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)

Surfactant-associated protein of Mr = 35,000, SAP-35, is the major glycoprotein present in mammalian pulmonary surfactants. In this study, canine SAP-35 and several of its COOH-terminal peptides were purified and characterized by amino acid composition and NH2-terminal sequencing analysis. These proteins were then studied in terms of their specific lipid-binding characteristics and surface activity when combined with a synthetic phospholipid mixture, SM, chosen as an approximation of lung surfactant phospholipids. Purified, delipidated SAP-35 bound SM strongly. In contrast, SAP-21 (a non-collagenous fragment generated by collagenase digestion) bound phospholipid weakly; SAP-18 (an acidic COOH-terminal fragment comprising residues Gly-118 to Phe-231) did not bind phospholipid, demonstrating the importance of hydrophobic amino acid residues Gly-81 to Val-117 and the NH2-terminal collagenous domain in interaction of the SAP-35 with phospholipids. In surface activity experiments, purified SAP-35 enhanced the adsorption of SM phospholipids in terms of both rate and overall surface tension lowering. However, the adsorption facility of the SM-SAP-35 mixture did not approach that of either whole surfactant or the surfactant extract preparations, calf lung surfactant extract or surfactant-TA, used in exogenous surfactant replacement therapy for the neonatal respiratory distress syndrome. In addition, the dynamic surface activity of the SM-SAP-35 mixture was well below that of natural surfactant or surfactant extracts. This was also true of mixtures of SM phospholipids combined with the SAP-18 and SAP-21 fragments of SAP-35.
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PMID:Phospholipid binding and biophysical activity of pulmonary surfactant-associated protein (SAP)-35 and its non-collagenous COOH-terminal domains. 302 34

The zona pellucida (ZP) from pig oocytes was isolated using two different methods. In the first method, the ZP was isolated using sieving procedures. In the second method, an enzymatic step with collagenase was used in addition to sieving procedures. Several commercial collagenase preparations were tested. The macromolecular composition of the ZP isolated by these two methods was determined by two-dimensional polyacrylamide-gel electrophoresis after disulfide bond reduction. The ZP prepared by the sieving method contained four glycoprotein families with apparent molecular weights of 25,000, 55,000, 65,000, and 90,000. The ZP obtained using the enzymatic method was distinctly different, lacking the highest molecular-weight family (90,000) and containing at least three new constituents with apparent molecular weights of 70,000, 40,000, and less than 25,000. Commercial collagenase preparations, when analyzed by two-dimensional polyacrylamide-gel electrophoresis to assess homogeneity, contained numerous protein components. The trypsin-like protease concentration in the collagenase preparations was determined to be 3.4-42 X 10(-8) M as determined by activity measurement using benzoyl-DL-arginine-beta-naphthylamide as substrate or the active site titrant p-nitrophenyl-p'-guanidinobenzoate. Thus, the ZP prepared by the enzymatic method, using collagenase preparations, had an altered macromolecular composition, thereby rendering the ZP unsuitable for most structural, immunological, or sperm-binding studies.
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PMID:Proteolysis by collagenase preparations alters the macromolecular composition of the porcine zona pellucida. 302 48

A collagenase inhibitor was purified from bovine cartilage by a combination of gel filtration, ion exchange, concanavalin A-Sepharose affinity chromatography, and elution from preparative sodium dodecyl sulfate-polyacrylamide gels. The inhibitor was purified 370-fold and migrated as a single polypeptide with an Mr of 19,000 on SDS-polyacrylamide gels. It stained positively for carbohydrate with periodic acid-Schiff's reagent and bound to lectins, indicating that it is a glycoprotein. The inhibitory activity was stable to heating up to 60 degrees C and between pH 4 and 10. The inhibition of collagenase by the cartilage inhibitor could not be reversed by trypsin or mersalyl. The inhibitory activity did not require the presence of free sulfhydryl groups, and it could be removed from the cartilage extract by incubating with native collagen, suggesting that the inhibitor binds to collagen. The cartilage inhibitor was effective against human and mouse interstitial collagenases, but it did not inhibit trypsin or bacterial collagenase.
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PMID:A vertebrate interstitial collagenase inhibitor from bovine scapular cartilage: purification and characterization. 303 Apr 45

There are no known intestinal cell phenotypic markers characteristic of the transmural idiopathic inflammatory disease of the small bowel. To address this issue, we developed monoclonal antibodies specific for Crohn's disease tissue by generating a library of hybridomas specific for intestine following murine immunization with intestinal epithelial cells from a patient with Crohn's disease. The epithelial cells were initially harvested from a fresh operative specimen by gentle scraping and digestion with 1% collagenase on ice. Cells were then washed, evaluated for viability, and polytron homogenized. After immunization and subsequent fusion, hybridoma cell lines producing distinct antibody-binding patterns were identified by indirect immunoepifluorescence (IIEF). Wells representing distinct patterns were subcloned and carried to limiting dilution. Of the multiple hybridomas studied, the predominant target antibodies were goblet cell and glycoprotein-like molecules. Patterns of antibody binding identified by IIEF included: brush border, goblet cell, surface glycoprotein, enterocyte membranes, basilar crypt inclusions, circumferential goblet cell, and a heterogeneous goblet cell glycoprotein pattern. Molecular weight determination and cross-reactivity with various human tissues were studied by immunoblotting following sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. Several hybridomas were specific for the intestine but were not specific for Crohn's disease.
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PMID:Generation of monoclonal antibodies to involved ileum of Crohn's disease: characterization of a panel of antibodies with goblet cell membrane and brush border-specific reactivity. 304 35

The glycosaminoglycans (GAG), glycoproteins and collagen in bovine aorta and venous tissue have been studied. The concentration of hyaluronic acid and dermatan sulphate was significantly more in the venous tissue while chondroitin sulphates were higher in the aorta. Sequential extraction with phosphate buffered saline (PBS) collagenase, hyaluronidase and urea was also carried out with the two tissues. The GAG extractable by PBS and collagenase digestion were more in the aorta. The total aortic glycoproteins had significantly lower hexose and higher sialic acid. The PBS extractable glycoproteins of the venous tissue had more hexose and fucose. The glycoproteins released by collagenase digestion of the venous tissue had lower sialic acid and higher fucose, while glycoprotein released by hyaluronidase digestion had lower sialic acid and higher hexose and fucose. Urea extractable glycoproteins had lower fucose and sialic acid in the venous tissue. Venous tissue had higher total collagen and acid and salt soluble collagen while insoluble collagen was more in the aorta. The total GAG in the venous tissue had greater anticoagulant activity while the aortic GAG bound significantly more serum lipoproteins.
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PMID:Studies on the macromolecular components of the bovine aortic and venous tissue. 309 9

Alveolar proteinosis is a disease characterized by accumulation of proteinaceous material in the alveolar space of the lung. Two major collagenase-sensitive polypeptides, alveolar proteinosis peptides of 34 kDa kilodaltons (APP-34) and of 62 kDa (APP-62), were isolated from bronchioalveolar lavage of patients with alveolar proteinosis. These proteins co-purified during fast-performance liquid chromatography (FPLC) chromatofocusing and were separated from each other by electroelution following SDS-polyacrylamide gel electrophoresis. Immunoblot analysis of these proteins demonstrated that both shared antigenic sites with the normal human surfactant-associated protein of Mr 34,000 (SAP-34) using both polyclonal and monoclonal antibodies generated against SAP-34. Removal of asparagine-linked oligosaccharides from the 34 kDa and 62 kDa alveolar proteinosis proteins with endoglycosidase F resulted in polypeptides of 28 kDa from APP-34 and 56 kDa from APP-62. Amino acid analysis and tryptic peptide maps of the electroeluted APP-34 and APP-62 proteins were essentially identical and similar to that previously reported for human SAP-34, supporting the likely relationship of APP-34 and APP-62 as monomer and dimer of the normal SAP-34. APP-34 and APP-62 were both sensitive to bacterial collagenase, yielding collagenase-resistant fragments of 21 kDa, similar in migration and amino acid composition to the fragment generated by collagenase digestion of normal human SAP-34. High molecular weight aggregates of APP-34 and APP-62 were the result of sulfhydryl-dependent and non-sulfhydryl-dependent cross-linking. A domain in the C-terminal non-collagenous portion of the molecules which forms sulfhydryl-dependent oligomers was identified. The two major polypeptides accumulating in the airway of patients with alveolar proteinosis are monomeric (34 kDa) and dimeric (62 kDa) forms of the major surfactant-associated glycoprotein, SAP-34.
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PMID:Structural relationships of the major glycoproteins from human alveolar proteinosis surfactant. 310 39

Hepatic lipase, a glycoprotein synthesized and secreted by the hepatocyte, binds to sinusoidal endothelium where it is involved in metabolism of lipoprotein phospholipid and triglyceride. To better understand the regulation of hepatic lipase, we investigated the synthesis, post-translational processing, and secretion of the enzyme by isolated rat hepatocytes. Metabolically labeled [35S]methionine hepatic lipase protein, produced by the collagenase-dispersed hepatocytes, was immunoisolated from detergent-solubilized cells and incubation medium at designated times, using a polyclonal rabbit anti-rat hepatic lipase antibody raised against hepatic lipase purified to homogeneity from rat liver post-heparin perfusates. Following polyacrylamide gel electrophoresis and fluorography, radiolabeled hepatic lipase was quantitated by densitometry. Newly synthesized hepatic lipase was rapidly secreted and accumulated in the medium as a 59,000-dalton protein in a manner consistent with a constitutive process. An intracellular 53,000-dalton precursor of the mature 59,000-dalton hepatic lipase was identified by immunoprecipitation. The 53,000-dalton form could also be generated by endoglycosidase digestion of the secreted 59,000-dalton protein. In pulse-chase experiments, the 53,000-dalton protein was converted into the 59,000-dalton form. A 47,000-dalton form of hepatic lipase was immunoisolated from cell lysates only after tunicamycin treatment and could be generated from the secreted 59,000-dalton enzyme by prolonged endoglycosidase digestion. These data show that hepatic lipase is synthesized and rapidly secreted by isolated rat hepatocytes. Further, an intracellular 47,000-dalton precursor peptide can be identified after tunicamycin treatment, which may represent the hepatic lipase polypeptide, presumably after removal of its signal sequence; a 53,000-dalton partially glycosylated peptide exists as a major precursor form in the cell; and the mature 59,000-dalton hepatic lipase is present in the hepatocyte, but it is rapidly secreted.
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PMID:Hepatic lipase. Synthesis, processing, and secretion by isolated rat hepatocytes. 310 30

Synaptophysin is a major glycoprotein of Mr approximately 38,000 (in deglycosylated form: Mr approximately 34,000) characteristic of a certain class of small (30-80 nm diameter) neurosecretory vesicles, including presynaptic vesicles, but also vesicles of various neuroendocrine cells of both neuronal and epithelial phenotype. Using synaptophysin-specific antibodies we have isolated cDNA clones from rat nervous tissue libraries, which identify an approximately 2.5-kb mRNA in rat and human cells, including neuroendocrine tumours, that contains a reading frame for a polypeptide of 307 amino acids with a total mol. wt of 33 312. The deduced amino acid sequence, which was partly confirmed by comparison with sequences of two tryptic peptides obtained from purified synaptophysin, revealed four hydrophobic regions of 24 amino acids each, which are characterized, according to conformation prediction analyses, by marked alpha-helicity. The sequence shows a single potential N-glycosylation site, which is assigned to the vesicle interior, and a carboxy-terminal tail of 89 amino acids which contains glycine-rich tetrapeptide repeats, the epitope of monoclonal antibody SY38, and a number of collagenase-sensitive sites accessible on the surface of the intact vesicles. These features suggest that the polypeptide spans the vesicle membrane four times, with both N and C termini located on the outer, i.e. cytoplasmic, surface of the vesicles.
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PMID:Synaptophysin: molecular organization and mRNA expression as determined from cloned cDNA. 312 15

Type II pneumocytes secrete pulmonary surfactant and are known to synthesize SP-35, a collagenous surfactant-associated protein. Freshly isolated type II cells also synthesize other bacterial collagenase-sensitive and hydroxyproline-containing proteins, including a glycoprotein designated CP4. CP4 was isolated from rat pneumocyte culture medium by immune precipitation with polyclonal antibodies to rat surfactant proteins or by DEAE chromatography and reverse-phase or gel permeation HPLC. CP4 did not cross-react with polyclonal antibodies to SP-35 and was completely resolved from SP-35 by SDS-PAGE (Mr 43K reduced) or isoelectric focusing. Unlike SP-35, which consists of acidic isoforms assembled as disulfide-bonded dimers and multimers, CP4 was secreted as basic isoforms assembled as disulfide-bonded trimers. Differences in primary structure were demonstrated by CNBr and V8 protease peptide mapping. The secretion of both proteins was inhibited by 2,2'-dipyridyl, an inhibitor of posttranslational prolyl and lysyl hydroxylation and collagen triple helix formation. CP4 was isolated from EDTA extracts of rat surfactant. These studies provide evidence for the heterogeneity of pneumocyte-derived collagenous surfactant-associated proteins.
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PMID:CP4: a pneumocyte-derived collagenous surfactant-associated protein. Evidence for heterogeneity of collagenous surfactant proteins. 321 63

An 80 kDa glycoprotein was isolated from adult frog skeletal muscle by concanavalin (Con A) affinity chromatography and electrophoretic separation by molecular mass. Characteristics of the 80 kDa glycoprotein are that it: 1) binds non-covalently to gelatin-agarose or some other protein(s) bound to gelatin-agarose, 2) does not bind wheat germ agglutinin, 3) appears only at 80 kDa in both reducing and non-reducing electrophoretic separations, 4) is present in skeletal muscle but absent in smooth muscle and cardiac muscle, 5) is not collagenase or hyaluronidase-sensitive, and 6) is antigenically similar to a protein in embryonic chicken skeletal muscle. It was used to generate a polyclonal antiserum which was affinity-purified and used for immunolocalization. Indirect immunofluorescence procedures showed the antigen to be present on the surface of the skeletal muscle cells and concentrated at sites where cells are closely apposed to one another. Preparations in which adult muscle cells were depleted of basement membrane and endomysial proteins did not reduce the amount of 80 kDa protein present in skeletal muscle. These data indicate that this is a cell surface glycoprotein that may mediate attachment of the cell to extracellular proteins at sites where adjacent skeletal muscle cells are apposed.
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PMID:Identification of an 80 kDa glycoprotein located at sites of close apposition between skeletal muscle cells. 326 Aug 64


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