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)

The mechanism of the biosynthesis of albumin was studied in cell suspensions from rat liver. The cells were prepared by continuous perfusion of the liver in situ with 0.05% collagenase and 0.10% hyaluronidase and incubated under conditions optimized for the incorporation of amino acids into protein. Seven minutes after starting the incubation L-[1-14C]leucine was added, followed after 25 min by a 15 or 30-min chase with an 830-fold excess of non-radioactive L-leucine. Total protein, an albumin-like protein, and albumin were isolated from samples withdrawn immediately of total protein was found to remain constant after addition of the non-radioactive L-leucine, whereas that of the albumin-like protein decreased and that of albumin increased with incubation time. The increase in albumin radioactivity accounted for the decrease in radioactivity of the albumin-like protein, suggesting that the latter is a precursor of albumin. The precursor protein differed from albumin by an oligopeptide extension at the N-terminal end.
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PMID:Synthesis of albumin via a precursor protein in cell suspensions from rat liver. 126 47

Murine interleukin 1 (IL-1) is initially synthesized as a 270-amino acid precursor protein. Guided by amino-terminal end sequence analyses of mouse macrophage-derived IL-1, it was shown that expression of the carboxyl-terminal 156 amino acids (i.e., amino acids 115-270) of this precursor in Escherichia coli yields biologically active recombinant IL-1 (rIL-1) protein. To answer questions about precursor processing and the size of the smallest biologically active IL-1 fragment, we have engineered deletions of the rIL-1 (115-270) gene to encode two amino-terminal deletion analogs, rIL-1 (131-270) and rIL-1 (144-270), and a carboxyl-terminal deletion analog, rIL-1 (131-257, 270). The analogs were produced in E. coli, purified to homogeneity, and assayed for biological activity on murine thymocytes, human rheumatoid synovial cells, and human dermal fibroblasts and for their ability to bind to IL-1 receptors on murine EL-4 thymoma cells. The amino-terminal deletion analog rIL-1 (131-270) possessed a specific activity in the murine thymocyte proliferation assay equivalent to that of the 115-270 parent protein and exhibited significant biological activity in stimulating the production of collagenase and prostaglandin E2 by synovial cells and fibroblasts. The more extensive amino-terminal deletion analog rIL-1 (144-270) was inactive in all biological assays and failed to compete in the receptor binding assay. The carboxyl-terminal deletion analog rIL-1 (131-257, 270) competed less efficiently (by a factor of 100) in the receptor binding assay, retained weak biological activity on synovial cells and fibroblasts, and only demonstrated full intrinsic activity in the thymocyte proliferation assay when 100-200 times more protein was assayed. These results suggest that biologically active murine IL-1 polypeptides are at least 127 amino acids long and are derived from the carboxyl terminus of the 270-amino acid precursor. Furthermore, it appears that the integrity of the carboxyl terminus of the 270-amino acid precursor is important for activity but that different amino termini can be utilized to generate molecules with equivalent specific activities. This amino-terminal end flexibility supports a processing model for IL-1 maturation that partially explains IL-1 polypeptide heterogeneity.
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PMID:Structure-function analysis of murine interleukin 1: biologically active polypeptides are at least 127 amino acids long and are derived from the carboxyl terminus of a 270-amino acid precursor. 302 89

A polyclonal antiserum was prepared in rabbits against the structural glycoprotein (SGP) complex previously isolated from a bacterial collagenase digest of bovine corneal stroma (R. Alper, Curr. Eye Res. 2:479, 1983). Direct and indirect enzyme-linked immunosorbent assays indicated that the antiserum was specific for the SGP-complex and did not react with Types I, III and IV collagen, fibronectin, laminin or actin. Immunoblot experiments indicated that the antiserum reacted with all of the components of the SGP-complex as well as with the cell matrix laid down by bovine keratocytes in culture. An attempt was made to isolate individual antibodies from the antiserum by selective elution from immunoblots of the components of the SGP-complex separated by SDS-PAGE. It was found that regardless of the protein band from which the antibody was eluted, every antibody isolated reacted with every protein component of the SGP-complex suggesting that the SGP-complex may have been derived from a single precursor protein and that the observed heterogeneity of the SGP-complex may have been the result of proteolytic breakdown of the protein held together by disulfide bonds. When the anti-SGP antiserum was used to immunoprecipitate 14C-proline labeled proteins from the media of bovine keratocytes in culture, the major protein observed had a Mr of about 140,000 daltons, similar to that of GP-140 also known as CL-glycoprotein. These proteins have been shown to represent the tissue form of Type VI collagen. To test the hypothesis that the SGP-complex may be related to the GP-140 (CL-glycoprotein), ELISA and immunoblotting studies were performed comparing the properties of the anti-SGP serum with those of a polyclonal antibody specific for Type VI collagen. The SGP-complex reacted positively by ELISA with the anti-human Type VI collagen antiserum and, conversely, human Type VI collagen gave a positive ELISA reaction with an antiserum against the SGP-complex. The anti-human Type VI collagen antiserum reacted with most of the major components of the SGP-complex on immunoblots of SDS-PAGE gels. These data indicate that the SGP-complex is related to and probably is derived from the tissue form of Type VI collagen.
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PMID:The bovine corneal SGP-complex is related to the tissue form of type VI collagen. 335 3

beta A4 amyloid peptide, the main constituent of amyloid plaques and cerebrovascular amyloid deposits associated with Alzheimer's disease, derives from a large precursor protein (APP) by the action of beta- and gamma-secretases, the unidentified endoproteases which release the amino and carboxyl termini of beta A4, respectively. Several gamma-secretase cleavage sites exist which yield the more soluble (1-39/40) forms of beta A4 and the longer forms (1-42/43) which have a greater tendency to aggregate into amyloid plaques. gamma-Secretase activity may therefore be critical in amyloid formation. In this study, a synthetic peptide which encompasses the various gamma-secretase cleavage sites was used as a substrate to probe proteases of various classes and specificities. Elastase, collagenase, and cathepsin D cleaved at the amyloidogenic sites (after Ala42 or after Thr43) to release the carboxyl termini of the aggregating forms. In addition, collagenase and pepsin released the carboxyl terminus of the more soluble forms. Human brain fractions enriched in lysosomes contained a proteolytic activity that cleaved the substrate at the amyloidogenic site(s). This activity was more active at acidic pH and was inhibited by pepstatin, two characteristics of the lysosomal aspartyl proteinase cathepsin D. The same lysosomal fractions were found to contain APP carboxyl-terminal fragments which are potentially amyloidogenic. These were degraded, only in acidic conditions, by an endogenous protease activity inhibited by pepstatin. Thus, a cathepsin D-like activity from human brain is a candidate for APP gamma-secretase(s).
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PMID:Candidate gamma-secretases in the generation of the carboxyl terminus of the Alzheimer's disease beta A4 amyloid: possible involvement of cathepsin D. 757 16

We recently demonstrated the presence of matrix metalloproteinases (MMPs)-1, -2, and -3 in AA amyloid deposits, which lead us to speculate that MMPs may participate in amyloidogenesis by either processing the precursor protein, or by degrading the amyloid deposits. Here we investigated this theory by determining the ability of MMP-1, -2, and -3 to degrade human acute-phase serum amyloid A (SAA) and human AA amyloid fibril proteins (AFPs). The following in vitro degradation experiments were performed: using either recombinant MMP-1, -2, or -3 and SAA as a substrate; using either recombinant MMP-1, -2, or -3 and AFP as a substrate; and using THP-1 cells as the protease source and AFP as the substrate. All three MMPs were able to cleave SAA and AFP within the region spanning residues 51 to 57. The following cleavage sites were identified: at 57 to 58 for MMP-1; at 7 to 8 and 51 to 52 for MMP-2; at 7 to 8, 16 to 17, 23 to 24, 51 to 52, 55 to 56, 56 to 57, and 57 to 58 for MMP-3. Cell culture experiments showed that THP-1 cells were able to degrade AFPs. Degradation was significantly delayed after addition of a general metalloproteinase inhibitor (o-phenanthroline) to dextran sulfate-stimulated cells. This is the first study to show that human SAAs and AFPs are susceptible to proteolytic cleavage by MMPs. Immunocytochemistry and electron microscopy showed that degradation takes place in the pericellular or extracellular compartment.
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PMID:Proteolysis of AA amyloid fibril proteins by matrix metalloproteinases-1, -2, and -3. 1148 14

Fibromodulin, a member of the small leucine-rich proteoglycan family, has been recently suggested as a biologically significant mediator of fetal scarless repair. To assess the role of fibromodulin in the tissue remodeling, we constructed an adenoviral vector expressing human fibromodulin cDNA. We evaluated the effect of adenovirus-mediated overexpression of fibromodulin in vitro on transforming growth factors and metalloproteinases in fibroblasts and in vivo on full-thickness incisional wounds in a rabbit model. In vitro, we found that Ad-Fibromodulin induced a decrease of expression of TGF-beta(1) and TGF-beta(2) precursor proteins, but an increase in expression of TGF-beta(3) precursor protein and TGF-beta type II receptor. In addition, fibromodulin overexpression resulted in decreased MMP-1 and MMP-3 protein secretion but increased MMP-2, TIMP-1, and TIMP-2 secretion, whereas MMP-9 and MMP-13 were not influenced by fibromodulin overexpression. In vivo evaluation by histopathology and tensile strength demonstrated that Ad-Fibromodulin administration could ameliorate wound healing in incisional wounds. In conclusion, although the mechanism of scar formation in adult wounds remains incompletely understood, we found that fibromodulin overexpression improves wound healing in vivo, suggesting that fibromodulin may be a key mediator in reduced scarring.
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PMID:Effect of adenoviral mediated overexpression of fibromodulin on human dermal fibroblasts and scar formation in full-thickness incisional wounds. 1721 96

A major collagenase was purified about 96-fold from a crude enzyme sample of Streptomyces parvulus by chromatography on Q-Sepharose, Sephacryl S-200, and butyl-Toyopearl. The purified enzyme showed a relative molecular mass of approximately 52,000 on SDS-PAGE and a pH optimum at about 9.0, and was strongly inhibited by metal-chelating agents. It also cleaved 4-phenylazobenzyloxycarbonyl-Pro-Leu-Gly-Pro-D-Arg specifically at the Leu-Gly bond, with a K(m) value of 0.60 mM at pH 9.0 at 37 degrees C. Based on the amino acid sequences of the N-terminal region and internal tryptic peptides, the corresponding gene was cloned. The DNA sequence of the cloned gene indicated that the enzyme is produced as an 864-residue precursor protein with a 408-residue prepro sequence followed by a 456-residue mature enzyme moiety. The enzyme is most homologous with the collagenase from S. coelicolor, the identity being 73%, and it is thought to be a member of the Vibrio collagenase subfamily.
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PMID:Purification and characterization of a major collagenase from Streptomyces parvulus. 1912 67

Photoaging of the skin is marked by obvious wrinkles and mainly depends on degradation of the extracellular matrix (ECM) in the dermis. Matrix metalloproteinase (MMP)-1 is one of the most important factors involved in degradation of the ECM; however, its mechanism of activation is not fully understood. It has been thought that MMP-1 is expressed by dermal fibroblasts as an inactive precursor protein that is activated by proteinases produced by keratinocytes in the epidermis. In this study, we constructed a 3D model of the dermis using collagen-embedded fibroblasts with or without ultraviolet (UV)-A exposure to mimic photoaging in the dermis. Collagen lattices embedded with UV-A-irradiated fibroblasts miniaturized and collagen was degraded to a greater extent than collagen lattices embedded with non-irradiated fibroblasts. The results demonstrate that fibroblasts in this 3D model express activated MMP-1 in the absence of keratinocytes. Moreover, the results confirm that activation of MMP-1 depends on increased plasmin activity in this model and lattice miniaturization was inhibited by the plasmin inhibitor tranexamic acid. Our results suggest that plasmin acts as an activator of MMP-1 and the inhibition of plasmin prevents collagen degradation.
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PMID:Dermal fibroblasts can activate matrix metalloproteinase-1 independent of keratinocytes via plasmin in a 3D collagen model. 2949 67