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)

PZ-peptidase is an endopeptidase that cleaves the synthetic substrate developed for clostridial collagenase, 4-phenylazobenzyloxycarbonyl-L-Pro-L-Leu-Gly-L-Pro-D-Arg (PZ-peptide). The peptidase has been purified to homogeneity from chicken embryos. The enzyme has a pH optimum of 7.5 to 8.5, and isoelectric point of 5.0, and a molecular weight of 77,000. The kinetic parameters at pH 8 and 37 degrees are: Km = 2 X 10(-4) M and Vmax = 4.2 mumol/min/mg of protein. The enzyme is inhibited by p-hydroxymercuribenzoate (100%), N-ethylmaleimide (60%), and chelating agents (40 to 60%). Maximum activity is attained in the presence of reducing agents and Ca2+, Sr2+, or Mg2+. The peptidase has no detectable action on casein, serum albumin, collagen, collagen alpha chains, various collagen peptides (alpha1)(I)-CB2, alpha1(I)-CB3, alpha1(I)-CB4), (Gly-Pro-Pro)10, or (Gly-Pro-Pro)5. It does catalyze the hydrolysis of the Hyp--Gly bond in the 17-residue collagen peptide alpha1(II)-CB6-C2 and it partially digested a mixture of collagen peptides of molecular weight 350 to 2500. A role of this peptidase in collagen breakdown appears to be restricted to a late stage when degradation products would fall in the range of 5 to 30 residues.
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PMID:PZ-peptidase from chick embryos. Purification, properties, and action on collagen peptides. 1 6

1. The synthetic peptide, 2,4-dinitrophenyl-L-Pro-L-Leu-Gly-L-Ile-L-Ala-Gly-L-Arg-amide (DNP-peptide) was tested as a potential substrate for uterine collagenase. Rat uteri were homogenized and the insoluble fraction was extracted at 60 degrees C to obtain collagenase. The extracts were chromatographed on Sephadex G-150 to yield two peaks of DNP-peptide hydrolyzing activity. Peak I was completely inhibited by EDTA and had a molecular weight greater than 100 000. Peak II was inhibited about 90% by EDTA and had an apparent molecular weight of about 70 000. 2. Peak II coincided closely, but not exactly, with the peak of collagenase activity. It differed from collagenase in heat stability, binding properties on CM-Sephadex and failure to display latency. 3. Peak II represents a new endopeptidase activity. It has a pH optimum of 7 and it cleaves the DNP-peptide at the Gly-Ile and, possibly, the Leu-Gly bond. 4. The DNP-peptide is not a satisfactory substrate for the assay of impure collagenase preparations nor does it inhibit the action of collagenase on collagen substrate when added in 30-fold molar excess.
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PMID:Separation of collagenase and a metal-dependent endopeptidase of rat uterus that hydrolyzes a heptapeptide related to collagen. 22 33

An endopeptidase was purified to homogeneity from the cell extracts of Treponema denticola ATCC 35405 (a human oral spirochete) by a procedure that comprised dialysis, anion exchange fast protein liquid chromatography (FPLC), hydroxylapatite FPLC, immobilized metal affinity FPLC, FPLC chromatofocusing, and two consecutive gel permeation FPLC steps. The enzyme is a 62-kDa protein with an isoelectric point of 6.5-7.0. Experiments with enzyme inhibitors suggest that this enzyme is a metallopeptidase and that its activity is not dependent on sulfhydryl or serine residues. The enzyme is active on furylacryloyl-Leu-Gly-Pro-Ala (FALGPA; pH optimum near 6.25), bradykinin (Bk), and several Bk-related peptides. In FALGPA, the cleavage site is the Leu-Gly bond. An imino acid is absolutely necessary in position P'2. The shortest hydrolyzed peptide was FALGPA, the hydrolysis of which is strongly and competitively inhibited by Bk (Ki = 5.0 microM). The pyrophosphate ion and phosphoramidon also inhibited the hydrolysis of FALGPA. The enzyme does not hydrolyze all typical synthetic collagenase substrates, Azocoll, Azocasein, or Type I and Type IV collagens, or any other proteins tested. In Bk-related peptides, the hydrolyzed bond was Phe5-Ser6. Since a Bk antagonist and a Bk-potentiating pentapeptide also were good substrates, it is possible that the enzyme hydrolyzes Bks and related peptides only because of the coincidental, specific amino acid sequence of those substrates. A proposal is made that since a substantial portion of the amino acid sequence of FALGPA is present in collagen (and additionally acknowledging that the furylacryloyl residue structurally resembles that of proline), the natural substrates of this enzyme may be small, soluble collagen fragments produced by other enzymes from periodontal connective tissue, and that such peptides are important for the nutrition and pathogenicity of T. denticola.
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PMID:Purification and substrate specificity of an endopeptidase from the human oral spirochete Treponema denticola ATCC 35405, active on furylacryloyl-Leu-Gly-Pro-Ala and bradykinin. 132 Nov 41

Inflammation of the periodontium leads to connective tissue degradation and eventual tooth loss. The regulation of matrix metalloproteinases (MMPs) has been studied to determine their role in these processes and also during tissue remodelling. Analysis of gingival crevicular fluid has revealed the presence of collagenase and gelatinase that, in the acute stages of periodontal disease, are derived predominantly from polymorphonuclear leukocytes. These MMPs appear to be intimately associated with tissue destruction since the levels of the active forms of these enzymes obtained from either crevicular fluid or mouthrinse samples correlate with tissue destruction and, therefore, provide a sensitive means of demonstrating disease activity. Transforming growth factor-beta, an important regulator of connective tissue remodelling, has been implicated in the rapid remodelling of periodontal tissues. TGF-beta promotes tissue matrix formation by stimulating both the synthesis of matrix proteins (collagen, fibronectin and SPARC) and proteinase inhibitors (TIMP, PAI-1) and by decreasing the synthesis of MMPs, but not the 72 kDa-gelatinase. Nuclear run-on analyses have shown that TGF-beta reduces collagenase and stromelysin synthesis by suppressing gene transcription without altering mRNA stabilities. In contrast, the transcription of the gelatinase and TIMP genes was increased by TGF-beta, which also increased gelatinase mRNA stability. Remodelling of alveolar bone involves interaction between osteoblasts and osteoclasts. Osteoblasts, under the influence of osteotropic hormones (vit D3, PTH and retinoic acid), produce MMPs which appear to function in the removal of soft tissue that precludes access of osteoclasts to the mineralized tissue surface. Rat osteoblastic cells produce MMPs with activity on native collagen, native collagen 3/4-fragments and gelatin and, in addition, two forms of TIMP activity. The 3/4-collagen endopeptidase, purified to apparent homogeneity, also has significant collagenase and gelatinase activities and an amino terminal sequence almost identical to human 72 kDa-gelatinase. The production of this enzyme was stimulated by TGF-beta, which suppresses bone resorption, and by osteotropic hormones which stimulate bone resorption, supporting a bifunctional role for the gelatinase in connective tissue remodelling. Although there is strong evidence for the involvement of MMPs in the resorption of bone and in the inflammation-mediated destruction of periodontal tissues, the role of MMPs in the remodelling of mature soft connective tissues remains equivocal.
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PMID:Matrix metalloproteinases in periodontal tissue remodelling. 148 60

A metal-dependent peptidase was isolated from the homogenate of human uterus by standard chromatographic techniques and purified to apparent homogeneity. The peptidase hydrolysed the synthetic vertebrate collagenase substrate 2,4-dinitrophenyl-Pro-Gln-Gly-Ile-Ala-Gly-Gln-D-Arg (Dnp-peptide), the synthetic bacterial collagenase substrate 4-phenylazobenzyloxycarbonyl-Pro-Leu-Gly-Pro-D-Arg (PZ-peptide) and gelatinolytic peptides of gelatin, but was inactive against collagen type I, gelatin and casein. The cleavage site for the Dnp-peptide was the Gly-Ile bond. The enzyme was not only inhibited by metal chelators, such as EDTA, 1,10-phenantroline and dithiothreitol but also by thiol reagents, such as mersalylic acid and N-ethylmaleimid. However, E-64, an inhibitor for thiolproteinases, and leupeptin, an inhibitor for thiol- and serine proteases, did not exhibit any inhibitory activity. Pepstatin, an inhibitor for aspartate proteinases, and inhibitors for serine proteinases like phenylmethanesulfonyl fluoride and Trasylol were ineffective as well. The purified peptidase displayed a single band in the SDS-PAGE with an apparent molecular mass of 65 kDa. Employing isoelectric focusing an IP of 5.0 could be determined. The enzyme's properties are discussed in relation to the proteinase EC 3.4.24.11 and to proteinases of the collagenase family as well as the possibility to discriminate these three metalloproteinase classes by employing the Dnp-peptide.
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PMID:Isolation and properties of a metal-dependent endopeptidase from human uterus hydrolysing synthetic collagenase substrates. 165 Feb 34

The nuchal ligament of unborn calves contains a neutral endopeptidase that is biochemically and immunologically similar to the neutral endopeptidase (NEP), or enkephalinase, from human kidney. Enzymatic activity was inhibited more than 90% by phosphoramidon (1 microM). The specific activity in membrane fractions, as determined by hydrolysis of the dansylated substrate, DAPGN, was similar in tissue from fetuses of gestational ages ranging from 100 to 280 days. NEP activity in adult ligament tissue, however, was less than 10% of that in fetal tissue. Fibroblasts dissociated from ligament tissue by collagenase displayed less NEP activity than did preparations of intact ligament, and activity was even lower in cultured cells. By contrast, fibroblasts cultured from fetal calf lungs had NEP activity comparable to that in the ligament tissue. When ligament fibroblasts were cultured on subcellular matrices derived from fetal lung fibroblasts the NEP activity increased relative to those cultured on plastic alone. These studies confirm the presence of neutral endopeptidase (NEP) in the nuchal ligament of the fetal calf. The consistent activity through a range of gestational ages and the influence of the subcellular matrix suggest that this enzyme might be involved in growth of the ligament during fetal life.
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PMID:Neutral endopeptidase from nuchal ligament of fetal calves. 219 69

The neutral zinc metalloendopeptidase (NEP, EC 3.4.24.11) is an integral membrane protein found in brain tissue, polymorphonuclear leukocytes, and many epithelia. We show here that NEP is expressed on rabbit synovial fibroblasts and on simian virus 40 (SV40) DNA- and H-ras-transformed rabbit mammary epithelial cells. Treatment of these cells with 12-O-tetradecanoylphorbol-13-acetate (TPA) for 24 h decreased expression of NEP mRNA transcripts and decreased the biosynthetically labeled immunoprecipitable NEP antigen. In contrast to its effects on NEP, TPA treatment induced expression of the secreted metalloproteinase collagenase and the tissue inhibitor of metalloproteinases. TPA induced stromelysin, another secreted metalloproteinase, only in the fibroblasts. These data provide evidence that the expression of the membrane-bound NEP is regulated in several cell types.
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PMID:Phorbol diesters regulate expression of the membrane neutral metalloendopeptidase (EC 3.4.24.11) in rabbit synovial fibroblasts and mammary epithelial cells. 254 98

Parenchymal organoidal structures that were obtained from collagenase digestion of reduction mammoplasty specimens of apparently normal human breasts have been grown in short-term primary cultures, either on plastic or on floating gels of polymerized rat-tail collagen. Three morphologically distinct major cell types are readily observed in both systems: cuboidal cells, which occupy apical positions on collagen gels; larger, epithelioid, or basal cells on gels; and elongated cells which penetrate into the gel. In addition, a fourth cell type, that of large, flat cell, is observed less readily by phase contrast microscopy on the surface of cultures grown on plastic. Immunofluorescent and immunocytochemical staining of cultures on plastic or histologic sections of cultures on gels have been undertaken with antisera and other histochemical reagents that stain the different parenchymal cell types in vivo. Thus antisera to epithelial membrane antigen(s), monoclonal antibodies (MABs) to the defatted mammary milk fat globule membrane, peanut lectin, and keratin MAB LE61, which preferentially stain the epithelial cells of ducts in vivo, also stain the cuboidal/apical cells in vitro. The large, flat cells are stained intensely by the first three reagents but not by the last one. Antisera to collagen IV, laminin, fibronectin, actin, keratin MAB LP34, MABs to the common acute lymphoblastic leukemia antigen, and MAB LICR-LON-23.10, which showed enhanced staining for the ductal myoepithelial cells in vivo, also stain the epithelioid/elongated cells in vitro. However, the effect of the last four reagents is reduced considerably in most elongated cells, and MAB LP34 stains the large, flat cells intensely. Heterogeneous cells of intermediate morphologies and staining patterns between the cuboidal cells and large, flat cells are related to mammary epithelial cells. whereas the large epithelioid/elongated cells have some characteristics of myoepithelial cells, and that intermediate forms may exist in culture between the two parenchymal cell types.
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PMID:Characterization of human mammary cell types in primary culture: immunofluorescent and immunocytochemical indicators of cellular heterogeneity. 264 83

A. niger LCF 9 synthesizes a new aspergillopeptidase of potential interest in therapeutics. The properties and operating range of the enzyme were determined. It is a semi-alkaline aspergillopeptidase (EC 3.4.23.4) with one endopeptidase activity. Its pI is 4.10, its molecular weight is 21000 Da and its A1%(1 cm) at 280 nm is 9.75. It rapidly hydrolyzes casein and hemoglobin. Its optimal pH is 7.8 and optimal temperature is 45 degrees C. It is thermally labile above 40 degrees C but can be stabilized by adding calcium ions. It is inhibited by phenylmethylsulfonyl fluoride (PMSF), by ethylenediaminetetraacetic acid (EDTA) and by certain metals ions, e.g. copper, manganese and cobalt ions. It has no dipeptidase or tripeptidase activity and its esterase activity is weak. It has a high collagenase activity and is to our knowledge the only aspergillopeptidase that is active towards benzoyl-arginine p-nitroanilide (BAPNA).
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PMID:Properties of a new alkaline proteinase from Aspergillus niger. 269 84

The amino-terminal amino acid sequence and several internal peptide sequences of angiotensin I-converting enzyme (ACE; peptidyl-dipeptidase A, kininase II; EC 3.4.15.1) purified from human kidney were used to design oligonucleotide probes. The nucleotide sequence of ACE mRNA was determined by molecular cloning of the DNA complementary to the human vascular endothelial cell ACE mRNA. The complete amino acid sequence deduced from the cDNA contains 1306 residues, beginning with a signal peptide of 29 amino acids. A highly hydrophobic sequence located near the carboxyl-terminal extremity of the molecule most likely constitutes the anchor to the plasma membrane. The sequence of ACE reveals a high degree of internal homology between two large domains, suggesting that the molecule resulted from a gene duplication. Each of these two domains contains short amino acid sequences identical to those located around critical residues of the active site of other metallopeptidases (thermolysin, neutral endopeptidase, and collagenase) and therefore bears a putative active site. Since earlier experiments suggested that a single Zn atom was bound per molecule of ACE, only one of the two domains should be catalytically active. The results of genomic DNA analysis with the cDNA probe are consistent with the presence of a single gene for ACE in the haploid human genome. Whereas the ACE gene is transcribed as a 4.3-kilobase mRNA in vascular endothelial cells, a 3.0-kilobase transcript was detected in the testis, where a shorter form of ACE is synthesized.
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PMID:Two putative active centers in human angiotensin I-converting enzyme revealed by molecular cloning. 284


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