Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.24.3 (collagenase)
18,340 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

During the course of human pregnancy, there is a marked increase in insulin-like growth factor (IGF) binding protein (IGFBP)-3 protease activity in maternal serum that is first evident at 6 weeks of gestation, persists through term, and returns to nonpregnancy levels by day 5 postpartum. This protease activity cleaves IGFBP-3 into smaller fragments that have markedly reduced affinity for the IGFs. To date, the precise identity and cellular origin of the pregnancy-associated serum IGFBP-3 protease have not been established. To investigate whether placental and/or decidual tissues, which uniquely develop during pregnancy, may be sources of the pregnancy-associated serum IGFBP protease, we examined the secretion of IGFBP-3 protease in vitro by isolated human cytotrophoblasts or fibroblasts from second trimester placentae and by in vitro decidualized human endometrial stromal cells. Cytotrophoblasts were either cultured alone, which favors aggregation and fusion, or cocultured with decidualized endometrial stromal cells, which favors differentiation to an invasive phenotype. IGFBP-3 protease activity was detected in trophoblast, but not in placental fibroblast or decidualized endometrial cultures, and was also present in trophoblast-endometrial cocultures. Western ligand blot and Western immunoblot analyses showed that most of the endogenous IGFBP-3 in trophoblast cultures was in the form of low molecular weight fragments with reduced IGF binding affinity. The substrate specificity of the trophoblast-derived protease was identical to that in pregnancy serum, showing activity against IGFBP-2, -3, and -4, but being inactive against IGFBP-1. IGFBP-3 proteolysis by both pregnancy serum and trophoblast conditioned medium showed a major peak of activity at neutral pH. The trophoblast-derived activity caused time-and temperature-dependent proteolysis of IGFBP-3 into fragments of identical size as those produced by pregnancy serum, and also shared its sensitivity to protease inhibitors: highly sensitive to EDTA and o-phenanthroline, partially sensitive to the serine protease inhibitors AEBSF and aprotinin, and insensitive to alpha2-antiplasmin, and to aspartic and cysteine protease inhibitors. IGFBP-3 proteolysis by both pregnancy serum and trophoblast conditioned medium was also insensitive to tissue inhibitor of metalloproteinase-1, precluding the involvement of the matrix metalloproteinases. In contrast, both the pregnancy serum- and trophoblast-derived proteases were preferentially inhibited by a hydroxamic acid derivative with selective activity against the disintegrin-metalloproteinase tumor necrosis factor-alpha converting enzyme. This study shows that placental trophoblasts produce an IGFBP-3 protease with characteristics very similar to the activity found in pregnancy serum and indicates these cells at the maternal-fetal interface are a potential source of the pregnancy-associated serum IGFBP-3 protease. The findings further suggest that the main IGFBP-3 protease activity in both pregnancy serum and trophoblast conditioned medium may correspond to a disintegrin-metalloproteinase type enzyme.
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PMID:Human placental trophoblasts secrete a disintegrin metalloproteinase very similar to the insulin-like growth factor binding protein-3 protease in human pregnancy serum. 1065 Sep 48

The dominant proteins released by Ascaris suum during development in vitro from the L3 to L4 stage were identified as collagenous cuticular proteins by sequence analysis and susceptibility to digestion by collagenase. Under reducing sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), the collagen proteins separated into 3 groups with molecular weights estimated at 32 kDa, 54-60 kDa, and 71-91 kDa. The 32-kDa protein represents monomeric collagen; the 54-60- and 71-91-kDa components represent dimeric and trimeric forms, respectively, polymerized by nonreducible cross-links. Furthermore, the release of these forms of collagen was developmentally regulated, as exemplified by a sequential temporal progression from monomeric to dimeric to trimeric forms in association with the in vitro transition from L3 to L4. The data suggest that collagen released in vitro during development of A. suum L3 to L4 reflects the increased translation of collagen gene products and their initial assembly into higher molecular weight molecules associated with the synthesis of the L4 cuticle. A biotinylated dipeptidyl fluoromethylketone cysteine protease inhibitor (Bio-phe-ala-FMK) bound specifically to the 32-kDa collagen and, to a lesser extent, to a 30-kDa protein; binding was dependent on the presence of dithiothreitol (DTT) and was prevented by iodoacetamide. Because cysteine residues play an essential role in the initial assembly of the collagen monomers into the higher molecular weight oligomers present in the mature nematode cuticle, inhibition of molting of A. suum L3 to L4 by the cysteine protease inhibitor Z-phe-ala-FMK might be due to its binding to thiol groups of collagen monomers during a critical phase of collagen assembly. Prevention of cystine cross-links during this critical period of cuticle assembly by peptide-FMK inhibitors may represent a potential control mechanism having a novel mechanism of action.
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PMID:Cuticular collagen synthesis by Ascaris suum during development from the third to fourth larval stage: identification of a potential chemotherapeutic agent with a novel mechanism of action. 1169 81

Cathepsin L (CTSL) is a lysosomal cysteine protease with potent elastase and collagenase activities. Its high activity in the uterine lumen during the period of placental attachment has led to speculation that CTSL may play an important role during embryonic implantation in the pig. Cathepsins have also been implicated in blastocyst implantation in other species like cat, rat and man. We isolated a PAC clone containing the porcine CTSL gene and determined the complete DNA sequence of the gene, which spans about 5.6 kb and consists of eight exons. The CTSL transcript encodes a primary peptide of 334 amino acids sharing 73-78% identity with other mammalian cathepsin L precursor proteins. Based on fluorescence in situ hybridization and radiation hybrid mapping, the porcine CTSL gene was assigned to chromosome 10q11--> q12.
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PMID:Characterization and comparative mapping of the porcine CTSL gene indicates a novel synteny between HSA9q21-->q22 and SSC10q11-->q12. 1197 77

Cathepsin K, a lysosomal papain-like cysteine protease, forms collagenolytically highly active complexes with chondroitin sulfate and represents the most potent mammalian collagenase. Here we demonstrate that complex formation with glycosaminoglycans (GAGs) is unique for cathepsin K among human papain-like cysteine proteases and that different GAGs compete for the binding to cathepsin K. GAGs predominantly expressed in bone and cartilage, such as chondroitin and keratan sulfates, enhance the collagenolytic activity of cathepsin K, whereas dermatan, heparan sulfate, and heparin selectively inhibit this activity. Moreover, GAGs potently inhibit the collagenase activity of other cysteine proteases such as cathepsins L and S at 37 degrees C. Along this line MMP1-generated collagen fragments in the presence of GAGs are stable against further degradation at 28 degrees C by all cathepsins but cathepsin K, whereas thermal destabilization at 37 degrees C renders the fragments accessible to all cathepsins. These results suggest a novel mechanism for the regulation of matrix protein degradation by GAGs. It further implies that cathepsin K represents the only lysosomal collagenolytic activity under physiologically relevant conditions.
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PMID:Regulation of collagenase activities of human cathepsins by glycosaminoglycans. 1464 29

Growth and rupture of abdominal aortic aneurysms (AAAs) result from increased collagen turnover. Collagen turnover critically depends on specific collagenases that cleave the triple helical region of fibrillar collagen. As yet, the collagenases responsible for collagen degradation in AAAs have not been identified. Increased type I collagen degradation products confirmed collagen turnover in AAAs (median values: <1, 43, and 108 ng/mg protein in control, growing, and ruptured AAAs, respectively). mRNA and protein analysis identified neutrophil collagenase [matrix metalloproteinase (MMP)-8] and cysteine collagenases cathepsin K, L, and S as the principle collagenases in growing and ruptured AAAs. Except for modestly increased MMP-14 mRNA levels, collagenase expression was similar in growing and ruptured AAAs (anterior-lateral wall). Evaluation of posttranslational regulation of protease activity showed a threefold increase in MMP-8, a fivefold increase in cathepsins K and L, and a 30-fold increase in cathepsin S activation in growing and ruptured AAAs. The presence of the osteoclastic proton pump indicated optimal conditions for extracellular cysteine protease activity. Protease inhibitor mRNA expression was similar in AAAs and controls, but AAA protein levels of cystatin C, the principle cysteine protease inhibitor, were profoundly reduced (>80%). We found indications that this secondary deficiency relates to cystatin C degradation by (neutrophil-derived) proteases.
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PMID:Collagen degradation in the abdominal aneurysm: a conspiracy of matrix metalloproteinase and cysteine collagenases. 1732 67

Actinidin is a cysteine protease abundant in Kiwifruit. This enzyme is known as a meat-tenderizing protease. In this project, actinidin was purified from kiwifruit by salt precipitation and ion exchange chromatography. Collagenolytic effect of the purified enzyme was tested in four different buffer systems. Thereafter, the enzyme was used for isolation and culture of cells from three different tissues: endothelial cells from human umbilical vein, hepatocytes from rat liver, and thymic epithelial cells from rat thymus. Our results revealed that actinidin can hydrolyze collagen types I and II at neutral and alkaline buffers. Furthermore, actinidin compared with type II or IV collagenase isolated intact human umbilical vein endothelial cells, hepatocytes, and thymic epithelial cells with viability more than 90%. These results address a novel and valuable collagenase, which can be used efficiently for hydrolysis of collagen and isolation of different cell populations from various solid tissues.
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PMID:Kiwifruit actinidin: a proper new collagenase for isolation of cells from different tissues. 1845 44

Cathepsin K is a cysteine protease of the papain family that cleaves triple-helical type II collagen, the major structural component of the extracellular matrix of articular cartilage. In osteoarthritis (OA), the anabolic/catabolic balance of articular cartilage is disrupted with the excessive cleavage of collagen II by collagenases or matrix metalloproteinases. A polyclonal antibody against a C-terminal neoepitope (C2K) generated in triple-helical type II collagen by the proteolytic action of cathepsin K was prepared and used to develop an enzyme-linked immunosorbent assay to study the generation of this epitope and the effects of its presence in normal adult and osteoarthritic femoral condylar articular cartilage. The generation of the C2K epitope in explant culture and the effect of a specific cathepsin K inhibitor were studied. The neoepitope, which is not generated by the collagenase matrix metalloproteinase-13, increased with age in articular cartilage and was significantly elevated in osteoarthritic cartilage compared with adult nonarthritic cartilage. Moreover, in explants from three of eight OA patients, the generation of the neoepitope in culture was significantly reduced by a specific, nontoxic inhibitor of cathepsin K. These data suggest that cathepsin K is involved in the cleavage of type II collagen in human articular cartilage in certain OA patients and that it may play a role in both OA pathophysiology and the aging process.
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PMID:Cleavage of type II collagen by cathepsin K in human osteoarthritic cartilage. 1851 17

The involvement of the mannose-induced Acanthamoeba cytopathic protein (MIP-133) in tissue injury and activation of metalloproteinase of corneal and stromal cells was examined in vitro. Activation of MMP-1, MMP-2, MMP-3, and MMP-9 induced by MIP-133 on human corneal epithelial and stromal cell cultures was examined by reverse transcriptase polymerase chain reaction (RT-PCR) and ELISA. MMP-1, MMP-2, MMP-3, and MMP-9 mRNA were expressed in both cultured human corneal epithelial and stromal cells. When the epithelial cells were exposed to MIP-133 protein, the mRNA expression for MMP-1 and MMP-9 was unchanged. However, the transcript for MMP-2 and MMP-3 was decreased by 2-fold. By contrast, the expression of MMP-2 and MMP-3 was significantly upregulated (2- to 4-fold) in the corneal stromal cells 1, 4, and 8h after MIP-133 stimulation. At the protein level, there was no significant difference in the level of MMPs between the corneal epithelial cells before and after stimulation with MIP-133. By contrast, the levels of MMP-2 and MMP-3 were significantly higher in the corneal stromal cells stimulated with MIP-133. The supernatants from corneal stromal cells stimulated with MIP-133 were incubated with PMSF and MIP-133 antibody and the level of MMP-2 was measured by ELISA. Activation of MMP-2 by MIP-133 was inhibited in the supernatants pretreated with the serine protease inhibitor, PMSF, and anti-MIP-133. Supernatants pretreated with the cysteine protease inhibitor E6 or control antibody produced the same amount of MMP-2 as the untreated supernatants. To verify possible homology between MMPs and Acanthamoeba castellanii proteases, the mRNA from A. castellanii was prepared and analyzed for the expression of MMP genes by PT-PCR. The results showed that A. castellanii did not express mRNA for MMP-1, MMP-2, MMP-3, or MMP-9. Thus, A. castellanii mRNA does not cross-react with human MMPs. Furthermore, ELISA was used to determine the cross-reactivity of MMP antibodies with the MIP-133 protein. Monoclonal antibodies against MMPs did not cross-react with either the MIP-133 protein or BSA (negative control antigen). The results indicate that the MIP-133 protein modulates MMP-2 and -3 expression differently in human corneal epithelial and stromal cells.
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PMID:Modulation of corneal and stromal matrix metalloproteinase by the mannose-induced Acanthamoeba cytolytic protein. 1862 71

Cathepsin K is a highly potent collagenase and the predominant papain-like cysteine protease expressed in osteoclasts. Cathepsin K deficiencies in humans and mice have underlined the central role of this protease in bone resorption and, thus, have rendered the enzyme as an attractive target for anti-resorptive osteoporosis therapy. In the past decade, a lot of efforts have been made in developing highly potent, selective and orally applicable cathepsin K inhibitors. Some of these inhibitors have passed preclinical studies and are presently in clinical trials at different stages of advancement. The development of the inhibitors and preliminary results of the clinical trials revealed problems and lessons concerning the in situ specificity of the compounds and their tissue targeting. In this review, we briefly summarize the history of cathepsin K research and discuss the current development of cathepsin K inhibitors as novel anti-resorptives for the treatment of osteoporosis. We also discuss potential off-target effects of cathepsin K inhibition and alternative applications of cathepsin K inhibitors in arthritis, atherosclerosis, blood pressure regulation, obesity and cancer.
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PMID:Cathepsin K inhibitors for osteoporosis and potential off-target effects. 1938 76

Local recurrence continues to limit survival in medulloblastoma patients, largely related to the persistence of invasive cells at the site of tumour resection and leptomeningeal dissemination. Given the relative dearth of understanding of causative mechanisms behind the invasiveness of medulloblastomas, and a general lack of validated in vitro models with which to study them, our objectives were (1) to obtain quantitative data on the invasiveness of five distinct medulloblastoma cell lines within a 3-dimensional in vitro collagen-based model; and (2) to characterize some of the mechanisms behind invasion, specifically striving to identify proteolytic processes that occur as medulloblastoma cells disrupt and thereby invade the normal tissue surrounding them, and specific inhibitors of these proteolytic enzymes. Five different medulloblastoma cell lines (UW228-1, 2 and 3; Daoy, and Madsen) were implanted onto a 3-dimensional, type I collagen gel assay to assess tumour invasion distance and mean doubling time over 5 days. Proteolytic activity was assessed against collagen types I and IV by measuring the degradation of 3H-collagen I and IV to products soluble in 100% w/v trichloroacetic acid; and general (neutral) proteolytic activity evaluated by measuring the degradation of 3H-albumin. In other experiments, cells were pre-exposed to a variety of protease inhibitors, including inhibitors of metalloproteinases and cysteine, serine and aspartic proteases, and then plated to identify any inhibition of invasion. Inter-group differences in mean invasion distance were assessed by means of Student's t-tests for non-paired subjects, with P < 0.05 set as the threshold for statistical significance. For the inhibitor studies, an inhibition index, called the inhibitory concentration 50, IC-50, was calculated by performing a regression analysis for each inhibitor tested over a range of concentrations, for each cell line. Within hours of implantation, individual cells readily detached from the surface of the cell aggregates and invaded the collagen matrix, to distances of up to 1,200 mum and at rates of up to 300-mum per day; the UW228-1 cell line clearly was less invasive than the other four cell lines. Proteolytic activity was identified against collagen type I, but not against collagen type IV or albumin; but there was no apparent correlation between invasion distance and either cell doubling time or the amount of collagen type I proteolytic activity. Both metalloproteinase inhibitors suppressed tumour invasion, as did one of two cysteine protease inhibitors; but there was no tumour suppression with either serine or aspartic protease inhibition. MMP-1 and 2, and TIMP-1 and 2 all were detectable by Western blot analysis. Medulloblastoma cell invasiveness within the 3-dimensional model used here appears to depend upon a combination of metalloproteinase and cysteine protease activity, a finding that may suggest areas for potential future clinical investigation and therapy.
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PMID:The invasiveness of five medulloblastoma cell lines in collagen gels. 1984 23


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