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:3.4.21.1 (chymotrypsin)
10,938 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Age-related increase in protein oxidation in brain coupled to an impairment of proteasomal activity may underline neuronal loss but differences in susceptibility between species and brain regions remain unexplained. We now investigate differences in proteasomal activity, measured as chymotrypsin-, trypsin- and peptidylglutamyl-like hydrolysing activities between brain regions in rats, mice and common marmosets. In aged rats and mice, proteasomal activity was decreased in the cortex, striatum, cerebellum, globus pallidus and substantia nigra overall when compared to young animals. However, in the aged brain only chymotrypsin-like activity was decreased in the cortex and the globus pallidus while only trypsin-like activity was reduced in the cerebellum. In contrast, in the striatum, both chymotrypsin-like and trypsin-like activities were reduced and in the substantia nigra, all the three catalytic activities of proteasome were significantly impaired. Chymotrypsin-like and trypsin-like activities were significantly higher in all the brain regions of marmosets compared to those of mice and rats. Peptidylglutamyl-like activity was only significantly higher in the cerebellum and striatum of marmoset compared to rodents. The data suggest that there is higher proteasome activity in common marmoset brain compared to rat and mouse and that the basal ganglia are more prone to an age-related decrease in proteasomal activity. This may explain the involvement of altered ubiquitin-proteasome system activity in Parkinson's disease and the relationship to ageing.
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PMID:Proteasomal activity in brain differs between species and brain regions and changes with age. 1588 31

Heat shock proteins (Hsps) with chaperoning function work together with the ubiquitin-proteasome pathway to prevent the accumulation of misfolded, potentially toxic proteins, as well as to control catabolism of the bulk of cytoplasmic, cellular protein. There is evidence for the involvement of both systems in neurodegenerative disease, and a therapeutic target is the heat shock transcription factor, Hsf1, which mediates upregulation of Hsps in response to cellular stress. The mechanisms regulating expression of proteasomal proteins in mammalian cells are less well defined. To assess any direct effect of Hsf1 on expression of proteasomal subunits and activity in mammalian cells, a plasmid encoding a constitutively active form of Hsf1 (Hsf1act) was expressed in mouse embryonic fibroblasts lacking Hsf1 and in cultured human myoblasts. Plasmid encoding an inactivatible form of Hsf1 (Hsf1inact) served as control. In cultures transfected with plasmid hsf1act, robust expression of the major stress-inducible Hsp, Hsp70, occurred but not in cultures transfected with hsf1inact. No significant changes in the level of expression of representative proteasomal proteins (structural [20Salpha], a nonpeptidase beta subunit [20Sbeta3], or 2 regulatory subunits [19S subunit 6b, 11 Salpha]) or in chymotrypsin-, trypsin-, and caspaselike activities of the proteasome were measured. Thus, stress-induced or pharmacological activation of Hsf1 in mammalian cells would upregulate Hsps but not directly affect expression or activity of proteasomes.
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PMID:Proteasome activity or expression is not altered by activation of the heat shock transcription factor Hsf1 in cultured fibroblasts or myoblasts. 1618 68

Although the structure of an enzyme is often depicted as static, it is dynamic. Hence, a population of chemically identical enzymes has not one, but a distribution of structures at any moment in time. Does this have an effect on the activity of the enzyme? This article reviews experiments designed to test the hypothesis that this distribution of structures results in a distribution of enzyme activities. The experiments reviewed here use different enzymes, falvin adenine dinucleotide, beta-galactosidase, alkaline phosphatase, exonuclease I, lactate dehydrogenase I, alpha-chymotrypsin, the 20S proteasome, and horseradish peroxidase. All experiments come to the same conclusion, when measured individually, apparently identical enzymes show a distribution in rates of activity.
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PMID:Diversity in the activity of individual enzymes. 1637 26

The inducible kinin B1 receptor is emerging as an attractive therapeutic target for the treatment of pain and inflammation. Although many studies described its regulation at the transcriptional level, little is known about the maturation of the B1 receptor. Using two human embryonic kidney (HEK) 293 cell lines stably expressing rabbit B1 receptors tagged with the yellow fluorescent protein at the C terminus (B1R-YFP) or the N-terminal myc epitope (myc-B1R), we showed that receptors are mainly retained in a perinuclear compartment and detectable as low-glycosylated species under control conditions. Interference with the ubiquitin-proteasome pathway function (proteasome inhibitors, coexpression with dominant-negative ubiquitin) blocked B1 receptor degradation and amplified its intracellular accumulation. A potent nonpeptide antagonist specifically increased the abundance of highly glycosylated B1R-YFP forms at the cell surface (accessible to chymotrypsin digestion in intact cells); this compound augmented low-glycosylated receptors in brefeldin A-treated cells, supporting the hypothesis that it reaches a newly synthesized receptor in the endoplasmic reticulum. Cell-impermeant peptide or low-affinity nonpeptide B1 receptor antagonists failed to influence the level of highly glycosylated receptors. Chemical chaperones stabilized all B1R-YFP species and up-regulated endogenous B1 receptors expressed at the surface of rabbit smooth muscle cells. Although myc-B1Rs behaved similarly to B1R-YFP in most aspects, antibody-based detection assays failed to reveal highly glycosylated species of this construct. Taken together, these results show that B1 receptors overexpressed in HEK 293 cells are degraded by the proteasome. Furthermore, a pharmacological chaperone highlights the existence of a highly N-glycosylated form of the rabbit kinin B1 receptor at the cell surface.
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PMID:A nonpeptide antagonist reveals a highly glycosylated state of the rabbit kinin B1 receptor. 1640 68

Isoprenoid inhibitors are being evaluated as agents for the treatment of cancer. Their antitumor activity is attributed to inhibition of post-translational modification of Ras, which is crucial for its translocation and attachment to the plasma membrane, and ultimate involvement in signal transduction. However, whether blocking of Ras is solely responsible for the observed antitumor activity is unresolved. In this report, we propose an alternate mechanism. Using breast tumor models, we show that agents possessing a lactone moiety, including statins (such as lovastatin) and the isoprenoid inhibitors (such as FTI-277 and GGTI-298), mediate their cell cycle inhibitory activities by blocking the chymotrypsin activity of the proteasome in vitro. This results in the accumulation of cyclin-dependent kinase inhibitors p21 and p27 with subsequent G(1) arrest. Cells devoid of p21 were refractory to the growth-inhibitory activity of lovastatin, FTI-277, and GGTI-298. However, in these p21 null cells, isoprenylation of key substrates of farnesyl transferase (such as Ras) and of geranylgeranyl transferase (such as RAP-1) were inhibited by FTI-277 and GGTI-298, respectively, suggesting that although both these isoprenoid inhibitors reached and inhibited their intended targets, inhibition of the isoprenylation of Ras and RAP-1A are not sufficient to mediate G(1) arrest. We also show that the cell cycle effects can be attributed to the functional lactone moiety of the aforementioned agents. Collectively, our data suggest that FTI and GGTI and other agents containing an active lactone moiety mediate G(1) arrest via inhibition of the proteasome and up-regulation of p21, independent of the inhibition of isoprenylation of Ras or RAP-1.
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PMID:Farnesyl and geranylgeranyl transferase inhibitors induce G1 arrest by targeting the proteasome. 1642 40

Previously, we showed that ester carbon-containing tea polyphenols, including (-)-epigallocatechin gallate [(-)-EGCG] and (-)-epicatechin-3-gallate [(-)-ECG], potently inhibit proteasomal chymotrypsin-like activity. In addition, our in silico docking study suggested that a particular pose of (-)-EGCG could lead to potential covalent modification of the N-terminal threonine (Thr 1) of the proteasome beta5 subunit in the chymotrypsin-like active site. It has been suggested that some major biotransformation reactions, such as methylation, could result in reduced biological activity of (-)-EGCG in vivo. We hypothesize that methylation reduces binding of (-)-EGCG to the beta5 subunit of the proteasome and, therefore, decreases its proteasomal chymotrypsin-like-inhibitory potency. Here, we report that, while methylation has no effect on nucleophilic susceptibility of (-)-EGCG and (-)-ECG, it may disrupt the ability of these polyphenols to interact with Thr 1 of the proteasome beta5 subunit. In silico docking shows that methylation results in the tea polyphenols' ester carbon being moved away or blocked entirely from Thr 1. Additionally, methylation impairs the ability of (-)-EGCG and (-)-ECG to dock in a consistent low energy pose. These observations, no change in nucleophilic susceptibility, moving or blocking the ester carbon from Thr 1, and lack of a consistent docking pose, suggest that methylation disrupts the ability of (-)-EGCG and (-)-ECG to bind to the proteasome beta5 subunit, which may then diminish their proteasomal chymotrypsin-inhibitory and, therefore, other biological activities.
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PMID:Methylation of green tea polyphenols affects their binding to and inhibitory poses of the proteasome beta5 subunit. 1696 15

Withaferin A (WA) is a steroidal lactone purified from medicinal plant "Indian Winter Cherry" that is widely researched for its variety of properties, including antitumor effects. However, the primary molecular target of WA is unknown. By chemical structure analysis, we hypothesized that Withaferin A might be a natural proteasome inhibitor. Computational modeling studies consistently predict that C1 and C24 of WA are highly susceptible toward a nucleophilic attack by the hydroxyl group of N-terminal threonine of the proteasomal chymotrypsin subunit beta5. Furthermore, WA potently inhibits the chymotrypsin-like activity of a purified rabbit 20S proteasome (IC50=4.5 microM) and 26S proteasome in human prostate cancer cultures (at 5-10 microM) and xenografts (4-8 mg/kg/day). Inhibition of prostate tumor cellular proteasome activity in cultures and in vivo by WA results in accumulation of ubiquitinated proteins and three proteasome target proteins (Bax, p27, and IkappaB-alpha) accompanied by androgen receptor protein suppression (in androgen-dependent LNCaP cells) and apoptosis induction. Treatment of WA under conditions of the aromatic ketone reduction, or reduced form of Celastrol, had significantly decreased the proteasome-inhibitory and apoptosis-inducing activities. Treatment of human prostate PC-3 xenografts with WA for 24 days resulted in 70% inhibition of tumor growth in nude mice, associated with 56% inhibition of the tumor tissue proteasomal chymotrypsinlike activity. Our results demonstrate that the tumor proteasome beta5 subunit is the primary target of WA, and inhibition of the proteasomal chymotrypsin-like activity by WA in vivo is responsible for, or contributes to, the antitumor effect of this ancient medicinal compound.
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PMID:The tumor proteasome is a primary target for the natural anticancer compound Withaferin A isolated from "Indian winter cherry". 2581 35

The beta subunits (beta1, beta2, and beta5) of 20S proteasome and HslV/ClpQ are ATP-dependent threonine proteases present in eukaryotes and prokaryotes, respectively that control levels of key regulatory proteins in the cell. The orthologue of prokaryotic HslV protease in Plasmodium falciparum (PfHslV) is a novel drug target candidate that has no homolog in the human host. In the present study, the PfHslV was expressed, localized and biochemically characterized. The recombinant PfHslV harbored threonine protease specific activity as well as chymotrypsin like and peptidyl glutamyl peptide hydrolase activities. All the three activities could be inhibited by respective specific inhibitors. The protein was localized in the cytosol of the parasite as a soluble protein by Western immunoblotting of parasite fractions and by immuno-fluorescence microscopy. Activity of the protease in the parasite was ascertained by following the degradation of GFP in a transgenic parasite line expressing fusion protein of GFP and Arc-repressor gene, a known target of HslV protease in the prokaryotes. A model structure of PfHslV was constructed based on the crystal structure of Escherichia coli HslV to assess the structural homology. Availability of the structure model of PfHslV may facilitate identification or designing of novel and specific drugs against PfHslV. The in vitro protease assays with recombinant PfHslV and the transgenic parasite line generated in the present study may be exploited in the screening of novel inhibitors to evaluate their anti-malarial activity.
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PMID:Characterization and localization of Plasmodium falciparum homolog of prokaryotic ClpQ/HslV protease. 1727 Feb 90

The proteasome has been successfully targeted for the treatment of multiple myeloma and mantle cell lymphoma; however, in other hematologic malignancies, bortezomib has been less effective as a single agent. Here, we describe effects of NPI-0052, a novel proteasome inhibitor, in leukemia model systems. In cell lines, NPI-0052 inhibits all 3 proteolytic activities associated with the proteasome: chymotrypsin-, trypsin-, and caspase-like. NPI-0052 also induces DNA fragmentation in leukemia lines and in mononuclear cells from a Ph + acute lymphoblastic leukemia (ALL) patient. Caspase-3 activation by NPI-0052 was seen in wild-type Jurkat cells, but was significantly lessened in Fas-associated death domain (FADD)-deficient or caspase-8-deficient counterparts. NPI-0052-induced apoptosis was further probed using caspase-8 inhibitors, which were more protective than caspase-9 inhibitors. N-acetyl cysteine (NAC) also conferred protection against NPI-0052-induced apoptosis, indicating a role for oxidative stress by NPI-0052. In support of the drug's in vitro activities, biweekly treatment with NPI-0052 lessened total white blood cell (WBC) burden over 35 days in leukemic mice. Interestingly, combining NPI-0052 with either MS-275 or valproic acid (VPA) induced greater levels of cell death than the combination of bortezomib with these histone deacetylase inhibitors (HDACi). These effects of NPI-0052, alone and in combination with HDACi, warrant further testing to determine the compound's clinical efficacy in leukemia.
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PMID:NPI-0052, a novel proteasome inhibitor, induces caspase-8 and ROS-dependent apoptosis alone and in combination with HDAC inhibitors in leukemia cells. 1735 34

Pristimerin is a natural product derived from the Celastraceae and Hippocrateaceae families that were used as folk medicines for anti inflammation in ancient times. Although it has been shown that pristimerin induces apoptosis in breast cancer cells, the involved mechanism of action is unknown. The purpose of the current study is to investigate the primary target of pristimerin in human cancer cells, using prostate cancer cells as a working model. Nucleophilic susceptibility and in silico docking studies show that C6 of pristimerin is highly susceptible towards a nucleophilic attack by the hydroxyl group of N-terminal threonine of the proteasomal chymotrypsin subunit. Consistently, pristimerin potently inhibits the chymotrypsin-like activity of a purified rabbit 20S proteasome (IC50 2.2 micromol/L) and human prostate cancer 26S proteasome (IC50 3.0 micromol/L). The accumulation of ubiquitinated proteins and three proteasome target proteins, Bax, p27 and I kappa B-alpha, in androgen receptor (AR)-negative PC-3 prostate cancer cells supports the conclusion that proteasome inhibition by pristimerin is physiologically functional. This observed proteasome inhibition subsequently led to the induction of apoptotic cell death in a dose- and kinetic-dependent manner. Furthermore, in AR-positive, androgen-dependent LNCaP and AR-positive, androgen-independent C4-2B prostate cancer cells, proteasome inhibition by pristimerin results in suppression of AR protein prior to apoptosis. Our data demonstrate, for the first time, that the proteasome is a primary target of pristimerin in prostate cancer cells and inhibition of the proteasomal chymotrypsin-like activity by pristimerin is responsible for its cancer cell death-inducing property.
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PMID:Pristimerin induces apoptosis by targeting the proteasome in prostate cancer cells. 1754 80


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