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

Human rheumatoid synovial cells in culture secrete at least three related metalloproteinases that digest extracellular matrix macromolecules. One of them, termed matrix metalloproteinase 2 (MMP-2), has been purified as an inactive zymogen (proMMP-2). The final product is homogeneous on SDS/PAGE with Mr = 72,000 under reducing conditions. The NH2-terminal sequence of proMMP-2 is Ala-Pro-Ser-Pro-Ile-Ile-Lys-Phe-Pro-Gly-Asp-Val-Ala-Pro-Lys-Thr, which is identical to that of the so-called '72-kDa type IV collagenase/gelatinase'. The zymogen can be rapidly activated by 4-aminophenylmercuric acetate to an active form of MMP-2 with Mr = 67,000, and the new NH2-terminal generated is Tyr-Asn-Phe-Phe-Pro-Arg-Lys-Pro-Lys-Trp-Asp-Lys-Asn-Gln-Ile. However, following 4-aminophenylmercuric acetate activation, MMP-2 is gradually inactivated by autolysis. Nine endopeptidases (trypsin, chymotrypsin, plasmin, plasma kallikrein, thrombin, neutrophil elastase, cathepsin G, matrix metalloproteinase 3, and thermolysin) were tested for their abilities to activate proMMP-2, but none had this ability. This contrasts with the proteolytic activation of proMMP-1 (procollagenase) and proMMP-3 (prostromelysin). The optimal activity of MMP-2 against azocoll is around pH 8.5, but about 50% of activity is retained at pH 6.5. Enzymic activity is inhibited by EDTA, 1,10-phenanthroline or tissue inhibitor of metalloproteinases, but not by inhibitors of serine, cysteine or aspartic proteinases. MMP-2 digests gelatin, fibronectin, laminin, and collagen type V, and to a lesser extent type IV collagen, cartilage proteoglycan and elastin. Comparative studies on digestion of collagen types IV and V by MMP-2 and MMP-3 (stromelysin) indicate that MMP-3 degrades type IV collagen more readily than MMP-2, while MMP-2 digests type V collagen effectively. Biosynthetic studies of MMPs using cultured human rheumatoid synovial fibroblasts indicated that the production of both proMMP-1 and proMMP-3 is negligible but it is greatly enhanced by the treatment with rabbit-macrophage-conditioned medium, whereas the synthesis of proMMP-2 is constitutively expressed by these cells and is not significantly affected by the treatment. This suggests that the physiological and/or pathological role of MMP-2 and its site of action may be different from those of MMP-1 and MMP-3.
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PMID:Matrix metalloproteinase 2 from human rheumatoid synovial fibroblasts. Purification and activation of the precursor and enzymic properties. 226 96

Mast cells have been implicated in the pathogenesis of the matrix degradation observed in the cartilaginous and osseous structures of the rheumatoid joint. We previously reported that human mast cell tryptase, a 134-kD granule-associated neutral protease, is present in rheumatoid synovium and can activate collagenase in crude culture medium in vitro. the present study attempts to depict the precise mechanism of this activation. To express full activation of latent collagenase, matrix metalloproteinase 3 (MMP-3) or stromelysin, can be activated by tryptase in a time and dose-dependent manner. Tryptase was not capable of generating active collagenase in the crude media from cultured rheumatoid synoviocytes depleted of proMMP-3 by immunoadsorption. In addition, the function of the tissue inhibitor of metalloproteinases (TIMP) was not altered by tryptase, and SDS-PAGE analysis revealed no degradation of TIMP by tryptase. The tryptase dependent activation of synoviocyte procollagenase thereby appears to be entirely dependent upon its ability to activate proMMP-3.
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PMID:Synovial procollagenase activation by human mast cell tryptase dependence upon matrix metalloproteinase 3 activation. 255 80

The treatment of crude culture medium from human rheumatoid synovial cells with 4-aminophenylmercuric acetate (APMA) or trypsin results in the activation of procollagenase. This process was shown to be dependent on the presence of matrix metalloproteinase 3 (MMP-3). MMP-3 can directly activate procollagenase without changing the apparent molecular weight of procollagenase. This activity was accelerated in the presence of APMA. We propose that MMP-3 plays an important role in connective tissue destruction through the activation of procollagenase in addition to its direct action on components of the extracellular matrix.
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PMID:Evidence that human rheumatoid synovial matrix metalloproteinase 3 is an endogenous activator of procollagenase. 284 49

Tissue inhibitor of metalloproteinases (TIMP) from cultured bovine dental pulp inhibits human rheumatoid synovial matrix metalloproteinase 3 (MMP-3) with a stoichiometry of 1:1 on a molar basis. Among the serine proteinases examined, human neutrophil elastase, trypsin and alpha-chymotrypsin destroyed the inhibitory activity of TIMP against MMP-3 by degrading the inhibitor molecule into small fragments. In contrast, the inhibitory activity of TIMP was not significantly reduced by the actions of cathepsin G, pancreatic elastase and plasmin. These data indicate that neutrophils which infiltrate tissues in various inflammatory conditions may play an important role in regulating TIMP activity in vivo through the action of neutrophil elastase.
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PMID:Inactivation of tissue inhibitor of metalloproteinases by neutrophil elastase and other serine proteinases. 316 16

A neutral metalloproteinase has been isolated and purified from adherent rheumatoid synovial cells in culture. This protease, named matrix metalloproteinase 3, (MMP-3) degrades gelatin, proteoglycan, fibronectin, type IV collagen, laminin, and the N propeptide of type I procollagen. It can be separated from MMP-2 (a potent gelatinase), and MMP-1, an interstitial collagenase. MMP-3 is released from cells as a proenzyme of 55 Kda. Activation by trypsin or organic mercurials produces 2 active species of 45 Kda and 28 Kda. The enzyme contains zinc as an intrinsic component and requires calcium for conformational stability. In concert, active MMP-1, -2, and -3 can destroy all significant structural proteins of joint structures.
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PMID:Matrix metalloproteinases 1, 2, and 3 from rheumatoid synovial cells are sufficient to destroy joints. 330 38

The precursor of matrix metalloproteinase 9 (pro-MMP-9, progelatinase B) noncovalently binds to tissue inhibitor of metalloproteinases (TIMP)-1 through the C-terminal domain of each molecule. We have isolated the proMMP-9.TIMP-1 complex from the medium of human fibrosarcoma HT-1080 cells and investigated the activation processes of the complex by 4-aminophenylmercuric acetate, trypsin, and matrix metalloproteinase 3 (MMP-3, stromelysin 1). The treatment of the proMMP-9.TIMP-1 complex with 4-aminophenylmercuric acetate or trypsin converts proMMP-9 to lower molecular weight species corresponding to active forms, but no gelatinolytic activity is detected. The lack of enzymic activity results from binding of TIMP-1 to the activated MMP-9. The treatment of the proMMP-9.TIMP-1 complex with a possible physiological proMMP-9 activator, MMP-3, does not reveal any gelatinolytic activity unless the molar ratio of MMP-3 to the complex exceeds 1. This is due to the inhibition of MMP-3 by TIMP-1 forming a ternary proMMP-9.TIMP-1.MMP-3 complex. The formation of the ternary complex weakens the interaction between proMMP-9 and TIMP-1, resulting in partial dissociation of the complex into proMMP-9 and the TIMP-1.MMP-3 complex. When MMP-3 is in excess, the propeptide is completely processed, and the full activity of MMP-9 is detected. Similarly, the proMMP-9.TIMP-1 complex inhibits MMP-1 (interstitial collagenase) and in turn renders the proMMP-9 activable by a catalytic amount of MMP-3. These results suggest that formation of the proMMP-9.TIMP-1 complex regulates extracellular matrix breakdown in tissue by switching the predominant MMP activity from one type to another.
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PMID:Steps involved in activation of the pro-matrix metalloproteinase 9 (progelatinase B)-tissue inhibitor of metalloproteinases-1 complex by 4-aminophenylmercuric acetate and proteinases. 762 79

The precursor of matrix metalloproteinase 3 (MMP-3/ stromelysin 1) is activated in vitro by proteinases or mercurial compounds by stepwise processes which include the initial formation of short-lived intermediates and the subsequent intermolecular cleavage of the His82-Phe83 bond to generate the fully activated mature MMP-3 (Nagase, H., Enghild, J. J., Suzuki, K., and Salvesen, G. (1990) Biochemistry 29, 5783-5789). To study the enzymatic properties of the intermediates we have mutated either His82 or Phe83 to Arg to obtain a stable MMP-3 intermediate. The mutant proteins were expressed in Chinese hamster ovary K-1 cells using a mammalian expression system. The proMMP-3(H82R) mutant was activated by chymotrypsin, elastase, and 4-aminophenylmercuric acetate to the 45-kDa MMP-3 with similar mechanism and kinetics as the wild-type. In contrast, the activation of the proMMP-3(F83R) mutant by proteinases or 4-aminophenylmercuric acetate resulted in 46-kDa forms, which retained 13, 14, or 15 amino acids of the pro-domain depending on the activators. The proteinase-activated MMP-3(F83R) intermediates exhibited little enzymatic activity, but they were partially active after treatment with SH-reacting reagents. These molecules could bind to the tissue inhibitor of metalloproteinases-1 and alpha 2-macroglobulin. However, the SH group of Cys75 of the intermediates was not modified by SH-reagents, indicating that the enzymatic activity generated by SH-reagents resulted from molecular perturbation of the enzyme rather than their interaction with Cys75. When gelatin and transferrin were digested with the 46-kDa intermediates the products were different from those generated by the wild-type MMP-3, suggesting an alteration in substrate specificity. The treatment of proMMP-3 with trypsin resulted in the formation of a 45-kDa MMP-3 with an NH2-terminal Thr85, whose activity and substrate specificity were similar to those of the 46-kDa MMp-3(F83R) obtained from the proMMP-3(F83R) mutant. These observations indicate that the correct processing at the His82-Phe83 bond is critical for expression of the full activity and the specificity of MMP-3.
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PMID:Characterization of the 46-kDa intermediates of matrix metalloproteinase 3 (stromelysin 1) obtained by site-directed mutation of phenylalanine 83. 863 80

Human kallikrein-related peptidases (KLKs) are 15 homologous serine proteases involved in several (patho)physiological processes, including cancer. Secreted as precursors, they are activated upon proteolytic release of a short pro-peptide. We searched for interconnection of KLKs within extracellular proteolytic networks leading to activation of protease zymogens and found that (i) pro-KLK activation by other KLKs is scarce, with the exception of pro-KLK11, which is efficiently activated by KLK4 and 5; (ii) pro-KLK4 is activated by matrix metalloproteinase 3; and (iii) trypsin-like KLKs efficiently activate the serine protease urokinase. Our observations provide new insights into the regulation of these important tumor-associated proteases.
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PMID:Interdependence of kallikrein-related peptidases in proteolytic networks. 2030 17

Osteoarthritis is characterized by a loss of extracellular matrix that leads to cartilage degradation and joint space narrowing. Specific proteases, including the aggrecanases ADAMTS-4 and matrix metalloproteinase 3, are important in initiating and promoting cartilage degradation in osteoarthritis. This study investigated protease-specific and disease-specific cleavage patterns of particular extracellular matrix proteins by comparing new peptide fragments, neopeptides, in specific exogenous protease-driven digestion of a crude cartilage proteoglycan extract and an in-vitro model of early osteoarthritis. Additionally, equine cartilage explants were treated with interleukin-1 and the media collected. Proteolytic cleavage products following trypsin digestion were then identified using tandem mass spectrometry. Complete sequences of proteolytically cleaved neopeptides were determined for the major cartilage proteoglycans aggrecan, biglycan, decorin, fibromodulin plus cartilage oligomeric matrix protein. The generation of neopeptides varied with enzyme specificity; however, some peptides were common to all samples. Previous known and novel cleavage sites were identifies. The identification of novel peptide fragments provides a platform for the development of antibodies that could assist in the identification of biomarkers for osteoarthritis (OA), as well as the identification of basic biochemical processes underlying OA.
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PMID:Characterization of neopeptides in equine articular cartilage degradation. 2612 2

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