Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.21.4 (trypsin)
 42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

1. A latent neutral proteinase was found in culture media of mouse bone explants. Its accumulation during the cultures is closely parallel to that of procollagenase; both require the presence of heparin in the media. 2. Latent neutral proteinase was activated by several treatments of the media known to activate procollagenase, such as limited proteolysis by trypsin, chymotrypsin, plasmin or kallikrein, dialysis against 3 M-NaSCN at 4 degrees C and prolonged preincubation at 25 degrees C. Its activation often followed that of the procollagenase present in the same media. 3. Activation of neutral proteinase (as does that of procollagenase) by trypsin or plasmin involved two successive steps: the activation of a latent endogenous activator present in the media followed by the activation of neutral proteinase itself by that activator. 4. The proteinase degrades cartilage proteoglycans, denatured collagen (Azocoll) and casein at neutral pH; it is inhibited by EDTA, cysteine or serum. Collagenase is not inhibited by casein or Azocoll and is less resistant to heat or to trypsin than is the proteinase. Partial separation of the two enzymes was achieved by gel filtration of the media but not by fractional (NH4)2SO4 precipitation, by ion exchange or by affinity chromatography on Sepharose-collagen. These fractionations did not activate latent enzymes. 5. Trypsin activation decreases the molecular weight of both latent enzymes (60 000-70 000) by 20 000-30 000, as determined by gel filtration of media after removal of heparin. 6. The latency of both enzymes could be due either to a zymogen or to an enzyme-inhibitor complex. A thermostable inhibitor of both enzymes was found in some media. However, combinations of either enzyme with that inhibitor were not reactivated by trypsin, indicating that this inhibitor is unlikely to be the cause of the latency.
 ...
PMID:The simultaneous release by bone explants in culture and the parallel activation of procollagenase and of a latent neutral proteinase that degrades cartilage proteoglycans and denatured collagen. 20 18

A method is described for the purification of clostridial collagenase from a crude enzyme preparation employing cation exchange chromatography on SP Sephadex, anion exchange chromatography on DEAE cellulose and gel filtration on Sephacryl S-200. Emphasis was placed on purity using continuous shallow gradients for the ion exchange separations to increase resolution and monitoring eluates both with respect to ultraviolet light absorption at 230 nm and analytical disc gel acrylamide electrophoresis. In addition, protein fractions were assayed for collagenolytic and non-specific proteolytic activity. The purity of the final preparation was assessed by acrylamide electrophoresis, gel filtration and amino acid analysis. The isolated enzyme hydrolyzed between 30 and 40% of rat tail tendon collagen in 1 h at 37 degrees C and lacked measurable trypsin or elastase-like activity.
 ...
PMID:A modified procedure for the purification of clostridial collagenase. 21 75

Antibodies to fibronectin and to distinct types of procollagens and collagens were used in immunofluorescent staining to localize these proteins in cell cultures. Normal human skin or lung fibroblasts produced a fibrillar pericellular matrix in which fibronectin and procollagen (types I and III) showed extensive codistribution. Fibronectin and procollagen were synthesized by the same cells as judged by double-stain immunofluorescence. Pericellular procollagen was specifically digested with collagenase without an effect on the fibrillar distribution of matrix fibronectin. Brief treatment with trypsin removed both matrix proteins. The human tumor cell lines HT-1080 (fibrosarcoma) and RD (rhabdomyosarcoma) produced little or no matrix fibronectin or procollagen. At sites of cell contact, simian virus 40-transformed lung fibroblasts (VA13) produced small amounts of pericellular fibrillar matrix fibronectin that codistributed with procollagen type I. Intracellular fibronectin and procollagen were visualized in all of these human sarcoma cell lines. When chicken embryo fibroblasts infected with a T class mutant (NY68) of Rous sarcoma virus temperature-sensitive for transformation were maintained at the nonpermissive temperature (41 degrees ) the cells had normal phenotype and a fibrillar matrix containing fibronectin and procollagen was present. At the permissive temperature (35 degrees ), the cells showed transformed phenotype and the matrix was lost. The failure to produce a pericellular fibronectin/collagen matrix may account for several phenotypic characteristics of transformed cultured fibroblasts.
 ...
PMID:Codistribution of pericellular matrix proteins in cultured fibroblasts and loss in transformation: fibronectin and procollagen. 21 6

Chemical and enzymatic properties of four collagenases newly isolated from anaerobic Clostridium histolyticum, aerobic Achromobacter iophagus, and from two lower eucaryotes, the fungus Entomophthora coronata and the insect Hypoderma lineatum are reviewed. The problems of their biosynthesis and precursors, namely the effect of induction of collagenase and neutral proteinase in Achromobacter by their macromolecular substrates are discussed. The two bacterial collagenases are Zn-metallo-enzymes; the highly purified Clostridium collagenase contains cyst(e)ine, serine phosphate and tryptophan additionally to amino acids reported previously. Achromobacter collagenase has the highest specific activity of all collagenases; it yields by autolysis enzymatically active degraded forms. The active dimer is composed of two identical subunits of molecular weight 35,000. Similarities between Achromobacter collagenase, thermolysin and Bacillus subtilis neutral proteinase in molecular weight, amino acid composition, and amino acids important for the active sites are discussed. The two collagenases from low eucaryotes are serine proteinases; Hypoderma collagenase is homologous to the trypsin family in the amino terminal sequence. The initial cleavage of native collagen by highly purified bacterial collagenases occurs in the central helical part of the alpha chains and not progressively from the amino terminal end. One of the two initial cleavages produced by Achromobacter collagenase is situated in the region cleaved specifically by vertebrate collagenases, but with different bond specificity. The same is true for the insect collagenase. Entomophthora collagenase is a proteinase of broad specificity which also cleaves collagen in its helical parts. All four collagenases also degrade other proteins according to their bond specificity.
 ...
PMID:Some newly characterized collagenases from procaryotes and lower eucaryotes. 22 May 20

The specificity of human skin collagenase and of an enzyme from an invasive tumor were studied by using types I, II, III, IV, and V (AB) collagen as substrates. Human skin collagenase degraded types I, II, and III collagen, producing the characteristic 3/4 and 1/4 cleavage products, but failed to degrade type IV or V collagen. Collagenase prepared from the invasive tumors showed maximal activity after trypsin treatment. The tumor enzyme degraded type IV (basement membrane) collagen, producing fragments consistent with a single cleavage site but did not attack types I, II, III, and V collagen. Because type IV collagen prepared by pepsinization of placenta was also digested, it is likely that cleavage of type IV collagen by the tumor collagenase occurs within a largely helical domain. A type IV collagenase could play a significant role in tumor metastases and in normal tissues where basement membrane turnover takes place.
 ...
PMID:Preferential digestion of basement membrane collagen by an enzyme derived from a metastatic murine tumor. 22 20

1. The addition of heparin to the culture fluid of mouse tibiae or calvaria did not cause any significant resorption of bone collagen or mineral. However, heparin (or analogue sulfated polyanions), enhanced greatly the amount of latent, trypsin-activatable collagenase (i.e. procollagenase) released by the bones in the medium without influencing that of directly active collagenase which was always very low. Heparin appeared to act by increasing the production of the enzyme which is immediately excreted. Procollagenase and collagenase are not stored in bone tissue, even under conditions where it is in active resorption. 2. Parathyroid hormone induced in the explants a resorption of both mineral and collagen that was inhibited by calcitonin. These hormones, however, had no influence on the release of procollagenase or collagenase either in the presence or in the absence of heparin. 3. Once activated, bone collagenase digested the collagen of the bone explants, and more extensively after their demineralization. Thus the latent collagenase that accumulates around non-resorbing bones has to be considered as a precursor, (and not as a residue), of active enzyme. 4. Active collagenase added to incipient cultures of bones disappeared with a half-life of 24 h. The lost enzyme could, however, not be reactivated by trypsin and thus was not transformed into latent procollagenase.
 ...
PMID:Collagenase, procollagenase and bone resorption. Effects of heparin, parathyroid hormone and calcitonin. 22 39

Biphasic kinetic data were obtained when trypsin (EC 3.4.21.4) which had previously been complexed with a thiol-containing inhibitor (present in Ehrlich ascites tumour cells) was incubated with incremental additions of periodate. At low concentrations of periodate the trypsin was re-activated whilst at higher concentrations of periodate the trypsin was irreversibly inhibited. This biphasic reactivation followed by inhibition was also demonstrated when trypsin was first inhibited by dithiothreitol and followed by incremental addition of periodate. Similar results were obtained with chymotrypsin (EC 3.4.21.1). Incremental additions of either dithiothreitol or periodate caused inhibition of both these enzymes. The biphasic kinetic data can be explained in terms of reduction and oxidation of a significant disulphide bond in both trypsin and chymotrypsin which can be cleaved by thiols in a disulphide exchange reaction [1]. This bond is thought to maintain the active centres of each of these enzymes in a conformation sterically favourable for enzymic cleavage of specific peptide bonds in the protein substrates (polymeric collagen fibrils and casein) employed in this study.
 ...
PMID:Inhibition of trypsin and chymotrypsin by thiols. Biphasic kinetics of reactivation and inhibition induced by sodium periodate addition. 22 48

In this study, we investigated the possible regulatory role of collagen in collagenase production by cultured human skin fibroblasts and human and rabbit synovial cells. Addition of types I, II or III collagen in solution to the culture media markedly stimulated trypsin-activable collagenase activity in these cultures. In the human cell cultures the stimulatory effect of collagen was further enhanced by a soluble factor isolated from human monocyte culture media (Dayer, Russell and Krane, 1977). Both native and denatured forms of collagen stimulated enzyme production; their relative efficacy varied among the different types. The native form of both types I and II collagen showed a greater effect on collagenase production than the corresponding denatured form, whereas with type III collagen the denatured form was more effective.
 ...
PMID:Stimulation of collagenase production by collagen in mammalian cell cultures. 22 68

The collagenase from the larvae Hypoderma lineatum, with a molecular weight of 24 000 and isoelectric point of 4.1, was obtained in homogeneous form by ion-exchange chromatography. It is stoichiometrically inhibited by diisopropylfluorophosphate. On the other hand it is unaffected by ethylenediaminetetraacetate, p-chloromercuribenzoate, dithiothreitol, N-tosyllysine chloromethyl ketone, N-tosylphenylalanine chloromethyl ketone and ovomucoid trypsin inhibitor. The enzyme which degrades native collagen in its helical parts, has a specific activity on thermally reconstituted collagen fibrils of 150 micrograms collagen degraded x min-1 x (mg enzyme)-1 at 37 degrees C. It hydrolyses casein but has no esterolytic activity characteristic of trypsin, chymotrypsin nor elastase. It has no action on the synthetic peptide 4-phenylazobenzyloxycarbonyl-L-prolyl-L-leucyl-L-glycyl-L-prolyl-D-arginine. The amino acid composition of Hypoderma collagenase indicates a distinct similarity with the serine proteinases of the trypsin family and with another athropode serine collagenase, that of the fiddler crab Uca pugilator. This suggests that eucaryotic collagenases with digestive rather than morphogenic function represent a new category of members of the trypsin family.
 ...
PMID:Chemical and enzymatic characterization of the collagenase from the insect Hypoderma lineatum. 23 30

The synovial fluid (SF) of RA patients contains large amounts of PMN which are well equipped with neutral enzymes to degrade articular cartilage: elastase and cathepsin G, which both destroy proteoglycans and native collagen, as well as 2 types of collagenoases. Indirect evidence suggests that PMN might be important in the destruction of RA articular cartilage. In 19 SF of RA patients no free elastase or collagenase was found. Using immune histochemical methods, we observed that PMN and macrophages of SF contain both elastase and alpha 1-anti-trypsin and alpha 2-macroglobulin. Peripheral PMN - but not monocytes - contain elastase, however both types of cells lack alpha 1-antitrypsin and alpha 2-macroglobulin. Elastase is demonstratable in the superficial layer of pannus free RA articular cartilage. These findings suggest that neutral proteinases from PMN in RA SF are generally neutralized by physiologic inhibitors and removed by phagocytes. The enzyme-inhibitor interaction might be bypassed during "frustrated phagocytosis" so that enzymes like PMN elastase can damage RA articular cartilage.
 ...
PMID:[Chronic polyarthritis: role of polymorphonuclear leukocytes in the destruction of pannus-free articular cartilage]. 23 68


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