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)

Insoluble elastins were isolated from control and aneurytic aortas by a sequential extraction procedure involving the use of purified collagenase. Marked differences in amino acid analyses and susceptibilities to pancreatic elastase were observed between normal and pathological samples. The incorporation of either 14C-lysine or 14C-glucosamine into proteins of the vessel wall was also studied. In addition, high amounts of elastase-type activity was extractable from pathological aorta specimens which may contribute significantly to the loss of elastic tissue evidenced by ultra-structural studies and confirmed by the biochemical technics. We propose therefore that increased elastase-type protease activity in these pathological aortas does significantly contribute to the weakening of the aortic wall and also may well be the main cause of the rupture of aneurysms observed occasionally.
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PMID:Studies on elastic tissue of aorta in aortic dissections and Marfan syndrome. 703 99

The biosynthesis of collagen and fibronectin molecules by cultivated glomerular epithelial or mesangial cells was studied at confluency using radioactive proline or lysine as precursors. Collagen represented 0.5% of the total protein synthesized by the glomerular epithelial cells. About 60% of this collagenous protein were associated to the cell layer, whereas about 40% were secreted into the culture medium. Two major collagenous polypeptides were observed with apparent molecular weights of 185K and 170K, and were identified as two gene products of type IV procollagen. They exhibited ratios of 3- to 4-hydroxyproline, of total hydroxyproline to proline, and of hydroxylysine to lysine characteristic of type IV procollagen. They were degraded by bacterial collagenase. The patterns of peptides obtained after digestion of the 185K and 170K chains of this type IV procollagen with pepsin and V8 protease were identical to those obtained after digestion of type IV procollagen chains purified from a murine tumor (EHS sarcoma). Finally. a purified antibody to type IV collagen specifically immunoprecipitated the collagenous protein produced by the glomerular epithelial cells. By contrast, the mesangial cells synthesized about 5% of collagenous protein. 90% of this collagen were secreted into the cultured medium, whereas about 10% remained associated to the cell layer. Type I, III and IV procollagens were synthesized by the mesangial cells. Fibronectin was found in the medium and cell layer of both epithelial and mesangial cells. Fibronectin molecules were identified by their resistance to bacterial collagenase, their susceptibility to pepsin digestion, and their specific adherence to collagen. It was composed of disulfide-linked peptides of 220K daltons. The data therefore demonstrate that: (a) the glomerular epithelial and mesangial cells synthesize fibronectin molecules and type IV procollagen in vitro; (b) the cultivated mesangial cells also synthesize type I and III collagens. The implications of these findings in certain pathological circumstances, such as diabetes mellitus, are now being investigated.
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PMID:Synthesis of collagen and fibronectin by glomerular cells in culture. 732 12

Triacylglycerol metabolism has been studied in kidney cortex tubules from starved rats, prepared by collagenase treatment. Triacylglycerol was determined by a newly developed fully enzymic method. Incubation of tubules in the absence of fatty acids led to a decrease of endogenous triacylglycerol by about 50% in 1h. Addition of albuminbound oleate or palmitate resulted in a steady increase of tissue triacylglycerol over 2h. The rate of triacylglycerol synthesis was linearly dependent on oleate concentration up to 0.8mm, reaching a saturation at higher concentrations. Triacylglycerol formation from palmitate was less than that from oleate. This difference was qualitatively the same when net synthesis was compared with incorporation of labelled fatty acids. Quantitatively, however, the difference was less with the incorporation technique. Gluconeogenic substrates, which by themselves had no effect on triacylglycerol concentrations, stimulated neutral lipid formation from fatty acids. Glucose and lysine did not have such a stimulatory effect. Inhibition of gluconeogenesis from lactate by mercaptopicolinic acid likewise inhibited triacylglycerol formation. This inhibitory effect was seen with oleate as well as with oleate plus lactate. When [2-(14)C]lactate was used the incorporation of label into triacylglycerol was found in the glycerol moiety exclusively. Addition of dl-beta-hydroxybutyrate (5mm) to the incubation medium in the presence of oleate or oleate plus lactate led to a significant increase in triacylglycerol formation. In contrast with the gluconeogenic substrates, dl-beta-hydroxybutyrate had no stimulatory effect on fatty acid uptake. The results suggest that renal triacylglycerol formation is a quantitatively important metabolic process. The finding that gluconeogenic substrates, but not glucose, increase lipid formation, indicates that the glycerol moiety is formed by glyceroneogenesis in the proximal tubules. The effect of ketone bodies seems to be caused by the sparing action of these substrates on fatty acid oxidation. The decrease of triacylglycerol in the absence of exogenous substrates confirms previous conclusions that endogenous lipids provide fatty acids for renal energy metabolism.
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PMID:Triacylglycerol metabolism in isolated rat kidney cortex tubules. 737 17

The formation of adducts between methyltetrahydrophthalic anhydride (MTHPA), an important industrial chemical and potent allergen, and collagen from guinea pig lung tissue was investigated. Collagen peptides were obtained from the lung tissue by homogenization, defatting, washing, and digestion with collagenase. In experiments in vitro, lung tissue was exposed to 8.4 mumol (50 microCi) of 14C MTHPA. The amount of adducts was 97 nmol MTHPA/g of wet tissue as determined from the bound radioactivity. In a study in vivo, four guinea pigs were injected intratracheally with 8.4 mumol of 14C MTHPA each. The amount of adducts was 0-1.2 nmol MTHPA/g of wet tissue (determined by bound radioactivity). N epsilon-methyltetrahydrophthaloyl-L-lysine (MTHPL) was synthesized and characterized by NMR, UV, and mass spectrometry (MS). A method to analyze MTHPL, after derivatization with methanol and pentafluorobenzoyl chloride, using gas chromatography-MS was developed. Analysis of Pronase-digested MTHPA-exposed lung tissue showed a concentration of 19 nmol MTHPL/g wet lung in vitro and between 0 and 0.15 nmol MTHPL/g wet lung in vivo. Thus, 20% in vitro and 12-15% in vivo of the bound radioactivity was found as adducts with lysine. These results are a first step toward studies of allergenic epitopes in proteins and methods for biological monitoring of exposure to acid anhydrides.
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PMID:Lysine adducts between methyltetrahydrophthalic anhydride and collagen in guinea pig lung. 748 35

The ability of TSH to stimulate synthesis and release of thyroid hormones in ovine thyroid follicles in vitro depends partially on a synergy with insulin-like growth factors (IGFs). The cellular availability of IGFs may be influenced by the release of several IGF binding proteins (IGFBPs). The purposes of these studies was to 1) further characterize the species of IGFBPs synthesized by thyroid follicles, 2) examine the ability of TSH and cortisol to alter IGFBP gene expression and protein release, and 3) investigate the actions of exogenous IGFBPs on thyroid cell function. Adult sheep thyroid follicles were isolated after collagenase digestion, grown to confluence in Coon's modified Ham's F12M medium (OH) with the addition of transferrin, glycylhistidyl-lysine, somatostatin (3H), cortisol and insulin, and maintained in serum-free test media with or without further supplements for up to 48 h. Conditioned media were analyzed for IGFBP presence by Western ligand blotting, and by immunoblotting using specific antisera against bovine IGFBP-2 and human IGFBP-5. IGFBP mRNAs from follicles were identified by Northern blot hybridization using [32P]labeled complementary DNAs encoding ovine IGFBP-1 or -2, and rat IGFBP-4, -5, or -6. Uptake and organification of Na[125I] were measured by incorporation into trichloroacetic acid-precipitable material. Isolated thyroid follicles synthesized four species of IGFBPs in either 0H or 3H medium as detected by ligand blotting, of sizes 40-46, 34, 28, and 18 kilodaltons (kDa), respectively. The 32 kDa IGFBP was identified immunologically as IGFBP-2, whereas the 28 kDa and 18 kDa species were identified as IGFBP-5. Northern blot hybridization of total RNA from cells in 3H medium demonstrated an IGFBP-2 messenger RNA (mRNA) [1.4 kilobase (kb)], an IGFBP-4 mRNA (2.6 kb), and two IGFBP-5 mRNAs (6 kb and 1.8 kb). No IGFBP-1 or -6 mRNAs were detected. Incubation of cultured follicles with TSH (30-500 microU/ml) caused a dose-dependent decrease in the abundance of all IGFBP mRNAs and released proteins, which were reduced further by TSH together with cortisol (10 nM). When the inhibitory effect of TSH and cortisol was removed, IGFBP-2 mRNA increased within 3 h and was 7-fold greater within 12 h. IGFBP-2 did not appear in the conditioned medium until 12 h after TSH removal, along with the other IGFBP species.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Hormonal regulation and biological actions of insulin-like growth factor binding proteins in isolated ovine thyroid follicles. 750 36

Enzymes and chemicals were used to analyse the biochemical structure of the antigenic epitope recognized by GDA-J/F3 monoclonal antibody (MoAb) in the human sperm tail fibrous sheath. Treatment of sperm dried onto slides with trypsin or dispase enzymes abolished their immunofluorescence staining with GDA-J/F3 MoAb, thus indicating the proteinaceous nature of the antigen. The proteolytic cleavage of GDA-J/F3 protein by trypsin, which also caused sperm decapitation, indicated the presence of peptide bonds involving the carboxyl groups of the basic amino acids, arginine and/or lysine. The epitope was also glycosylated as demonstrated by its sensitivity to sodium metaperiodate treatment which was dose-dependent. The GDA-J/F3 antigenic epitope lacked sialic acid since pre-treatment of spermatozoa with sialidase enzyme (neuraminidase) had no effect on their reactivity with the antibody. The lack of collagenous domains in the GDA-J/F3 antigen was demonstrated by the failure of collagenase to abrogate sperm immunostaining with the MoAb. Furthermore, type VII collagen of the skin basement membrane (BM) was previously thought of as a potential target antigen for GDA-J/F3 MoAb. This was ruled out since several monoclonal and polyclonal antibodies failed to detect the antigen in the spermatozoa using immunofluorescence and Western blotting. These data, therefore, show that the target antigen for GDA-J/F3 MoAb is a non-collagenous asialo-glycoprotein, and by inference provide the first evidence for the glycosylation of the sheath proteins as another step of post-translational modification occurring during sperm tail development.
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PMID:The target antigen for GDA-J/F3 monoclonal antibody in the human sperm tail fibrous sheath is a non-collagenous asialo-glycoprotein: implications and significance. 752 22

1. The kinetics of the degradation of the kinins bradykinin and Met-Lys-bradykinin, angiotensins I and II and the tachykinin substance P by PMNL-collagenase (MMP 8), PMNL-gelatinase (MMP 9) and by the recombinant catalytic domain of MMP 8 (rcd-PMNL-c) was examined by RP-HPLC. The resulting fragments were identified by automated Edman degradation or by amino acid analysis. 2. The initial degradation rates of substance P at a substrate concentration of 25 microM were 5 min-1 for MMP 9 and 150 min-1 for MMP 8. The kinetic constants KM and kcat were determined by concentration-dependent measurements. For MMP 8/substance P the constants were KM = 78 +/- 14 microM and kcat = 412 +/- 67 min-1. For MMP 9/substance P the constants were KM = 91 +/- 15 microM and kcat = 25 +/- 4 min-1. Both enzymes cleaved substance P between Gln6 and Phe7 and between Gly9 and Leu10. 3. Under the same conditions, MMP 8 degraded angiotensin I at an initial rate of 20 h-1, resulting mainly in the vasoactive fragments angiotensin II and angiotensin(1-7). At a substrate concentration of 25 microM and an enzyme/substrate ratio of 1:100, angiotensin II was degraded very slowly (19% in 24 h) by MMP 8. Under these conditions, MMP 9 degraded angiotensin I to a lesser extent than MMP 8 (25% in 24 h) and was unable to cleave angiotensin II. 4. Under the same conditions, bradykinin and Met-Lys-bradykinin were cleaved by PMNL-collagenase at a rate of 20% in 24 h, producing BK(1-7) and BK(1-8). PMNL-gelatinase was unable to cleave the kinins under these conditions. 5. In all cases, rcd-PMNL-c produced the same fragments as wild type PMNL-collagenase, but at a significantly lower rate.
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PMID:Degradation of kinins, angiotensins and substance P by polymorphonuclear matrix metalloproteinases MMP 8 and MMP 9. 753 73

In addition to the known 94-kd gelatinase (matrix metalloproteinase 9, MMP-9), HL-60 leukemia cells release a hither-to undescribed 45-kd metalloproteinase into the culture medium. This enzyme cleaves the synthetic substrate Pro-Gln-Gly-Ile-Ala-Gly-Gln-Arg, which represents the cleavage site for collagenases in collagen type I not between isoleucine and alanine--the typical cleavage site for collagenases--but between alanine and glycine. The enzymatic activity was purified through a combination of zinc-chelate-Sepharose column chromatography, precipitation with Fractogel TSK-AF Red and gelatin-Sepharose, and subsequent sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Microsequence analysis of the NH2-terminus of the purified 45-kd proteinase revealed the sequence Asp-Ile-Ser-Lys-Tyr-Thr-Thr-Thr-, which could not be found in other proteins when searched in several protein data bases. Incubation of the enzyme immobilized on nitrocellulose membranes with polyclonal antibodies to collagenase and stromelysin or gelatinases revealed no cross-reactivity. The proteolytic activity was not increased by treatment with trypsin, 8M urea, acid, or organomercurials. The proteinase, which was inhibited by chemical inhibitors of metalloproteinases, such as phenanthrolene or EDTA, is able to degrade several matrix constituents, such as collagen type IV, fibronectin, gelatin, and proteoglycans. In contrast to all known MMPs, the proteolytic activity of the 45-kd enzyme was not abolished upon incubation with recombinant tissue inhibitors of matrix metalloproteinases (TIMP) 1 or 2. Thus, the novel enzyme may influence extracellular matrix (ECM) turnover in vivo because its activity is not influenced by specific inhibitors of MMPs.
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PMID:Leukemic cells (HL-60) produce a novel extracellular matrix-degrading proteinase that is not inhibited by tissue inhibitors of matrix metalloproteinases (TIMPs). 782 72

Matrix metalloproteinase 7 (MMP-7) has been purified as an inactive zymogen of M(r) 28,000 (proMMP-7) from the culture medium of CaR-1 human rectal carcinoma cells. The NH2-terminal sequence of proMMP-7 is Lys-Pro-Lys-Pro-Gln-Glu, which is identical to that of matrilysin. The zymogen is activated by 4-aminophenylmercuric acetate (APMA), yielding an intermediate form of M(r) 21,000 and an active species of M(r) 19,000 which shows the new NH2-terminal sequence of Tyr78-Ser-Leu-Phe-Pro-Asn-Ser. Although trypsin fully activates the zymogen, the activation rate by plasmin or leukocyte elastase is confined to approximately 50%. ProMMP-7 can be activated by MMP-3 (stromelysin 1) to its full activity in a single-step mechanism and generates the same NH2 terminus obtained by APMA activation, whereas MMP-1 (tissue collagenase), MMP-2 (gelatinase A), and MMP-9 (gelatinase B) do not have such an effect. On the other hand, proMMP-1 is activated by MMP-7 to an activity similar to that obtained by APMA and the activation by MMP-7 is enhanced up to approximately 6.5 fold in the presence of APMA. This enhanced activity is donated by specific cleavage at the Gln80-Phe81 bond of proMMP-1. MMP-7 can also activate proMMP-9 up to approximately 50% of the full activity with a new NH2 terminus of Leu16-Arg-Thr-(Asn)-Leu. Incubation of proMMP-2 or proMMP-3 with MMP-7 results in no activation of these proMMPs. MMP-7 degrades type IV collagen, laminin-1, fibronectin, proteoglycan, type I gelatin, and insoluble elastin. These results suggest that in vivo MMP-7 may play a role in degradation of extracellular matrix macromolecules in concert with MMP-1, -3, and -9 under pathological conditions.
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PMID:Matrix metalloproteinase 7 (matrilysin) from human rectal carcinoma cells. Activation of the precursor, interaction with other matrix metalloproteinases and enzymic properties. 789 11

Affinity-based purification and characterization of the collagenolytic serine protease 1 from Uca pugilator (fiddler crab) hepatopancreas shows that the enzyme cleaves the native bovine alpha 1(I) collagen chain carboxyl-terminal to Gln and Arg residues adjacent to the metallocollagenase site. Cleavage carboxyl-terminal to Leu residues is observed in the alpha 2(I) chain and at a secondary site in alpha 1(I). These sites correlate with the preferences observed toward p-nitroanilide substrates varying at the P1 position, for which the specificity (kcat/Km) is Arg > Leu, Phe, Lys > Gln > Ala. Furthermore, collagen cleavage after Gln was found exclusively between two Gln-Arg bonds. The P'1-P'3 specificity of collagenase, as determined by nucleophile acyl transfer, indicated a strong preference for Arg in the P'1 position. Crab collagenase cleaves peptide bonds adjacent to Leu and Gln at the P1 position more efficiently than trypsin, chymotrypsin, or elastase. Moreover, the efficiency of collagenase toward P1-Arg substrates is equivalent to that of trypsin. Crystals of crab collagenase have been grown complexed with the protein inhibitor ecotin. These crystals diffract to better than 2.8 A resolution and belong to the space group P3(2)21 with unit cell dimensions of a = b = 89.0 A, c = 291.7 A.
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PMID:The substrate specificity of Uca pugilator collagenolytic serine protease 1 correlates with the bovine type I collagen cleavage sites. 803 25


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