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

---

Query: EC:3.4.21.4 (trypsin)
42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Proteoglycans of bovine compact bone were purified by chromatography of the formic acid precipitate of an EDTA extract. The sequential chromatographic steps consisted of gel filtration on Sepharose CL-6B in 4-M guanidine HCl, ion-exchange chromatography on DEAE-Sephacel in 4-M urea and rechromatography on Sepharose CL-6B in 4-M guanidine HCl. The preparation consisted of a relatively small proteoglycan (Kav = 0.4 on Sepharose CL-6B) containing about 40% protein, 21% hexuronic acid, 23% galactosamine and lesser amounts of other monosaccharides. The core protein was shown by gradient NaDodSO4 gel electrophoresis, electrotransfer and immunodetection to be monodispersed with an Mr = 45,000. Analysis of glycopeptides obtained after papain digestion of the proteoglycan and separation from glycosaminoglycan chains by gel chromatography, indicated that both N-linked and O-linked oligosaccharides were present. The glycosaminoglycan chains liberated by papain digestion eluted from Sepharose CL-6B as a broad peak with Kav = 0.50, slightly ahead of the position of elution of bovine nasal cartilage glycosaminoglycans (Kav = 0.52); the bone glycosaminoglycans are thus slightly larger than those from cartilage and smaller than the ones attached to fetal bone proteoglycans. These chains were totally susceptible to chondroitinase AC II, a procedure that yielded unsaturated disaccharides corresponding predominantly to chondroitin-4-sulfate, and to a lesser extent chondroitin-6-sulfate. Antisera raised against adult bone proteoglycans cross-reacted with core protein of bone proteoglycan (obtained after chondroitinase digestion) but not with papain digested proteoglycan. In addition, they cross-reacted with core protein and trypsin-liberated, chondroitin sulfate rich region (AlTAl) derived from cartilage proteoglycans and, to a lesser extent, rat bone proteoglycans. No cross-reactivity could be detected to Smith-degraded cartilage proteoglycans, bone acidic glycoproteins or serum proteins.
...
PMID:Proteoglycans of adult bovine compact bone. 293 15

Previous studies have reported an increase in heparan sulfate glycosaminoglycan (HSGAG) during skeletal muscle differentiation in culture. We have investigated this phenomenon further in relation to the heparan sulfate proteoglycans (HSPG) produced by myogenic cultures. Pulse-chase analysis indicated an approx. 3-fold increase in heparan sulfate synthesis in myotube cultures over that in proliferating or aligning myoblast cultures. Muscle fibroblast culture heparan sulfate synthesis was higher than that of myoblasts but was lower than myotubes. The turnover rates appeared to be the same for all stages of development, with a t1/2 of approx. 5 h. Enrichment for heparan sulfate by Sepharose CL-4B and DEAE-Sephacel chromatography indicated an increase in the hydrodynamic size of the proteoglycan produced by myotubes over that from myoblasts, with a shift in Kav from 0.14-0.19 to 0.07. Fibroblasts synthesized the smallest proteoglycan, with a Kav of 0.22. All of the proteoglycans contained similar sized glycosaminoglycan chains with an estimated molecular weight of 30,000-40,000. Localization of the heparan sulfate proteoglycan in myotube cultures by trypsin sensitivity indicated much of the intact proteoglycan to be closely associated with the cell surface, while internalized material appeared in a degraded form.
...
PMID:Biosynthesis of heparan sulfate proteoglycans of developing chick breast skeletal muscle in vitro. 294 1

Rat glomerular heparan sulfate (HS) and dermatan sulfate (DS) proteoglycan synthesis was studied in vitro and in vivo. Incorporation of [35S]sulfate into macromolecules was linear over 16 h in vitro, and DS was the predominant glycosaminoglycan (GAG), while HS dominated in vivo incubations. Proteoglycans were found in the bottom 2/5 (high density) CsCl gradient fractions and eluted as two overlapping peaks from DEAE-Sephacel columns. The proportion of low density 35S-glycoproteins and 35S-proteoglycans increased with time. Two high buoyant density HS proteoglycans were extracted from glomeruli and eluted in DEAE peak I. The first, HS-tIA, had an Mr of 130 X 10(3) with Mr 12.5 X 10(3) GAG chains. This proteoglycan was released from the tissue by trypsin and was partially displaced by heparin treatment. In addition, it was rapidly released into the medium of label-chase experiments after which it migrated slightly more rapidly than HS-tIA in gels, with HS chains similar in length to its tissue counterpart. The second, HS-tIB, had an Mr of 8.6 X 10(3) with little or no attached protein. This proteoglycan was characterized as intracellular as it resisted release by trypsin treatment or heparin extraction in medium and was not detected in the medium of label-chase experiments. Two tissue DS proteoglycans were characterized. The first, DS-tIA, co-purified with HS-tIA and was the predominant proteoglycan synthesized during 4-h in vitro incubations. Like HS-tIA, it was rapidly released into medium and displaced from cell surfaces or tissue "receptors" by heparin or trypsin treatments. A second, Sepharose CL-6B-excluded DS proteoglycan from DEAE peak II, DS-tII, accumulated in tissue over 16 h in vitro. This proteoglycan was self-associating and contained clusters of iduronic acid residues along its Mr 26 X 10(3) DS chains. It resisted extraction from the tissue with heparin, trypsin, and detergent. No DS-tII was detected in the incubation medium. Instead, medium proteoglycans eluted as single Sepharose CL-6B-included peaks. DS chains from medium proteoglycans were shorter (Mr 18 X 10(3)) and had more regularly spaced iduronic acid residues than GAGs from DS-tII. The length and sulfation patterns of DS-mII GAG were similar to GAG from DS-tIA. Thus, glomeruli rapidly synthesized and released Sepharose CL-6B-included heparin-displaceable DS and HS proteoglycans while retaining a Sepharose CL-6B-excluded self-associating DS proteoglycan and an intracellular HS.
...
PMID:Partial characterization of heparan and dermatan sulfate proteoglycans synthesized by normal rat glomeruli. 294 88

A rat hepatocyte cell line was cultured in Higuchi's medium with fetal calf serum and insulin and labeled with 35SO2/4-. The cells were treated with a number of ligands to displace the heparan 35SO4 proteoglycan (HSPG) from the pericellular matrix. Maximum release was obtained with D-mannose-6-PO4 (50 mM), D-glucose-6-PO4 (50 mM), myo-inositol-2-PO4 (2-5 mM), myo-inositol hexaphosphate (2-5 mM), and DL-myo-inositol-1-PO4 (1-2 mM). D-myo-Inositol-1,3,4-(PO4)3 (1 mM) and L-myo-inositol-1-PO4 (2 mM) were intermediate in their ability to release the cell surface HSPG, whereas heparin (2 mg/ml), yeast phosphomannan (4 mg/ml), D-xylose-1-PO4 (50 mM), D-glucose-6-SO4 (50 mM), and myo-inositol hexasulfate (5 mM) were ineffective. When 35SO2/4- was added to cell cultures, the total cell surface HSPG increased linearly, but the percentage of the total cell surface [35SO4]HSPG that was released by myo-inositol-PO4 increased with time during the labeling period, reaching a maximum of 65% after 5 h. When cells were labeled for 12 h without insulin in the medium, the maximum amount of cell surface HSPG that was released by myo-inositol-PO4 was reduced to 30%. However, when cells labeled in the absence of insulin were treated with phosphatidylinositol-specific phospholipase C and then myo-inositol-PO4, the release of the cell surface [35SO4]HSPG was increased to 73%. When the [35SO4]HSPG that was released from the cell surface by treatment with myo-inositol-PO4 was added to cultures of unlabeled hepatocytes, it was taken up very rapidly and a portion of the internalized HSPG was converted to free heparan SO4 chains which appeared in the nucleus. Uptake was Ca2+- and Mg2+-independent. The amount of [35SO4]HSPG taken up was markedly reduced when the myo-inositol-PO4-releasable [35SO4]HSPG was pretreated with trypsin, thermolysin, alkaline borohydride, or alkaline phosphatase. When the cells were grown in inositol-deficient medium or in the presence of myo-inositol-PO4, the amount of heparan SO4 found in the nucleus was markedly reduced, and the cells no longer exhibited contact inhibition. These effects of myo-inositol deficiency on the growth and nuclear heparan SO4 were accentuated by addition of LiCl to the cultures to prevent phosphatidylinositol synthesis from the endogenous myo-inositol-PO4.(ABSTRACT TRUNCATED AT 400 WORDS)
...
PMID:Involvement of phosphatidylinositol and insulin in the coordinate regulation of proteoheparan sulfate metabolism and hepatocyte growth. 295 71

The proteoglycan (PG) on the surface of NMuMG mouse mammary epithelial cells consists of at least two functional domains, a membrane-intercalated domain which anchors the PG to the plasma membrane, and a trypsin-releasable ectodomain which bears both heparan and chondroitin sulfate chains. The ectodomain binds cells to collagen types I, III, and V, but not IV, and has been proposed to be a matrix receptor. Because heparin binds to the adhesive glycoproteins fibronectin, an interstitial matrix component, and laminin, a basal lamina component, we asked whether the cell surface PG also binds these molecules. Cells harvested with either trypsin or EDTA bound to fibronectin; binding of trypsin-released cells was inhibited by the peptide GRGDS but not by heparin, whereas binding of EDTA-released cells was inhibited only by a combination of GRDS and heparin, suggesting two distinct cell binding mechanisms. In the presence of GRGDS, the EDTA-released cells bound to fibronectin via the cell surface PG. Binding via the cell surface PG was to the COOH-terminal heparin binding domain of fibronectin. In contrast with the binding to fibronectin, EDTA-released cells did not bind to laminin under identical assay conditions. Liposomes containing the isolated intact cell surface PG mimic the binding of whole cells. These results indicate that the mammary epithelial cells have at least two distinct cell surface receptors for fibronectin: a trypsin-resistant molecule that binds cells to the sequence RGD and a trypsin-labile, heparan sulfate-rich PG that binds cells to the COOH-terminal heparin binding domain. Because the cell surface PG binds cells to the interstitial collagens (types I, III, and V) and to fibronectin, but not to basal lamina collagen (type IV) or laminin, we conclude that the cell surface PG is a receptor on epithelial cells specific for interstitial matrix components.
...
PMID:Cell surface proteoglycan binds mouse mammary epithelial cells to fibronectin and behaves as a receptor for interstitial matrix. 296 12

Cultured mouse mammary (NMuMG) cells produce heparan sulfate-rich proteoglycans that are found at the cell surface, in the culture medium, and beneath the monolayer. The cell surface proteoglycan consists of a lipophilic membrane-associated domain and an extracellular domain, or ectodomain, that contains both heparan and chondroitin sulfate chains. During culture, the cells release into the medium a soluble proteoglycan that is indistinguishable from the ectodomain released from the cells by trypsin treatment. This medium ectodomain was isolated, purified, and used as an antigen to prepare an affinity-purified serum antibody from rabbits. The antibody recognizes polypeptide determinants on the core protein of the ectodomain of the cell surface proteoglycan. The reactivity of this antibody was compared with that of a serum antibody (BM-1) directed against the low density basement membrane proteoglycan of the Englebarth-Holm-Swarm tumor (Hassell, J. R., W. C. Leyshon, S. R. Ledbetter, B. Tyree, S. Suzuki, M. Kato, K. Kimata, and H. Kleinman. 1985. J. Biol. Chem. 250:8098-8105). The BM-1 antibody recognized a large, low density heparan sulfate-rich proteoglycan in the cells and in the basal extracellular materials beneath the monolayer where it accumulated in patchy deposits. The affinity-purified anti-ectodomain antibody recognized the cell surface proteoglycan on the cells, where it is seen on apical cell surfaces in subconfluent cultures and in fine filamentous arrays at the basal cell surface in confluent cultures, but detected no proteoglycan in the basal extracellular materials beneath the monolayer. The amino acid composition of the purified medium ectodomain was substantially different from that reported for the basement membrane proteoglycan. Thus, NMuMG cells produce at least two heparan sulfate-rich proteoglycans that contain distinct core proteins, a cell surface proteoglycan, and a basement membrane proteoglycan. In newborn mouse skin, these proteoglycans localize to distinct sites; the basement membrane proteoglycan is seen solely at the dermal-epidermal boundary and the cell surface proteoglycan is seen solely at the surfaces of keratinocytes in the basal, spinous, and granular cell layers. These results suggest that although heparan sulfate-rich proteoglycans may have similar glycosaminoglycan chains, they are sorted by the epithelial cells to different sites on the basis of differences in their core proteins.
...
PMID:Mouse mammary epithelial cells produce basement membrane and cell surface heparan sulfate proteoglycans containing distinct core proteins. 296 52

Rabbit synovial fibroblasts induced to undergo a specific switch in gene expression by agents that alter cell morphology secreted the neutral proteinase precursor procollagenase (apparent Mr of 53,000 and 57,000). A major Mr = 51,000 polypeptide that was always induced coordinately with procollagenase has now been identified as the proenzyme form of a metal-dependent proteinase active at neutral pH. We have named this proteinase stromelysin. Prostromelysin and procollagenase were the most prominent [35S]methionine-labeled secreted proteins of the induced fibroblasts. By the use of casein degradation as an assay for enzyme activity, stromelysin was isolated with high yield from the conditioned culture medium of 12-O-tetradecanoylphorbol 13-acetate-treated fibroblasts and migrated as an active form of Mr = 21,000 that was immunologically identical to the proteoglycan-degrading proteinase purified from rabbit bone. Immunoglobulin G from antiserum raised to purified rabbit bone proteoglycanase immunoprecipitated the Mr = 51,000 proenzyme form from conditioned medium of induced rabbit cells and also immunoprecipitated an Mr = 55,000 polypeptide from induced human fibroblasts. When rabbit prostromelysin was activated by trypsin or 4-aminophenylmercuric acetate, the proenzyme was converted to an active form of Mr = 41,000. During the course of the purification, prostromelysin was converted to an additional activatable form of Mr = 35,000 and additional active forms of Mr = 21,000-25,000, which had related peptide maps distinct from collagenase. All of these forms were immunologically cross-reactive. Purified stromelysin degraded casein, cartilage proteoglycans, fibronectin, alpha 1-proteinase inhibitor, and immunoglobulin G2a and had limited activity on laminin, elastin, type IV collagen, and gelatin, but did not degrade type I collagen. Stromelysin was inhibited by EDTA, 1,10-phenanthroline, and the specific glycoprotein tissue inhibitor of metalloproteinases isolated from human amniotic fluid and was therefore classified as a metalloproteinase.
...
PMID:Stromelysin, a connective tissue-degrading metalloendopeptidase secreted by stimulated rabbit synovial fibroblasts in parallel with collagenase. Biosynthesis, isolation, characterization, and substrates. 299 74

Pig synovial and human skin fibroblast procollagenases were treated with highly purified tryptase, the major proteinase of human mast cells, to determine whether this trypsin-like proteinase could activate the latent form of collagenase and so be involved in connective tissue breakdown. No significant activation of either human or pig procollagenase was found, but the highest concentration of tryptase partially destroyed procollagenase. Tryptase did not degrade type I collagen or proteoglycan. These data indicate that human mast cell tryptase does not contribute to connective tissue breakdown via procollagenase activation or via proteoglycan degradation.
...
PMID:Human lung mast cell tryptase fails to activate procollagenase or degrade proteoglycan. 299 69

Studies on the effect of leukocyte elastase on the metabolism of chondrocytes in culture have demonstrated that these cells possess a specific cell surface receptor for leukocyte-derived elastase. Purified elastase from rabbit and human leukocytes is capable of modulating the metabolism of the cell by causing a marked decrease in both proteoglycan and protein biosynthesis. Addition of 125I-labeled elastase to chondrocytes maintained in suspension culture has shown that binding occurs, and that it is saturable and is inhibited by the addition of unlabeled enzyme. We ascertained that the active site of the enzyme was necessary for binding to the chondrocyte, since phenylmethylsulfonyl fluoride-inactivated leukocyte elastase failed to bind. Pancreatic elastase had only a slight affinity for the receptor, whereas trypsin and bovine serum albumin failed to bind to any significant extent. Autoradiographic studies and the use of inhibitors of endocytosis, such as dansyl cadaverine, confirmed that endocytosis of elastase was the secondary event after cell binding.
...
PMID:Receptor-mediated binding of leukocyte elastase by chondrocytes. 303 97

Culture medium from rabbit uterine cervical fibroblasts contained a procollagenase and a neutral proproteinase which acts as a procollagenase activator. These two proenzymes have been purified by a combination of ion-exchange, affinity and gel chromatographies. The purified neutral proproteinase showed Mr 60,000 with sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. This neutral proproteinase was activated by trypsin, 4-aminophenylmercuric acetate (APMA) and plasmin, and the active species of the proteinase had Mr 53,000 when activated by APMA; kallikrein and urokinase did not activate this proproteinase. The purified neutral proteinase was inhibited by EDTA, 1,10-phenanthroline and rabbit plasma, but not by serine proteinase inhibitors, suggesting that this proteinase is a metal-dependent proteinase. The purified enzyme could also degrade gelatin, casein, proteoglycan and type IV procollagen. The purified procollagenase had Mr 55,000 and was activated by trypsin, APMA and the active neutral proteinase. These activations were accompanied by decrease in Mr, and the activated species had an Mr which was approx. 10,000 less than that of the procollagenase. In particular, procollagenase activation with neutral proteinase depended on incubation time and proteolytic activity of proteinase. These results indicate that activation of procollagenase by the rabbit uterine neutral proteinase is related to limited proteolysis in the procollagenase molecule.
...
PMID:Procollagenase activator produced by rabbit uterine cervical fibroblasts. 303 65


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>

---