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:3.4.25.1 (proteasome)
28,817 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The 20 S proteasome core purified from Saccharomyces cerevisiae is inhibited by reduced glutathione (GSH), cysteine (Cys), or the GSH precursor gamma-glutamylcysteine. Chymotrypsin-like activity was more affected by GSH than trypsin-like activity, whereas the peptidylglutamyl-hydrolyzing activity (caspase-like) was not inhibited by GSH. Cys-sulfenic acid formation in the 20 S core was demonstrated by spectral characterization of the Cys-S(O)-4-nitrobenzo-2-oxa-1,3-diazole adduct, indicating that 20 S proteasome Cys residues might react with reduced sulfhydryls (GSH, Cys, and gamma-glutamylcysteine) through the oxidized Cys-sulfenic acid form. S-Glutahionylation of the 20 S core was demonstrated in vitro by GSH-biotin incorporation and by decreased alkylation with monobromobimane. Compounds such as N-ethylmaleimide (-S-sulfhydril H alkylating), dimedone (-SO sulfenic acid H reactant), or 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole (either -SH or -SOH reactant) highly inhibited proteasomal chymotrypsin-like activity. In vivo experiments revealed that 20 S proteasome extracted from H(2)O(2)-treated cells showed decreased chymotrypsin-like activity accompanied by S-glutathionylation as demonstrated by GSH release from the 20 S core after reduction with NaBH(4). Moreover, cells pretreated with H(2)O(2) showed decreased reductive capacity assessed by determination of the GSH/oxidized glutathione ratio and increased protein carbonyl levels. The present results indicate that at the physiological level the yeast 20 S proteasome is regulated by its sulfhydryl content, thereby coupling intracellular redox signaling to proteasome-mediated proteolysis.
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PMID:20 S proteasome from Saccharomyces cerevisiae is responsive to redox modifications and is S-glutathionylated. 1240 93

The thermal stability of a cysteine-free alkaline protease (Alp) secreted by the eukaryote Aspergillus oryzae was improved both by the introduction of engineered twin disulfide bridges (Cys-69/Cys-101 and Cys-169/Cys-200), newly constructed as part of this study, and by the addition of calcium ions. We performed an extensive kinetic analysis of the increased thermal stability of the mutants as well as the role of calcium dependence. The thermodynamic activation parameters for irreversible thermal inactivation, the activation free energy (deltaG), the activation enthalpy (deltaH), and the activation entropy (deltaS) were determined from absolute reaction rate theory. The values of deltaH and deltaS were significantly and concomitantly increased as a result of introducing the twin disulfide bridges, for which the increase in the value of deltaH outweighed that of deltaS, resulting in significant increases in the value of deltaG. The enhancement of the thermal stability obtained by introducing the twin disulfide bridges is an example of the so-called low-temperature stabilization of enzymes. The stabilizing effect of calcium ions on wild-type Alp is similar to the results we obtained by introducing the engineered twin disulfide bridges.
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PMID:Kinetic analysis of enhanced thermal stability of an alkaline protease with engineered twin disulfide bridges and calcium-dependent stability. 1245 55

The proposed pathway and mechanism of substrate entry and product egress in the hexameric D3 symmetric tricorn protease from Thermoplasma acidophilum were explored by crystallographic studies of ligand complexes and by structure-based mutagenesis. Obstruction of the pore within the 7-bladed beta-propeller (beta7) domain by alkylation or oxidation of an engineered double cysteine mutant strongly decreased enzymatic activities. In line herewith, the crystal structure of the tricorn protease in complex with a trideca-peptide inhibitor modifying the catalytic Ser965 revealed part of the peptide trapped inside the channel of the beta7 domain. The cysteine mutation widening the lumen of the 6-bladed beta-propeller (beta6) domain enhanced catalytic activity, which was restored to normal values after its alkylation. A charge reversal mutant at the putative anchor site of the substrate C terminus, R131E-R132E, drastically reduced the proteolytic activity. The complex crystal structure of a peptide inhibitor with a diketo group at the cleavage site mapped the substrate recognition site and confirmed the role of Arg131-Arg132 as an anchor site. Our results strongly suggest the wider beta7 domain to serve as a selective filter and guide of the substrate to the sequestered active site, while the narrower beta6 domain routes the product to the surface. Moreover, we identified the role of Arg131-Arg132 in anchoring the substrate C terminus.
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PMID:Navigation inside a protease: substrate selection and product exit in the tricorn protease from Thermoplasma acidophilum. 1247 Sep 58

Proteins comprising the first nucleotide-binding- and R-domains of wild-type and Delta F508 cystic fibrosis transmembrane conductance regulator (CFTR) have been synthesised by in vitro transcription/translation. The kinetics and extent of degradation of wild-type and Delta F508 cytoplasmic domain proteins in rabbit reticulocyte lysates, in which proteasome activity was inhibited, were similar, with a half-life of approximately 4h. The results show for the first time, that the benzo(c)quinolizinium compounds, MPB-07 and MPB-91, selectively inhibit degradation of the Delta F508 cytoplasmic domain protein. Studies using protease inhibitors demonstrated that both Delta F508 and wild-type proteins are substrates for cysteine proteases. The studies provide evidence that benzo(c)quinolizinium compounds protect a proteolytic cleavage site by direct binding to the first cytoplasmic domain of Delta F508-CFTR and this is a likely mechanism for increasing Delta F508-CFTR trafficking in intact cells.
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PMID:Benzo(c)quinolizinium drugs inhibit degradation of Delta F508-CFTR cytoplasmic domain. 1250 15

Oxidized low-density lipoproteins (oxLDL) exhibit proinflammatory properties and play a role in atherosclerosis plaque formation, rupture, and subsequent thrombosis. OxLDL alter the activity of the transcription factor NF-kappaB that is involved in the expression of immune and inflammatory genes. In contrast, high-density lipoproteins (HDL) are anti-atherogenic and exhibit anti-inflammatory properties. This work aimed to investigate how oxLDL activate NF-kappaB and whether and how HDL may prevent NF-kappaB activation. In cultured rabbit smooth muscle cells, mitogenic concentrations of mildly oxLDL trigger a rapid and transient NF-kappaB activation, which is strongly inhibited by HDL. Growth factors, phosphatidylinositol 3-kinase/Akt, and sphingosine kinase pathways are not implicated in the oxLDL-induced NF-kappaB activation and are not targets of HDL. OxLDL induce reactive oxygen species (ROS) generation and proteasome activation, which are implicated in NF-kappaB activation, as suggested by the inhibitory effect of the antioxidants N-acetyl-L-cysteine and pyrrolidinedithiocarbamate and the proteasome inhibitor PSI. HDL were able to prevent the intracellular ROS rise triggered by oxLDL or H2O2, thereby inhibiting the subsequent proteasome activation, IkappaB degradation, and NF-kappaB activation. In conclusion, the oxLDL-induced NF-kappaB activation involves ROS generation and proteasome activation, both events being inhibited by HDL. This 'antioxidant' and potentially anti-inflammatory effect of HDL may participate in their general anti-atherogenic properties.
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PMID:HDL counterbalance the proinflammatory effect of oxidized LDL by inhibiting intracellular reactive oxygen species rise, proteasome activation, and subsequent NF-kappaB activation in smooth muscle cells. 1258 48

Iron regulatory protein 2 coordinates cellular regulation of iron metabolism by binding to iron responsive elements in mRNA. The protein is synthesized constitutively but is rapidly degraded when iron stores are replete. This iron-dependent degradation requires the presence of a 73-residue degradation domain, but its functions have not yet been established. We now show that the domain can act as an iron sensor, mediating its own covalent modification. The domain forms an iron-binding site with three cysteine residues located in the middle of the domain. It then reacts with molecular oxygen to generate a reactive oxidizing species at the iron-binding site. One cysteine residue is oxidized to dehydrocysteine and other products. This covalent modification may thus mark the protein molecule for degradation by the proteasome system.
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PMID:Iron regulatory protein 2 as iron sensor. Iron-dependent oxidative modification of cysteine. 1259 20

Novel N-arylsulfonyldipeptidyl aldehyde derivatives were prepared by DMSO oxidation from the corresponding dipeptide alcohol, and their potencies as calpain inhibitors were evaluated in vitro. Among them, N-(4-fluorophenylsulfonyl)-l-valyl-l-leucinal (8, SJA6017) potently inhibited calpains. 8 also inhibited cathepsin B and L but did not inhibit other cysteine proteases (interleukin 1beta-converting enzyme), serine proteases (trypsin, chymotrypsin, thrombin, factor VIIa, factor Xa), or proteasome. Preliminary cytotoxicity studies of 8 exhibited a relatively safe profile.
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PMID:Structure-activity relationship study and drug profile of N-(4-fluorophenylsulfonyl)-L-valyl-L-leucinal (SJA6017) as a potent calpain inhibitor. 1259 66

Thymocytes undergo negative and positive selection during development in the thymus. During this selection process, the majority of thymocytes are eliminated by apoptosis through signaling via TCR or die by neglect, possibly mediated through glucocorticoids. In this study, we report that thymocytes require molecular oxygen to undergo apoptosis induced by dexamethasone (DEX), a synthetic glucocorticoid, and treatment with N-acetyl-L-cysteine (NAC), a thiol antioxidant, inhibits thymocyte apoptosis in vivo as well as ex vivo. We detected elevated intracellular levels of hydrogen peroxide (H(2)O(2)) during DEX-induced apoptosis, which is reduced by NAC treatment, indicating that the elevated levels of intracellular H(2)O(2) are proapoptotic. We also show that loss of mitochondrial membrane potential, cytochrome c release, as well as caspase-3 activation induced by DEX are attenuated by NAC treatment. We identified the production site for H(2)O(2) as the ubiquinone cycle at complex III of mitochondria by using various inhibitors of the mitochondrial electron transport chain, and we show that the cell death events mediated by mitochondria are also significantly reduced when the inhibitors were used. Through inhibition of the proteasome, we also show that the production of H(2)O(2) and the cell death events mediated by mitochondria are regulated by proteosomal activities in DEX-induced thymocyte apoptosis. We conclude that in DEX-treated thymocytes, the increased production of H(2)O(2) originates from mitochondria and is proapoptotic for cell death mediated by mitochondria. We also conclude that all the apoptotic events mediated by mitochondria are regulated by proteasomes.
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PMID:Glucocorticoid-induced apoptosis of thymocytes: requirement of proteasome-dependent mitochondrial activity. 1259 72

Cystathionine beta-synthase (CBS) catalyzes the first of two steps in the transsulfuration pathway that converts homocysteine to cysteine, a precursor of glutathione, a major intracellular antioxidant. Tumor necrosis factor-alpha (TNFalpha), which is known to enhance production of reactive oxygen species, increased CBS activity and glutathione levels in HepG2 cells. Western blot analysis revealed that the higher CBS activity correlated with cleavage of the enzyme to a truncated form. This cleavage was suppressed by inhibitors of superoxide production or by transfection with an expression vector for manganese superoxide dismutase. The commonly used proteasome inhibitors, MG132 and lactacystin but not N-acetyl-Leu-Leu-norleucinal, suppressed the TNFalpha-induced response. Targeted proteolysis of CBS was also observed in livers of mice injected with lipopolysaccharide, which is known to induce TNFalpha. Together, these data reveal a novel and previously unknown mechanism of regulation for homocysteine-linked glutathione homeostasis in cells challenged by oxidative stress.
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PMID:Tumor necrosis factor-alpha-induced targeted proteolysis of cystathionine beta-synthase modulates redox homeostasis. 1261 17

Abnormal proteolysis may be involved in the motor neuron degeneration of amyotrophic lateral sclerosis (ALS). Although several studies of the ubiquitin-proteasome system in ALS have been reported, the endosome-lysosome system has not been investigated in detail. To clarify the association of neurodegeneration with the endosome-lysosome system in ALS, we examined the pathological expression of cysteine proteases such as cathepsins B, H and L and an aspartate protease, cathepsin D, in the anterior horns of 15 ALS cases and 5 controls. In the ALS cases, cathepsin B immunoreactivity was preferentially decreased in the lateral parts of the anterior gray horns compared with the controls. Its immunoreactivity was increased in the cytoplasm of both shrunken and pigmented neurons but was weak in the neurons containing Bunina bodies. In addition, reactive astrocytes were also immunolabeled with cathepsin B. Cathepsin H and cathepsin L were detected in the cytoplasm of a small number of shrunken and pigmented neurons. Cathepsin D immunoreactivity was strong in the cytoplasm of all motor neurons. The immunoreactivity of cathepsins H, L and D was not significantly different between control and ALS cases. Western blot analysis showed that the 25-kDa activated form of cathepsin B was down-regulated in ALS. Our results suggest that cathepsin B is involved in the motor neuron degeneration in ALS.
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PMID:Involvement of cathepsin B in the motor neuron degeneration of amyotrophic lateral sclerosis. 1267 46


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