Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.6.4 (chondroitinase)
 2,039 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have analyzed the binding of thrombin, a serine protease with central roles in hemostasis, to the subendothelial extracellular matrix (ECM) produced by cultured endothelial cells. This substrate provides a thrombogenic surface where hemostasis is initiated. Binding was saturable and equilibrium was achieved after 3 h incubation with 125I-alpha-thrombin. Scatchard analysis of thrombin binding revealed the presence of 5.1 X 10(9) binding sites per squared millimeter ECM, with an apparent Kd of 13 nM. The catalytically blocked enzyme, diisofluorophosphate (DIP)-alpha-thrombin competed efficiently with 125I-alpha-thrombin, indicating that the binding was independent of its catalytic site. Moreover, high concentrations of the synthetic tetradecapeptide, representing residues 367-380 of thrombin B chain (the macrophage mitogenic domain of thrombin), competed with thrombin binding to ECM, indicating that the binding site may reside in the vicinity of "loop B" region. Thrombin binds to dermatan sulfate in the ECM, as demonstrated by the inhibition of 125I-alpha-thrombin binding to ECM pretreated with chondroitinase ABC, but not with heparitinase or chondroitinase AC. This stands in contrast to 125I-FGF (fibroblast growth factor) binding to ECM, which was inhibited by heparitinase but not by chondroitinase ABC, ECM-bound thrombin exhibits an exposed proteolytic site as monitored by the Chromozyme TH assay and by its ability to convert fibrinogen to a fibrin clot and to induce platelet activation as indicated by 14C-serotonin release. ECM-bound thrombin failed to form a complex with its major circulating inhibitor-antithrombin III (AT III), compared with rapid complex formation with soluble thrombin. We propose that thrombin binds to subendothelial ECM where it remains functionally active, localized, and protected from inactivation by circulating inhibitors.
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PMID:Binding of thrombin to subendothelial extracellular matrix. Protection and expression of functional properties. 279 47

Processing and metabolism of beta-amyloid precursor protein (APP) and generation of a variety of beta-amyloid (Abeta) peptides in the human brain is essentially associated with pathophysiology of Alzheimer's disease (AD). APP degradation activity of the 68 kDa serine protease, which was originally prepared from familial AD lymphoblastoid cells and harbors beta-secretase-like activity, was analyzed by Western blot using anti Abeta 1/40 antibody and anti APP cytoplasmic domain (CT) antibody. Native lymphocyte APP (LAPP) prepared from normal or AD-derived lymphoblastoid cells was degraded by the protease, generating a 16 kDa Abeta-bearing C-terminal fragment of APP. N-terminal amino acid sequencing of the fragment indicated that the protease cleaves LAPP at the Abeta-N-terminus. When the LAPP was treated with chondroitinase ABC prior to proteolysis, the activity to generate the fragment was inhibited, but pretreatment with heparitinase resulted in no effect. Native hippocampal APP prepared from normal brain, however, did not generate the 16 kDa peptide by the protease treatment. These results suggest that the process of APP degradation and Abeta-peptides generation, including beta-secretase activity, is associated with tissue specificity of both APP substrate and proteases. They also indicate that sulfated glycoconjugates attached to a portion of APP isoforms may play a role as a molecular determinant in the proteolysis.
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PMID:The 68K protease has beta-secretase-like activity for lymphocyte precursor protein but not for brain substrate. 1067 89

Atrial natriuretic peptide (ANP) is a cardiac hormone essential for normal blood pressure and cardiac function. Corin is a transmembrane serine protease that activates ANP. Recently, we identified proprotein convertase subtilisin/kexin-6 (PCSK6), also called PACE4, as the long-sought corin activator. Both corin and PCSK6 are expressed in cardiomyocytes, but corin activation occurs only on the cell surface. It remains unknown if cell membrane association is needed for PCSK6 to activate corin. Here we expressed corin deletion mutants in HEK293 cells to analyze the domain structures required for PCSK6-mediated activation. Our results show that soluble corin lacking the transmembrane domain was activated by PCSK6 in the conditioned medium but not intracellularly. Recombinant PCSK6 also activated the soluble corin under cell-free conditions. Moreover, PCSK6-mediated corin activation was not enhanced by cell membrane fractions. These results indicate that cell membrane association is unnecessary for PCSK6 to activate corin. Experiments with monensin that blocks PCSK6 secretion and immunostaining indicated that the soluble corin and PCSK6 were secreted via different intracellular pathways, which may explain the lack of corin activation inside the cell. We also found that the protein domains in the corin pro-peptide region were dispensable for PCSK6-mediated activation and that addition of heparan sulfate and chondroitin sulfate or treatment with heparinase or chondroitinase did not alter corin activation by PCSK6 in HEK293 cells. Together, our results provide important insights into the molecular and cellular mechanisms underlying PCSK6-mediated corin activation that is critical for cardiovascular homeostasis.
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PMID:Functional analysis of corin protein domains required for PCSK6-mediated activation. 2918 Mar 4