiology of PD is still unknown, but recent identification of mutations in familial cases of PD has advanced the understanding of the molecular mechanisms of this neurological disease. Mutations in the parkin gene, which encodes for ubiquitin-protein ligase (E3), have been implicated in autosomal recessive juvenile Parkinsonism, an early onset and common familial form of PD. Here we reported that Parkin selectively binds to RanBP2, which is localized in the cytoplasmic filament of the nuclear pore complex and belongs to the small ubiquitin-related modifier E3 ligase family. We also demonstrated that RanBP2 becomes a target for Parkin E3 ubiquitin-ligase and is processed via Parkin-mediated ubiquitination and subsequent proteasomal degradation. Furthermore, Parkin controls the intracellular levels of sumoylated HDAC4, as a result of the ubiquitination and degradation of RanBP2. Our findings suggested that the intracellular levels of RanBP2 and its functional activity may be modulated by Parkin-mediated ubiquitination and proteasomal pathways.

Amino Acid Motifs↗

PubMed2005

Dequalinium-induced protofibril formation of alpha-synuclein.

alpha-Synuclein is the major constituent of Lewy bodies, a pathological signature of Parkinson disease, found in the degenerating dopaminergic neurons of the substantia nigra pars compacta. Amyloidosis generating the insoluble fibrillar protein deposition has been considered to be responsible for the cell death observed in the neurodegenerative disorder. In order to develop a controlling strategy toward the amyloid formation, 1,1'-(1,10-decanediyl)-bis-[4-a-mino-2-methylquinolinium] (dequalinium), was selected and examined in terms of its specific molecular interaction with alpha-synuclein. The protein was self-oligomerized by dequalinium, which gave rise to the ladder formation on N-[2-hydroxy-1,1-bis(hydroxymethyl)ethyl]glycine/SDS-PAGE in the presence of a coupling reagent of N-(ethoxycarbonyl)-2-ethoxy-1,2-dihydroquinoline. The double-headed structure of dequalinium with the two cationic 4-aminoquinaldinium rings was demonstrated to be critical for the protein self-oligomerization. The dequalinium-binding site was located on the acidic C-terminal region of the protein with an approximate dissociation constant of 5.5 mum. The protein self-oligomerization induced by the compound has resulted in the protofibril formation of alpha-synuclein before it has developed into amyloids. The protofibrils were demonstrated to affect the membrane intactness of liposomes, and they have also been shown to influence cell viability of human neuroblastoma cells. In addition, dequalinium treatment of the alpha-synuclein-overexpressing cells exerted a significant cell death. Therefore, it is pertinent to consider that dequalinium could be used as a molecular probe to assess toxic mechanisms related to the amyloid formation of alpha-synuclein. Ultimately, the compound could be employed to develop therapeutic and preventive strategies toward alpha-synucleinopathies including Parkinson disease.

Amyloid↗

PubMed2005

Proteolytic cleavage of extracellular secreted {alpha}-synuclein via matrix metalloproteinases.

Although alpha-synuclein is the main structural component of the insoluble filaments that form Lewy bodies in Parkinson disease (PD), its physiological function and exact role in neuronal death remain poorly understood. In the present study, we examined the possible functional relationship between alpha-synuclein and several forms of matrix metalloproteinases (MMPs) in the human dopaminergic neuroblastoma (SK-N-BE) cell line. When SK-N-BE cells were transiently transfected with alpha-synuclein, it was secreted into the extracellular culture media, concomitantly with a significant decrease in cell viability. Also the addition of nitric oxide-generating compounds to the cells caused the secreted alpha-synuclein to be digested, producing a small fragment whose size was similar to that of the fragment generated during the incubation of alpha-synuclein with various MMPs in vitro. Among several forms of MMPs, alpha-synuclein was cleaved most efficiently by MMP-3, and MALDI-TOF mass spectra analysis showed that alpha-synuclein is cleaved from its C-terminal end with at least four cleavage sites within the non-Abeta component of AD amyloid sequence. Compared with the intact form, the protein aggregation of alpha-synuclein was remarkably facilitated in the presence of the proteolytic fragments, and the fragment-induced aggregates showed more toxic effect on cell viability. Moreover, the levels of MMP-3 were also found to be increased significantly in the rat PD brain model produced by the cerebral injection of 6-hydroxydopamine into the substantia nigra. The present study suggests that the extracellularly secreted alpha-synuclein could be processed via the activation of MMP-3 in a selective manner.

Alzheimer Disease↗

PubMed2004

Alpha-synuclein has structural and functional similarities to small heat shock proteins.

The aggregation and fibrillization of alpha-synuclein, a major component of Lewy bodies, is a key event in Parkinson's disease. Although the mechanisms of fibrils formation are largely investigated, physiological function of alpha-synuclein is not yet clearly elucidated. Here, we showed that C-terminal region of alpha-synuclein is similar to alpha-crystalline domain of small heat shock proteins. In our experiments, alpha-synuclein, like small heat shock proteins, protected cellular proteins from denaturation, and confer Escherichia coli cellular tolerances against thermal- and oxidative-stresses.

14-3-3 Proteins↗

PubMed2004

Beta-synuclein exhibits chaperone activity more efficiently than alpha-synuclein.

Beta-synuclein exhibits high sequence homology and structural similarity with alpha-synuclein, a protein implicated in the pathogenesis of Parkinson's disease. We investigated the chaperone function of beta-synuclein and its anti-fibrillar activity in comparison with alpha-synuclein. beta-Synuclein suppressed the heat-induced aggregation of aldolase, alcohol dehydrogenase, and citrate synthase, and its anti-aggregative activity was remarkably higher than that of alpha-synuclein. Heat-induced inactivation of citrate synthase was significantly protected by beta-synuclein. Moreover, beta-synuclein inhibited the amyloid formation of both Abeta(1-40) and alpha-synuclein. It is, therefore, suggested that beta-synuclein can prevent abnormal protein aggregations more effectively than alpha-synuclein by acting as a molecular chaperone.

Amino Acid Sequence↗

PubMed2004

Phthalocyanine tetrasulfonates affect the amyloid formation and cytotoxicity of alpha-synuclein.

Alpha-synuclein is a pathological component of Parkinson's disease by constituting the filamentous component of Lewy bodies. Phthalocyanine (Pc) effects on the amyloidosis of alpha-synuclein have been examined. The copper complex of phthalocyanine tetrasulfonate (PcTS-Cu(2+)) caused the self-oligomerization of alpha-synuclein while Pc-Cu(2+) did not affect the protein, indicating that introduction of the sulfonate groups was critical for the selective protein interaction. The PcTS-Cu(2+) interaction with alpha-synuclein has occurred predominantly at the N-terminal region of the protein with a K(d) of 0.83 microM apart from the hydrophobic NAC (non-Abeta component of Alzheimer's disease amyloid) segment. Phthalocyanine tetrasulfonate (PcTS) lacking the intercalated copper ion also showed a considerable affinity toward alpha-synuclein with a K(d) of 3.12 microM, and its binding site, on the other hand, was located at the acidic C-terminus. These mutually exclusive interactions between PcTS and PcTS-Cu(2+) toward alpha-synuclein resulted in distinctive features on the kinetics of protein aggregation, morphologies of the final aggregates, and their in vitro cytotoxicities. The PcTS actually suppressed the fibrous amyloid formation of alpha-synuclein, but it produced the chopped-wood-looking protein aggregates. The aggregates showed rather low toxicity (9.5%) on human neuroblastoma cells (SH-SY5Y). In fact, the PcTS was shown to effectively rescue the cell death of alpha-synuclein overexpressing cells caused by the lactacystin treatment as a proteasome inhibitor. The anti-aggregative and anti-amyloidogenic properties of PcTS were also demonstrated with alcohol dehydrogenase, glutathione S-transferase, and amyloid beta/A4 protein under their aggregative conditions. The PcTS-Cu(2+), on the other hand, promoted the protein aggregation of alpha-synuclein, which gave rise to the fibrillar protein aggregates whose cytotoxicity became significant to 35.8%. Taken together, the data provided in this study indicate that PcTS/PcTS-Cu(2+) could be considered as possible candidates for the development of therapeutic or prophylactic strategies against the alpha-synuclein-related neurodegenerative disorders.

Amyloid↗

PubMed2003

Parkin cleaves intracellular alpha-synuclein inclusions via the activation of calpain.

Mutations in the alpha-synuclein and parkin genes cause heritable forms of Parkinson's disease. In the present study, we examined the possible functional relationship between the parkin and alpha-synuclein genes in a conditionally immortalized embryonic hippocampal cell (H19-7) line. Whereas transient transfection of alpha-synuclein into neuronal H19-7 cells caused the formation of its intracytoplasmic inclusions and a significant cell death, the combined overexpression of parkin restored the alpha-synuclein-induced decrease in cell viability to control levels. In addition, the overexpression of parkin was found to generate selective cleavage of alpha-synuclein. Furthermore, the cytoprotective effect of parkin on alpha-synuclein-induced cell death was not inhibited in the presence of a proteasome inhibitor. Interestingly, the overexpression of parkin induced the activation of an intracellular cysteine protease, calpain, but not caspase, and the cytoprotective effect of parkin on alpha-synuclein cytotoxicity was significantly inhibited by the presence of calpain-specific inhibitors. In conclusion, our results suggest that parkin accelerates the degradation of alpha-synuclein via the activation of the nonproteasomal protease, calpain, leading to the prevention of alpha-synuclein-induced cell death in embryonic hippocampal progenitor cells.

Animals↗

PubMed2003

Oxidized glutathione stimulated the amyloid formation of alpha-synuclein.

alpha-Synuclein is the major filamentous constituent of Lewy bodies found in Parkinson's disease (PD). The amyloid formation of alpha-synuclein was significantly facilitated by oxidized glutathione (GSSG) as the lag period of the aggregation kinetics was shortened by 2.5-fold from its absence. Reduced glutathione (GSH), on the other hand, did not influence the lag phase although it increased the final amyloid formation. The GSSG stimulation was specific for not only alpha-synuclein but also its intactness. The preferred GSSG interaction of alpha-synuclein to GSH was also demonstrated with dissociation constants of 0.53 and 43.5 mM, respectively. It is suggested that the oxidative stress favoring the GSSG generation from GSH could result in the augmented amyloid formation of alpha-synuclein, which ought to be related to the pathogenesis of PD.

Amyloid↗

PubMed2003

Lipid interaction of alpha-synuclein during the metal-catalyzed oxidation in the presence of Cu2+ and H2O2.

Alpha-synuclein co-exists with lipids in the Lewy bodies, a pathological hallmark of Parkinson's disease. Molecular interaction between alpha-synuclein and lipids has been examined by observing lipid-induced protein self-oligomerization in the presence of a chemical coupling reagent of N-(ethoxycarbonyl)-2-ethoxy-1,2-dihydroquinoline. Lipids such as phosphatidic acid, phosphatidylinositol, phosphatidylserine, phosphatidylethanolamine, and even arachidonic acid induced the self-oligomerization whereas phosphatidylcholine did not affect the protein. Because the oligomerizations occurred from critical micelle concentrations of the lipids, the self interaction of alpha-synuclein was shown to be a lipid-surface dependent phenomenon with head group specificity. By employing beta-synuclein and a C-terminally truncated alpha-synuclein (alpha-syn97), the head-group dependent self-oligomerization was demonstrated to occur preferentially at the N-terminal region while the fatty acid interaction leading to the protein self-association required the presence of the acidic C-terminus of alpha-synuclein. In the presence of Cu2+ and H2O2, phosphatidylinositol (PI), along with other acidic lipids, actually enhanced the metal-catalyzed oxidative self-oligomerization of alpha-synuclein. The dityrosine crosslink formation responsible for the PI-enhanced covalent self-oligomerization was more sensitive to variation of copper concentrations than that of H2O2 during the metal-catalyzed oxidation. The enhancement by PI was shown to be due to facilitation of copper localization to the protein because actual binding affinity between copper and alpha-synuclein increased from Kd of 44.7 microm to 5.9 microm in the presence of the lipid. Taken together, PI not only affects alpha-synuclein to be more self-interactive by providing the lipid surface, but also enhances the metal-catalyzed oxidative protein self-oligomerization by facilitating copper localization to the protein when the metal and H2O2 are provided. This observation therefore could be implicated in the formation of Lewy bodies as lipids and metal-catalyzed oxidative stress have been considered to be a part of pathological causes leading to the neurodegeneration.

Catalysis↗

PubMed2002

Structural and functional implications of C-terminal regions of alpha-synuclein.

Aggregation of alpha-synuclein is thought to play a major role in the pathogenesis of Parkinson's disease (PD), which is characterized by the presence of intracytoplasmic Lewy bodies (LB) in the brain. alpha-Synuclein and its deletion mutants are largely unfolded proteins with random coil structures as revealed by CD spectra, fluorescence spectra, gel filtration chromatography, and ultracentrifugation. On the basis of its highly unfolded and flexible conformation, we have investigated the chaperone-like activity of alpha-synuclein in vitro. In our experiments, alpha-synuclein inhibited the aggregation of model substrates and protected the catalytic activity of alcohol dehydrogenase and rhodanese during heat stress. In addition, alpha-synuclein inhibited the initial aggregation of reduced/denatured lysozyme on the refolding pathway. Interestingly, deletion of the C-terminal regions led to the abolishment of chaperone activity, although largely unstructured conformations are maintained. Moreover, alpha-synuclein could inhibit the aggregation of various Escherichia coli cellular proteins during heat stress, and C-terminal deletion mutants could not provide any protection to these cellular proteins. Results with synthetic C-terminal peptides and C-terminal deletion mutants suggest that the second acidic repeat, (125)YEMPSEEGYQDYEPEA(140), is important for the chaperone activity of alpha-synuclein, and C-terminal deletion leads to the facilitated aggregation with the elimination of chaperone activity.

Alcohol Dehydrogenase↗

PubMed2002

Evidence that alpha-synuclein functions as a negative regulator of Ca(++)-dependent alpha-granule release from human platelets.

alpha-Synuclein has been implicated in the pathogenesis of Parkinson disease (PD) and related neurodegenerative disorders. More recently, it has been suggested to be an important regulatory component of vesicle transport in neuronal cells. alpha-Synuclein is also highly expressed in platelets and is loosely associated with the membrane of the secretory alpha-granules. However, the functional significance of these observations is unknown. In this study, the possible function of alpha-synuclein in vesicle transport, with particular regard to alpha-granule release from the platelets, was investigated. The results showed that ionomycin- or thrombin-induced alpha-granule secretion was inhibited by exogenous alpha-synuclein addition in a dose-dependent manner. However, [(3)H]5-HT release from the dense granules and hexosaminidase release from the lysosomal granules were not affected. Two point mutants (A30P and A53T) found in some familial types of PD, in addition to beta-synuclein and alpha-synuclein112, effectively inhibited PF4 release from the alpha-granules. However, the deletion mutants, which completely lacked either the N-terminal region or the C-terminal tail, did not affect alpha-granule release. Interestingly, exogenously added alpha-synuclein appeared to enter the platelets but did not change the Ca(++) level in the platelets at the resting state and the increase in the Ca(++) level on stimulation. Electron microscopy also supported that alpha-synuclein inhibits alpha-granule release. These results suggest that alpha-synuclein may function as a specific negative regulator of alpha-granule release in platelets.

Blood Platelets↗

PubMed2002

Purification and characterization of a prothrombin-activating protease from Nephila clavata.

We report upon the purification and characterization of a novel prothrombin-activating enzyme from the body fluid (total homogenates of isolated digestive tract without eggs, spinnerets and silk glands) of the spider, Nephila clavata by a combination of acetone fractionation, ion exchange, and Soybean trypsin inhibitor-Sepharose chromatography. Analysis of the purified enzyme with SDS-PAGE and gel filtration revealed a single polypeptide chain with an apparent molecular weight of 24kDa. The proteolytic activity of the enzyme was stable up to 50 degrees C, however, it became unstable over 55 degrees C. The enzyme had an optimum pH of 8, and Ca(2+) was not required for the enzyme activity. According to inhibition profiles obtained with several serine protease inhibitors such as PMSF and benzamidine, the purified protease is a member of the serine proteases. Bz-Ile-Glu(gamma-OR)- Gly-Arg-pNA and Z-Arg-Gly-Arg-pNA which are known as substrates for factor Xa, were hydrolyzed favorably by the enzyme. And the Nephila protease could produce thrombin from prothrombin at nM range, and form the turbid ring using fibrinogen-agarose plate. The results obtained confirmed that the purified protease is a potent prothrombin-activating activity belonging to the family of serine protease.

Animals↗