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)

Squash inhibitors of serine proteinases form an uniform family of small proteins. They are built of 27-33 amino-acid residues and cross-linked with three disulfide bridges. The reactive site peptide bond (P1-P1') is between residue 5 (Lys, Arg or Leu) and 6 (always Ile). High resolution X-ray structures are available for two squash inhibitors complexed with trypsin. NMR solution structures have also been determined for free inhibitors. The major structural motif is a distorted, triple-stranded antiparallel beta-sheet. A similar folding motif has been recently found in a number of proteins, including: conotoxins from fish-hunting snails, carboxypeptidase inhibitor from potato, kalata B1 polypeptide, and in some growth factors (e.g. nerve growth factor, transforming growth factor beta 2, platelet-derived growth factor). Squash inhibitors are highly stable and rigid proteins. They inhibit a number of serine proteinases: trypsin, plasmin, kallikrein, blood clotting factors: Xa and XIIa, cathepsin G. The inhibition spectrum can be much broadened if specific amino-acid substitutions are introduced, especially at residues which contact proteinase. Squash inhibitors inhibit proteinases via the standard mechanism. According to the mechanism, inhibitors are substrates which exibit at neutral pH a high kcat/K(m) index for hydrolysis and resynthesis of the reactive site, and a low value of the hydrolysis constant.
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PMID:Squash inhibitor family of serine proteinases. 892 25

A cDNA encoding of the serine proteinase inhibitor (serpin), B-43, was cloned from the cDNA library of the bovine brain. It encoded 378 amino acids, and the MW of the protein was estimated to be 42.6 kDa, which is consistent with that of the native B-43 purified from the bovine brain. The homology search revealed that B-43 belongs to the ovalbumin branch of the serpin superfamily. Among them, B-43 was most homologous to human placental thrombin inhibitor (PI-6) and its murine counterpart, with the amino acid identity of 76% and 71%, respectively. Northern blot analysis showed that the size of the transcript was 1.4 kb, and that the expression of B-43 in the bovine brain varied depending on the brain regions, i.e. a lower level of expression was observed in the cerebral cortex and the hippocampus compared to the level of expression that was observed in the medulla oblongata. [35S]-labeled B-43 protein was synthesized in vitro by using a rabbit reticulocyte lysate system, which formed complexes with proteinases such as thrombin, trypsin, alpha-chymotrypsin, and 7S nerve growth factor (NGF), but not with urokinase or plasmin. These results, together with the immunohistochemical localization of B-43 in astrocytes and in some neurons which was observed in the previous study suggest that B-43 may be involved in the regulation of serine proteinases present in the brain or extravasated from the blood.
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PMID:Cloning of a serine proteinase inhibitor from bovine brain: expression in the brain and characterization of its target proteinases. 901 86

We reported previously that two epidermal growth factor receptor ligands, epidermal growth factor and transforming growth factor-alpha, inhibit medial septal cholinergic cell phenotypic expression (choline acetyltransferase and acetylcholinesterase activities) in vitro indirectly via (a) soluble molecule(s) released from astrocytes [Kenigsberg R. L. et al. (1992) Neuroscience 50, 85-97; Kenigsberg R. L. and Mazzoni I. E. (1995) J. Neurosci. Res. 41, 734-744; Mazzoni I. E. and Kenigsberg R. L. (1996) Brain Res. 707, 88-99]. In the present study, we found that this response to transforming growth factor-alpha is mediated, for the most part, by alpha 2-macroglobulin, a potent protease inhibitor with a wide spectrum of biological activities. In this regard, the effects of transforming growth factor-alpha on cholinergic cells can be blocked with immunoneutralizing antibodies raised against alpha 2-macroglobulin. Furthermore, western blot analysis reveals that although alpha 2-macroglobulin is present in conditioned media from control septal cultures, it is more abundant in those treated with transforming growth factor-alpha. In addition, exogenous alpha 2-macroglobulin, both in its native and trypsin-activated forms, can mimic transforming growth factor-alpha's effects on septal cholinergic cell expression. However, while the native antiprotease can slightly but significantly decrease choline acetyltransferase activity, trypsin-activated alpha 2-macroglobulin, in the nanomolar range, induces as marked a decrease in this enzyme activity as that noted with transforming growth factor-alpha. Furthermore, trypsin-activated alpha 2-macroglobulin, like epidermal growth factor/transforming growth factor-alpha, decreases choline acetyltransferase activity by arresting its spontaneous increase that occurs with time in culture, does so in a reversible manner and is not neurotoxic. In addition, trypsin-activated alpha 2-macroglobulin, in the nanomolar range, can affect choline acetyltransferase in a dual manner, up-regulating it at low concentrations while down-regulating it at higher ones. These responses are identical in mixed neuronal-glial and pure neuronal septal cultures. Furthermore, when concentrations of trypsin-activated alpha 2-macroglobulin, which alone decrease choline acetyltransferase, are added simultaneously with nerve growth factor, they serve to potentiate the nerve growth factor-induced increase in enzymatic activity. As GABAergic cell expression is not affected by alpha 2-macroglobulin, it appears that the effects of this protease inhibitor on medial septal neuronal expression are neurotransmitter-specific. Finally, trypsin-activated but not native alpha 2-macroglobulin promotes a dose-dependent aggregation of the septal neurons. This change in morphology, however, is not related to those noted in choline acetyltransferase activity. In summary, these data suggest that the expression of alpha 2-macroglobulin in astroglia from the medial septal nucleus can be controlled by epidermal growth factor receptor ligands to impact the functioning of basal forebrain cholinergic neurons.
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PMID:Transforming growth factor-alpha's effects on astroglial-cholinergic cell interactions in the medial septal area in vitro are mediated by alpha 2-macroglobulin. 933 Mar 64

Mast cells are the principal effector cells in IgE-dependent hypersensitivity reactions. Despite reports that rodent mast cells proliferate in the presence of nerve growth factor (NGF), human mast cells reportedly do not respond to this factor. To determine if human mast cells express the NGF receptors, TrkA tyrosine receptor and the low affinity NGF receptor (LNGFR), we first analyzed the mRNA expression by RT-PCR of TrkA and LNGFR in a human mast cell line (HMC-1) and in human mast cells cultured in the presence of stem cell factor. Both HMC-1 and cultured human mast cells were found to express TrkA but not LNGFR. TrkA protein was demonstrated by Western blot analysis of HMC-1 lysates. Using flow cytometric analysis and mast cell tryptase as a mast cell marker, both HMC-1 cells and cultured human mast cells were shown to coexpress tryptase and TrkA. Treatment of mast cells with NGF resulted in phosphorylation of TrkA on tyrosine residues as detected by immunoblotting with an antiphosphotyrosine antibody. Furthermore, NGF induced the immediate early gene c-fos in HMC-1 cells. HMC-1 cells and cultured human mast cells were also found to express NGF mRNA, and conditioned medium from HMC-1 cells stimulated neurite outgrowth from chicken embryonic sensory ganglia in culture. This effect was blocked by anti-NGF. Thus, mast cells express functional TrkA and synthesize NGF, suggesting a mechanism by which NGF may act as an autocrine factor for human mast cells, and by which mast cells and nerves may interact.
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PMID:Human mast cells express functional TrkA and are a source of nerve growth factor. 934 72

A cDNA clone for the serine proteinase inhibitor (serpin), neuroserpin, was isolated from a human whole brain cDNA library, and recombinant protein was expressed in insect cells. The purified protein is an efficient inhibitor of tissue type plasminogen activator (tPA), having an apparent second-order rate constant of 6. 2 x 10(5) M-1 s-1 for the two-chain form. However, unlike other known plasminogen activator inhibitors, neuroserpin is a more effective inactivator of tPA than of urokinase-type plasminogen activator. Neuroserpin also effectively inhibited trypsin and nerve growth factor-gamma but reacted only slowly with plasmin and thrombin. Northern blot analysis showed a 1.8 kilobase messenger RNA expressed predominantly in adult human brain and spinal cord, and immunohistochemical studies of normal mouse tissue detected strong staining primarily in neuronal cells with occasionally positive microglial cells. Staining was most prominent in the ependymal cells of the choroid plexus, Purkinje cells of the cerebellum, select neurons of the hypothalamus and hippocampus, and in the myelinated axons of the commissura. Expression of tPA within these regions is reported to be high and has previously been correlated with both motor learning and neuronal survival. Taken together, these data suggest that neuroserpin is likely to be a critical regulator of tPA activity in the central nervous system, and as such may play an important role in neuronal plasticity and/or maintenance.
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PMID:Neuroserpin, a brain-associated inhibitor of tissue plasminogen activator is localized primarily in neurons. Implications for the regulation of motor learning and neuronal survival. 940 89

Glial cells play an important role in maintaining neural function. In the present study, we examined the effects of a factor derived from human astrocytoma cells (1321N1) on differentiation of rat pheochromocytoma cells (PC-12). The conditioned medium which had been used for culture of 1321N1 cells caused the differentiation of PC-12 cells, suggesting that 1321N1 cells release a neurotrophic factor. The factor was apparently distinct from well-known neurotrophic factors, such as nerve growth factor (NGF), since it was resistant to boiling and trypsin treatment. The molecular size of the factor was assumed to be below 1000 through dialysis and ultrafiltration experiments. Furthermore, PC-12 cells were differentiated synergistically by the combined addition of NGF and the conditioned medium of 1321N1 cells. Partially purified fraction of the factor by Sephadex G-15 gel filtration column caused the prolonged activation of mitogen-activated protein kinase (MAPK). The differentiation of PC-12 cells induced by the fraction or NGF disappeared after the treatment with PD98059, a specific inhibitor of MAPK kinase (MEK), suggesting the involvement of MAPK in the differentiation. These results suggest that the new low-molecular factor derived from glial cells causes differentiation of PC-12 cells mediated through an activation of MAPK.
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PMID:A new factor derived from 1321N1 human astrocytoma cells causes differentiation of PC-12 cells mediated through mitogen-activated protein kinase cascade. 973 11

We have previously shown that fibroblast and keratinocyte supernatants up-regulate expression of mast cell characteristics in the human immature mast cell line HMC-1. This effect could not be induced in HMC-1 cells by the well-known mast cell growth factor stem cell factor (SCF), probably due to mutations of the SCF receptor c-Kit in these cells. Here we report the effects of several known fibroblast- and keratinocyte-derived growth factors, namely nerve growth factor (NGF), basic fibroblast growth factor, platelet-derived growth factor and transforming growth factor-beta, on mast cell differentiation, using HMC-1 cells as a model. NGF, at 0.1-50 ng/ml concentrations, caused a marked, dose-dependent up-regulation of tryptase, Fc epsilon RI and histamine within 10 days of culture, associated with an enhanced expression of mRNA for Fc epsilon RI and mast cell tryptase. On restriction analysis, only mast cell beta-tryptase, but not alpha-tryptase, could be demonstrated. Furthermore, the high-affinity NGF receptor (TrkA) was found at both the transcriptional and protein levels, while expression of the low-affinity NGF receptor was detectable at the mRNA level only. None of the other growth factors caused a significant alteration of the mast cell markers studied when added to HMC-1 cells at concentrations known to be biologically active in other culture systems. Immature human mast cells are thus induced to assume a more mature phenotype in vitro in response to NGF, most probably via stimulation of the high-affinity NGF receptor expressed on these cells. Besides SCF, NGF should therefore be considered as an additional mast cell growth factor that contributes to human mast cell maturation at tissue sites.
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PMID:Effects of nerve growth factor (NGF) and other fibroblast-derived growth factors on immature human mast cells (HMC-1). 977 35

A new protein with nerve growth promoting activity was purified from the crude venom of the Agkistrodon halys Pallas, a Chinese snake. Its amino-terminal sequence unexpectedly showed high homology with serine proteases, suggesting that it is a new member of the serine protease family. It also cross-reacted with antibodies against thrombin-like enzyme and possessed weak arginine esterase activity, amounting to about 3% of the activity of trypsin. However, its nerve growth promoting activity was comparable to that of nerve growth factor (NGF). It was named NGF-like protease (NLP). Northern blot analysis further demonstrated different patterns of induction of c-myc, vgf and trkA mRNA transcription in PC12 pheochromocytoma cells treated with NGF and NLP, respectively. These data suggested that NLP represents a novel potent neurotrophic factor.
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PMID:Identification of a serine protease with nerve growth promoting activity from snake venom. 985 63

Neuropsin is a novel serine protease, the expression of which is highly localized in the limbic areas of the mouse brain and which is suggested to be involved in kindling epileptogenesis and hippocampal plasticity. The 2.1-A resolution crystal structure of neuropsin provides the first three-dimensional view of one of the serine proteases highly expressed in the nervous system, and reveals a serine protease fold that exhibits chimeric features between trypsin and nerve growth factor-gamma (NGFgamma), a member of the kallikrein family. Neuropsin possesses an N-glycosylated "kallikrein loop" but forms six disulfide bonds corresponding to those of trypsin. The ordered kallikrein loop projects proline toward the active site to restrict smaller residues or proline at the P2 position of substrates. Loop F, which participates in forming the S3/S4 sites, is similar to trypsin rather than NGFgamma. The unique conformations of loops G and H form an S1 pocket specific for both arginine and lysine. These characteristic loop structures forming the substrate-binding site suggest the novel substrate specificity of neuropsin and give a clue to the design of its specific inhibitors.
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PMID:Crystal structure of neuropsin, a hippocampal protease involved in kindling epileptogenesis. 993 20

The gene expression and regulation of nerve growth factor (NGF) in atopic dermatitis (AD) and the human mast cell line (HMC)-1 was investigated at the molecular level. NGF-stimulation of HMC-1 cells resulted in increases in tryptase activity and histamine contents, paralleled by an increase of tryptase and histamine at the transcriptional level. Also, an increased expression of NGF was found in AD lesions, in association with increased systemic NGF plasma levels. Further cutaneous sources for increased NGF levels were keratinocytes and fibroblasts. These findings demonstrate an increased expression of NGF in AD and effects on tryptase and histamine. Mast cells may be major mediators of neurotrophin effects in AD.
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PMID:Gene expression and regulation of nerve growth factor in atopic dermatitis mast cells and the human mast cell line-1. 1574 47


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