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

---

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

An inhibitory protein for the 20S proteasome (also known as macropain, the multicatalytic proteinase complex and 20S proteinase) has been purified from bovine red blood cells. The inhibitor has an apparent molecular weight of 31,000 on SDS-PAGE and appears to form multimers under nondenaturing conditions. This protein inhibited all three of the putatively distinct catalytic activities of proteasome A (the active form of the proteinase) characterized by the hydrolysis of synthetic peptides such as Z-VLR-MNA, Z-GGL-AMC or Suc-LLVY-AMC and Z-LLE-beta NA. The inhibitor also prevented the hydrolysis of large protein substrates such as casein, lysozyme and bovine serum albumin. Proteasome L (the latent form of the proteinase) does not degrade these large protein substrates, but does hydrolyze the three synthetic peptides at rates similar to those by proteasome A. The inhibitor inhibited only two of these peptidase activities of proteasome L (hydrolysis of Z-GGL-AMC and of Z-LLE-beta NA or Suc-LLVY-AMC); it had no effect on the hydrolysis of Z-VLR-MNA. The inhibitor was specific for inhibition of the proteasome and had no effect on the activity of any other proteinase tested including trypsin, chymotrypsin, papain, subtilisin and both isoforms of calpain. Kinetic analysis indicates that the inhibitor interacted with the proteasome by a mechanism involving tight-binding. Because the proteasome appears to be a key component of the ATP/ubiquitin-dependent pathway of intracellular protein degradation, the inhibitor may represent an important regulatory protein of this pathway.
...
PMID:Purification and characterization of a protein inhibitor of the 20S proteasome (macropain). 131 59

Proteins conjugated to ubiquitin are degraded by a 26S (1500-kDa) proteolytic complex that, in reticulocyte extracts, can be formed by the association of three factors: CF-1, CF-2, and CF-3. One of these factors, CF-3, has been shown to be the proteasome, a 650-kDa multicatalytic protease complex. We have purified a 250-kDa inhibitor of the proteasome and shown that it corresponds to CF-2. In the presence or absence of ATP, this factor inhibited hydrolysis by the proteasome of both fluorogenic tetrapeptides and protein substrates. When the inhibitor, proteasome, and CF-1 were incubated together in the presence of ATP and Mg2+, degradation of ubiquitin-125I-lysozyme occurred. Both the inhibitory activity and the ability to reconstitute ubiquitin-125I-lysozyme degradation were very labile at 42 degrees C, but both activities were stabilized by ATP or a nonhydrolyzable ATP analog. SDS/PAGE indicated that the 250-kDa inhibitor fraction contained a major subunit of 40 kDa (plus some minor bands). The 125I-labeled inhibitor and purified proteasome formed a complex. When CF-1, ATP, and Mg2+ were also present, the 125I-labeled inhibitor along with the proteasome formed a complex of 1500 kDa. The inhibitor (CF-2) thus appears to be an ATP-binding component that regulates proteolysis within the 1500-kDa complex.
...
PMID:An ATP-stabilized inhibitor of the proteasome is a component of the 1500-kDa ubiquitin conjugate-degrading complex. 131 79

It is known that two types of high-molecular-mass protease complexes are present in the cytosol of mammalian cells; a 20S latent multicatalytic proteinase named the proteasome, and a large proteolytic complex with an apparent sedimentation coefficient of 26S that catalyzes ATP-dependent breakdown of proteins conjugated with ubiquitin. In this work, we first demonstrated that a low concentration of SDS was required for activation of the latent proteasome, whereas the 26S complex degraded substrates for proteasomes in the absence of SDS. Moreover, the 26S complex was greatly stabilized in the presence of 2 mM ATP and 20% glycerol. Based on these characteristics, we next devised a novel procedure for purification of the 26S proteolytic complexes from human kidney. In this procedure, the proteolytic complexes were precipitated from cytoplasmic extracts by ultracentrifugation for 5 h at 105000 x g, and the large 26S complexes were clearly separated from the 20S proteasomes by molecular-sieve chromatography on a Biogel A-1.5 m column. The 26S enzyme was then purified to apparent homogeneity by successive chromatographies on hydroxyapatite and Q Sepharose, then by glycerol density-gradient centrifugation. Electrophoretic and immunochemical analyses showed that the purified human 26S complex consisted of multiple subunits of proteasomes with molecular masses of 21-31 kDa and 13-15 protein components ranging in molecular mass over 35-110 kDa, which were directly associated with the proteasome. The purified 26S proteolytic complex degraded 125I-labeled lysozyme-ubiquitin conjugates in an ATP-dependent manner. The 26S enzyme also showed high ATPase activity, which was copurified with the complex. Vanadate and hemin strongly inhibited not only ATP cleavage, but also ATP-dependent breakdown of ubiquitinligated proteins, suggesting that the 26S complex hydrolyzes ATP and ubiquitinated proteins by closely linked mechanisms. These findings indicate that the 26S complex consists of a proteasome with proteolytic function and multiple other components including an ATPase that regulates energy-dependent, ubiquitin-mediated protein degradation.
...
PMID:Demonstration that a human 26S proteolytic complex consists of a proteasome and multiple associated protein components and hydrolyzes ATP and ubiquitin-ligated proteins by closely linked mechanisms. 131 98

Chemical composition of a substance produced by Klebsiella pneumoniae which is accumulated in chemically defined medium was determined. This substance (the protective factor) protects the bacterium from being killed by normal human serum. Protective factor was treated with DNase, lysozyme and an alkaline protease. The two former enzymes did not affect the protective factor. On the other hand, when alkaline protease was used, the protective activity was totally lost. Results showed that the protective factor is a protein.
...
PMID:[Characterization of a substance which protects Klebsiella pneumoniae from the bactericidal effect of normal human serum]. 134 14

A protein that greatly stimulates the multiple peptidase activities of the 20 S proteasome (also known as macropain, the multicatalytic protease complex, and 20 S protease) has been purified from bovine red blood cells and from bovine heart. The activator protein was a single polypeptide with an apparent molecular weight of 28,000, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and had a native molecular weight of approximately 180,000. This protein, which we have termed PA28, regulated all three of the putatively distinct peptidase activities displayed by each of two functionally different forms of the proteasome. This regulation usually included both an increase in the maximal reaction velocity and a decrease in the concentration of substrate required for half-maximal velocity and indicated that PA28 acted as a positive allosteric effector of the proteasome. PA28 failed, however, to stimulate the hydrolysis of large protein substrates such as casein and lysozyme. These results suggested that the hydrolysis of protein substrates occurred at a site or sites distinct from those that hydrolyzed small peptides and that the regulation of the two processes could be uncoupled. Evidence for direct binding of PA28 to the proteasome was obtained by glycerol density gradient centrifugation. PA28 may play an important regulatory role in intracellular proteolytic pathways mediated by the proteasome.
...
PMID:Identification, purification, and characterization of a protein activator (PA28) of the 20 S proteasome (macropain). 158 32

Haemoglobin damaged by exposure of red blood cells to oxidants is rapidly degraded by a proteolytic pathway which does not require ATP [Fagan, Waxman & Goldberg (1986) J. Biol. Chem. 261, 5705-5713]. By fractionating erythrocyte lysates, we have purified two proteases which hydrolyse oxidatively damaged haemoglobin (Ox-Hb). One protease hydrolysed small fluorogenic substrates in addition to Ox-Hb. Its molecular mass was approximately 700 kDa and it consisted of several subunits ranging in size from 22 to 30 kDa. This enzyme may be related to the high-molecular-mass multicatalytic proteinase previously isolated from a variety of tissue and cell types. The other Ox-Hb-degrading activity had an apparent molecular mass of 400 kDa on gel filtration, a subunit size of 110 kDa and an isoelectric point between 4.5 and 5.0. This protease also hydrolysed the small polypeptides insulin and glucagon, as well as other large proteins such as lysozyme. Insulin blocked the degradation of Ox-Hb and Ox-Hb blocked the hydrolysis of insulin by the purified protease. Thiol reagents and metal chelators strongly inhibited the hydrolysis of both Ox-Hb and insulin, whereas inhibitors of serine, aspartic and thiol proteases had little effect. These properties suggest that the Ox-Hb-degrading activity purified from rabbit erythrocytes is the cytosolic insulin-degrading enzyme that is believed to play a role in the metabolism of insulin in several tissues. We propose that this enzyme may also function as a key component in a cytoplasmic degradative pathway responsible for removing proteins damaged by oxidants.
...
PMID:Purification of a protease in red blood cells that degrades oxidatively damaged haemoglobin. 187 13

Mammalian cells contain two large proteolytic complexes, the 650-kDa proteasome (or multicatalytic protease) and the 1500-kDa (26 S) Ubiquitin-conjugate-degrading enzyme. Since the proteasome is also required for the ATP-dependent degradation of ubiquitinated proteins, we tested whether it may be a component of the larger complex. The proteasome normally is soluble in 38% ammonium sulfate. However, after preincubation of reticulocyte extracts with ATP, several proteasome activities appeared in the 38% ammonium sulfate pellet, including the ability to degrade hydrophobic peptides and 14C-casein. Also, following preincubation with ATP, the precipitable fraction could degrade 125I-lysozyme-ubiquitin (Ub) conjugates. The activities were not present after incubation without ATP or with a nonmetabolizable ATP analog. Nondenaturing gel electrophoresis indicated the ATP-dependent appearance of a new band which degraded proteasome substrates, and reacted with an anti-proteasome monoclonal antibody on Western blot. This new band appeared larger than the proteasome and migrated similarly to the larger Ub-conjugate-degrading complex. The formation of the larger complex required factor(s) present in the 38% ammonium sulfate pellet and either the 40-80% fraction or the purified proteasome from reticulocytes or muscle. After complex formation, hydrolysis of Ub-protein conjugates and also the non-ubiquitinated substrate, casein, was stimulated severalfold by ATP, but non-metabolizable ATP analogs had little or no effect. Thus, the proteasome corresponds to component CF-3 of Ganoth et al. (Ganoth, D., Leshinisky, E., Eytan, E., and Hershkov, A. (1989) J. Biol. Chem. 263 12412-12419) and undergoes an energy-dependent association with other factors to form the 1500-kDa, ATP-requiring proteolytic complex.
...
PMID:The proteasome (multicatalytic protease) is a component of the 1500-kDa proteolytic complex which degrades ubiquitin-conjugated proteins. 218 Sep 50

Eukaryotic cells contain a 700-kDa proteolytic complex (the "proteasome" or multicatalytic endopeptidase complex), whose role in intracellular protein breakdown is unclear. It has been suggested that the proteasome functions in the rapid degradation of oxidant-damaged proteins and in the ATP-dependent proteolytic pathway. To test these possibilities, oxidant-damaged hemoglobin and albumin were produced by treating hemoglobin and albumin with phenylhydrazine, with hydroxyl radicals, or with both hydroxyl and superoxide radicals. After oxidant damage, these proteins were degraded more rapidly in erythrocyte extracts and also by the purified proteasome. However, complete removal of proteasomes from these extracts by immunoprecipitation (or inhibitors of its proteolytic activity) did not reduce the breakdown of oxidant-damaged hemoglobin and decreased degradation of hydroxyl- and superoxide-treated proteins by only 30-40%. Thus, erythrocytes must contain another proteolytic system for degradation of oxidant-damaged proteins. In contrast, immunoprecipitation of proteasomes with polyclonal or monoclonal antibodies prevented the ATP/ubiquitin-dependent degradation of lysozyme and also blocked the ATP-stimulated degradation of ubiquitin-conjugated lysozyme in reticulocyte and skeletal muscle extracts. These data indicate a critical role of the proteasome in the degradation of ubiquitin-conjugated proteins and suggest that the proteasome is associated with or is a component of the larger ubiquitin-conjugate-degrading enzyme complex.
...
PMID:Involvement of the proteasome in various degradative processes in mammalian cells. 253 95

Soluble, cell-free extracts of BHK 21/C13 fibroblasts degraded a variety of exogenous proteins to acid-soluble peptides at pH 8.0. ATP stimulated the rates of proteolysis. Both the absolute rate of proteolysis and the magnitude of the ATP effect depended on the specific substrate. For example, casein was degraded approximately 10-fold faster than lysozyme, but lysozyme degradation was more highly stimulated by ATP than was casein degradation. Ubiquitin enhanced the ATP-stimulated proteolysis of each substrate in both postmicrosomal extracts and DEAE-cellulose fractionated extracts. In each extract, ubiquitin enhanced the ATP-stimulated degradation of lysozyme to a greater degree than that of casein. These results suggested that lysozyme was degraded by a pathway that was more dependent upon ubiquitin than was casein. Further evidence for this conclusion was obtained in studies using substrates whose amino groups were blocked by extensive methylation or carbamoylation. The high molecular weight proteinase, macropain, appears to be involved in the ATP-stimulated degradation of both substrates. Specific immunoprecipitation of macropain with polyclonal antibodies resulted in the inhibition of ATP-stimulated proteinase activity both in the absence and presence of ubiquitin. These results indicate that macropain plays a role in both ubiquitin-mediated and ubiquitin-independent ATP-stimulated proteolysis in BHK cell extracts.
...
PMID:An enzyme related to the high molecular weight multicatalytic proteinase, macropain, participates in a ubiquitin-mediated, ATP-stimulated proteolytic pathway in soluble extracts of BHK 21/C13 fibroblasts. 284 2

We constructed expression vectors for Acremonium chrysogenum using a Fusarium alkaline protease promoter region and tested their potential as secretion systems for foreign proteins using the human (h)-lysozyme gene as an indicator. The gene encoding h-lysozyme was linked to the coding region of (1) the carboxy terminal of the alkaline protease pre peptide, (2) the carboxy terminal of the prepro peptide, (3) three amino acids of the mature protein preceded by the prepro peptide and (4) the carboxy terminal of chicken lysozyme signal peptide, inserted into the genomic DNAs of A. chrysogenum and expressed under the control of the alkaline protease promoter. The transformants of A. chrysogenum with each of these plasmids secreted enzymatically active h-lysozyme. A maximum yield in excess of 40 mg l-1 was obtained when h-lysozyme was linked to the carboxy terminal of alkaline protease prepro peptide. The majority of the amino terminal sequence of the purified h-lysozyme from the culture supernatant was identical with that of authentic h-lysozyme, but it showed some heterogeneity.
...
PMID:Secretion of active human lysozyme by Acremonium chrysogenum using a Fusarium alkaline protease promoter system. 766 38


1 2 3 4 5 6 Next >>

---