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)

Nob1p, which interacts with Nin1p/Rpn12, a subunit of the 19S regulatory particle (RP) of the yeast 26S proteasome, has been identified by two-hybrid screening. NOB1 was found to be an essential gene, encoding a protein of 459 amino acid residues. Nob1p was detected in growing cells but not in cells in the stationary phase. During the transition to the stationary phase, Nob1p was degraded, at least in part, by the 26S proteasome. Nob1p was found only in proteasomal fractions in a glycerol gradient centrifugation profile and immuno-coprecipitated with Rpt1, which is an ATPase component of the yeast proteasomes. These results suggest that association of Nob1p with the proteasomes is essential for the function of the proteasomes in growing cells.
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PMID:Nob1p, a new essential protein, associates with the 26S proteasome of growing saccharomyces cerevisiae cells. 1067 11

Two proteins were isolated, in a stable form, from bovine brain by ion exchange chromatography, gel filtration and ultracentrifugation on glycerol gradient. They were identified as 20S and 26S proteasomes on the basis of molecular mass, migration velocity on non-denaturing gels, immunoreactivity, multipeptidase activity and the 26S proteasome also for dependence on ATP for the degradation of short peptides and ubiquitinylated proteins. However, the 26S proteasome has some properties not yet described for its counterpart of other tissues and from brain of this and other species. In particular, the ATP concentration required by the 26S proteasome to reach maximal peptidase activity was approximately 40-fold lower than the one required for maximal proteolytic activity on polyubiquitinylated substrates. Moreover, plots of substrate concentration vs. velocity gave a saturation curve for the 26S proteasome only, which, for the trypsin-like and post-glutamyl peptide hydrolase activities fitted the Michaelis-Menten equation, whereas for the chymotrypsin-like activity indicated multibinding site kinetics with positive cooperativity (n = 2.32+/-0.38). As concerns the 20S proteasome, its electrophoretic pattern on native gel revealed a single protein band, a feature, to our knowledge, not yet described for the brain particle of any species.
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PMID:Structural and functional characterization of 20S and 26S proteasomes from bovine brain. 1071 20

We have identified a 26S proteasome-associated ubiquitin carboxyl-terminal hydrolase (UCH) in Schizosaccharomyces pombe. The gene (designated uch2+) encodes a protein containing a UCH catalytic domain at its N-terminus and a short extension at its C-terminus. uch2+ is nonessential as the uch2 null mutant strain showed no significant difference from the wild-type strain. The GFP-tagged Uch2p is localized predominantly to the nuclear periphery, which is similar to the 26S proteasome localization. Deletion of the C-terminal extension of Uch2p resulted in a drastic change of its subcellular localization: it showed a generally diffused distribution instead of a perinuclear pattern. Glycerol gradient centrifugation analysis and coimmunoprecipitation studies of fission yeast extracts using anti-Mts4p antiserum suggest that Uch2p is associated with the 26S proteasome and the association of Uch2p with the 26S proteasome is mediated by its C-terminal extension.
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PMID:Identification of a 26S proteasome-associated UCH in fission yeast. 1087 38

A major metabolic effect of insulin is inhibition of cellular proteolysis, but the proteolytic systems involved are unclear. Tissues have multiple proteolytic systems, including the ATP- and ubiquitin-dependent proteasome pathway. The effect of insulin on this pathway was examined in vitro and in cultured cells. Insulin inhibited ATP- and ubiquitin-dependent lysozyme degradation more than 90% by reticulocyte extract, in a dose-dependent manner (IC50 approximately 50 nM). Insulin did not reduce the conjugation of ubiquitin to lysozyme and was not itself ubiquitin-conjugated. In HepG2 cells, insulin increased ubiquitin-conjugate accumulation 80%. The association between the 26S proteasome and an intracellular protease, the insulin-degrading enzyme (IDE), was examined by a purification scheme designed to enrich for the 26S proteasome. Copurification of IDE activity and immunoreactivity with the proteasome were detected through several chromatographic steps. Glycerol gradient analysis revealed cosedimentation of IDE with the 20S proteasome and possibly with the 26S proteasome. The proteasome-associated IDE was displaced when the samples were treated with insulin. These results suggest that insulin regulates protein catabolism, at least in part, by decreasing ubiquitin-mediated proteasomal activity, and provides a new target for insulin action. The displacement of IDE from the proteasome provides a mechanism for this insulin action.
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PMID:Insulin inhibits the ubiquitin-dependent degrading activity of the 26S proteasome. 1087 52

We have previously cloned a cDNA encoding TBP-1, a protein present in the rat spermatid manchette and outer dense fibers of the developing sperm. TBP-1 contains a heptad repeat of six-leucine zipper fingers at the amino terminus and highly conserved ATPase and DNA/RNA helicase motifs toward the carboxyl terminus. TBP-1 is one of the 20 subunits forming the 19S regulatory complex of the 26S proteasome, an ATP-dependent multisubunit protease found in most eukaryotic cells. We now report the isolation of the 26S proteasome from rat testis and sperm tail and its visualization by whole-mount electron microscopy using negative staining. The 26S proteasome from rat testis was fractionated by Sephacryl S-400/Mono-Q chromatography using homogenates suspended in a 10% glycerol-supplemented buffer. Chromatographic fractions were analyzed by immunoblotting using a specific anti-TBP-1 serum. During the purification of Sak57, a keratin filament present in outer dense fibers from epididymal sperm, we detected a substantial amount of 26S proteasomes. Intact 26S proteasomes from rat testis display a rod-shaped particles about 45 nm in length and 11-17 nm in diameter. Each particle consists of a 20S barrel-shaped component formed by four rings (alphabetabetaalpha), capped by two polar 19S regulatory complexes, each identified by an element known as the "Chinese dragon head motif". TBP-1 is an ATPase-containing subunit of the 19S regulatory cap. Rat sperm preparations displayed both dissociated 26S proteasomes and Sak57 filaments. We hypothesize that 26S proteasomes in the perinuclear-arranged manchette are in a suitable location for recognition, sequestration, and degradation of accumulating ubiquitin-conjugated somatic and transient testis-specific histones during spermiogenesis. In the sperm tail, the 26S proteasome may have a role in the remodeling of the outer dense fibers and other tail components during epididymal transit.
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PMID:Structural features of the 26S proteasome complex isolated from rat testis and sperm tail. 1098 18

The proteasome is an eukaryotic multi-subunit protease complex composed of one 20S core component and two 19S regulatory complexes. The regulatory complex contains 6 putative ATPases. We investigated tissue and cell distribution of one of these ATPases, MSS1 (mammalian suppressor of sgv1). MSS1 was ubiquitously present in rat tissues as was the 20S core component of proteasome. However, the ratio of MSS1 to 20S varied greatly among tissues and MSS1 was concentrated in the thymus. Glycerol gradient sedimentation analysis revealed that MSS1 is included in protein complexes whose density is lighter than that of the proteasome. MSS1 was distributed in mammalian cells ubiquitously, while proteasome was rather concentrated in the nuclei. Hence, a novel molecular status of MSS1 distinct from proteasome is implicated. Interestingly, multiple basal transcription factors for RNA polymerase II, including TBP, TFIIB, TFIIH, and TFIIF, were found to be associated with MSS1. These results suggest that MSS1, in addition to proteolysis, plays a role in DNA metabolism including transcriptional regulation.
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PMID:Tissue and cell distribution of a mammalian proteasomal ATPase, MSS1, and its complex formation with the basal transcription factors. 1111 27

Tonicity-responsive genes are regulated by the TonE enhancer element and the tonicity-responsive enhancer binding protein (TonEBP) transcription factor with which it interacts. Urea, a permeant solute coexistent with hypertonic NaCl in the mammalian renal medulla, activates a characteristic set of signaling events that may serve to counteract the effects of NaCl in some contexts. Urea inhibited the ability of hypertonic stressors to increase expression of TonEBP mRNA and also inhibited tonicity-inducible TonE-dependent reporter gene activity. The permeant solute glycerol failed to reproduce these effects, as did cell activators including peptide mitogens and phorbol ester. The inhibitory effect of urea was evident as late as 2 h after the application of hypertonicity. Pharmacological inhibitors of known urea-inducible signaling pathways failed to abolish the inhibitory effect of urea. TonEBP action is incompletely understood, but evidence supports a role for proteasome function and p38 action in regulation; urea failed to inhibit proteasome function or p38 signaling in response to hypertonicity. Consistent with its effect on TonEBP expression and action, urea pretreatment inhibited the effect of hypertonicity on expression of the physiological effector gene, aldose reductase. Taken together, these data 1) define a molecular mechanism of urea-mediated inhibition of tonicity-dependent signaling, and 2) underscore a role for TonEBP abundance in regulating TonE-mediated gene transcription.
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PMID:Urea inhibits hypertonicity-inducible TonEBP expression and action. 1129 34

Although proteasomes are mainly located in the cytosol, it is known that significant amounts are also associated with endoplasmic reticulum (ER) membranes where they may play a role in the degradation of specific ER membrane proteins. The present studies were undertaken to compare ER and cytosolic proteasomal activities in WB rat liver cells. N-Heptyl-beta-thioglucopyranoside (HTG) extracts of membrane or cytosol fractions were chromatographed in glycerol/ATP buffers on size-exclusion and ion-exchange columns and the elution profiles of proteasomal peptidase activity and immunoreactive components of the 20S complex, 19S complex, and PA28 were compared. Cytosol fractions showed a single peak of chymotrypsin-like peptidase activity (Cht-L), which was inhibited completely by 5 microM lactacystin (LC) or SDS (0.03%) and corresponded to 26S proteasomes based upon the presence of both 20S and 19S components. By comparison, membrane fractions contained two major peaks of Cht-L activity. The first peak shared the same properties as the peak activity observed in cytosol fractions. However, the second peak was stimulated by SDS and was LC-insensitive (5 microM) and contained trypsin-like (T-L) and peptide-glutamyl peptidase (PGPH) but no cathepsin or calcium-activated protease activities. PA28 activator protein was present in both membrane and cytosol fractions. Thus, the principal difference between cytosolic and membrane activity was that the latter fractions contained a novel membrane-associated LC-insensitive protease(s) catalyzing three of the major peptidase activities of the proteasome.
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PMID:Characterization of membrane-associated proteasomes in WB rat liver epithelial cells. 1136 Oct 31

Loss of skeletal muscle is a major factor in the poor survival of patients with cancer cachexia. This study examines the mechanism of catabolism of skeletal muscle by a tumour product, proteolysis-inducing factor (PIF). Intravenous administration of PIF to normal mice produced a rapid decrease in body weight (1.55 +/- 0.12 g in 24 h) that was accompanied by increased mRNA levels for ubiquitin, the Mr 14 000 ubiquitin carrier-protein, E2, and the C9 proteasome subunit in gastrocnemius muscle. There was also increased protein levels of the 20S proteasome core and 19S regulatory subunit, detectable by immunoblotting, suggesting activation of the ATP-ubiquitin-dependent proteolytic pathway. An increased protein catabolism was also seen in C(2)C(12)myoblasts within 24 h of PIF addition with a bell-shaped dose-response curve and a maximal effect at 2-4 nM. The enhanced protein degradation was attenuated by anti-PIF antibody and by the proteasome inhibitors MG115 and lactacystin. Glycerol gradient analysis of proteasomes from PIF-treated cells showed an elevation in chymotrypsin-like activity, while Western analysis showed a dose-related increase in expression of MSSI, an ATPase that is a regulatory subunit of the proteasome, with a dose-response curve similar to that for protein degradation. These results confirm that PIF acts directly to stimulate the proteasome pathway in muscle cells and may play a pivotal role in protein catabolism in cancer cachexia.
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PMID:Activation of ATP-ubiquitin-dependent proteolysis in skeletal muscle in vivo and murine myoblasts in vitro by a proteolysis-inducing factor (PIF). 1146 Oct 93

Prolonged treatment of rat adipocytes with TNF alpha increases lipolysis through a mechanism mediated, in part, by down-regulation of inhibitory G proteins (G(i)). Separately, down-regulation of G(i) by prolonged treatment with an A(1)-adenosine receptor agonist, N(6)-phenylisopropyl adenosine (PIA) increases lipolysis. To investigate the role of proteolysis in TNF alpha and PIA-mediated G(i) down-regulation and stimulation of lipolysis, we used the protease inhibitors lactacystin (proteasome inhibitor) and calpeptin (calpain inhibitor). Rat adipocytes were preincubated for 1 h with lactacystin (10 microM) or calpeptin (50 microM), before 30-h treatment with either TNF alpha (50 ng/ml) or PIA (300 nM). We then measured lipolysis (glycerol release), abundance of alpha-subunits of G(i)1 and G(i)2 in plasma membranes (Western blotting) and protease activities (in specific fluorogenic assays). TNF alpha and PIA stimulated lipolysis approximately 2-fold and caused G(i) down-regulation. Although neither lactacystin nor calpeptin affected basal lipolysis, lactacystin completely inhibited both TNF alpha and PIA-stimulated lipolysis (the 50% inhibitory concentration was approximately 2 microM), whereas calpeptin had no effect. Similarly, lactacystin but not calpeptin blocked both PIA and TNF alpha-induced G(i) down-regulation. These findings provide further evidence that the chronic lipolytic effect of TNF alpha and PIA is secondary to G(i) down-regulation and suggest that the mechanism involves proteolytic degradation mediated through the proteasome pathway.
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PMID:Inhibition of proteasome activity blocks the ability of TNF alpha to down-regulate G(i) proteins and stimulate lipolysis. 1171 99


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