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 bacterial HslVU ATP-dependent protease is a homolog of the eukaryotic 26 S proteasome. HslU ATPase forms a hexameric ring, and HslV peptidase is a dodecamer consisting of two stacked hexameric rings. In HslVU complex, the HslU and HslV central pores are aligned, and the proteolytic active sites are sequestered in an internal chamber of HslV, with access to this chamber restricted to small axial pores. Here we show that the C-terminal tails of HslU play a critical role in the interaction with and activation of HslV peptidase. A synthetic tail peptide of 10 amino acids could replace HslU in supporting the HslV-mediated hydrolysis of unfolded polypeptide substrates such as alpha-casein, as well as of small peptides, suggesting that the HslU C terminus is involved in the opening of the HslV pore for substrate entry. Moreover, deletion of 7 amino acids from the C terminus prevented the ability of HslU to form an HslVU complex with HslV. In addition, deletion of the C-terminal 10 residues prevented the formation of an HslU hexamer, indicating that the C terminus is required for HslU oligomerization. These results suggest that the HslU C-terminal tails act as a molecular switch for the assembly of HslVU complex and the activation of HslV peptidase.
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PMID:The C-terminal tails of HslU ATPase act as a molecular switch for activation of HslV peptidase. 1201 Oct 53

Both exercise and insulin-like growth factor I (IGF-I) are known to have major hypertrophic effects in skeletal muscle; however, the interactive effect of exogenous IGF-I and exercise on muscle protein turnover or the ubiquitin-proteasome pathway has not been reported. In the present study, we have examined the interaction between endurance exercise training and IGF-I treatment on muscle protein turnover and the ubiquitin-proteasome pathway in the postexercise period. Adult male rats (270-280 g) were randomized to receive 5 consecutive days of progressive treadmill exercise and/or IGF-I treatment (1 mg. kg body wt(-1). day(-1)). Twenty-four hours after the last bout of exercise, the rate of protein breakdown in incubated muscles was significantly reduced compared with that in unexercised rats. This was associated with a significant reduction in the chymotrypsin-like activity of the proteasome and the rate of ubiquitin-proteasome-dependent casein hydrolysis in muscle extracts from exercised compared with unexercised rats. In contrast, the muscle expression of the 20S proteasome subunit beta-1, ubiquitin, and the 14-kDa E2 ubiquitin-conjugating enzyme was not altered by exercise or IGF-I treatment 24 h postexercise. Exercise had no effect on the rates of total mixed muscle protein synthesis in incubated muscles 24 h postexercise. IGF-I treatment had no effect on muscle weights or the rates of protein turnover 24 h after endurance exercise. These results suggest that a suppression of the ubiquitin-proteasome proteolytic pathway after endurance exercise may contribute to the acute postexercise net protein gain.
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PMID:IGF-I has no effect on postexercise suppression of the ubiquitin-proteasome system in rat skeletal muscle. 1201 37

HslVU is a bacterial homolog of the proteasome, where HslV is the protease that is activated by HslU, an ATPase and chaperone. Structures of singly and doubly capped HslVU particles have been reported, and different binding modes have been observed. Even among HslVU structures with I-domains distal to HslV, no consensus mode of activation has emerged. A feature in the Haemophilus influenzae HslVU structure, insertion of the C termini of HslU into pockets in HslV, was not seen in all other structures of the enzyme. Here we report site-directed mutagenesis, peptide activation, and fluorescence experiments that strongly support the functional relevance of the C terminus insertion mechanism: we find that mutations in HslV that disrupt the interaction with the C termini of HslU invariably lead to inactive enzyme. Conversely, synthetic peptides derived from the C terminus of HslU bind to HslV with 10(-5) M affinity and can functionally replace full HslU particles for both peptide and casein degradation but fail to support degradation of a folded substrate. Thus, the data can be taken as evidence for separate substrate unfoldase and protease stimulation activities in HslU. Enhanced HslV proteolysis could be due to the opening of a gated channel or allosteric activation of the active sites. To distinguish between these possibilities, we have mutated a series of residues that line the entrance channel into the HslV particle. Our mutational and fluorescence experiments demonstrate that allosteric activation of the catalytic sites is required in HslV, but they do not exclude the possibility of channel opening taking place as well. The present data support the conclusion that the H. influenzae structure with I-domains distal to HslV captures the active species and point to significant differences in the activation mechanism of HslV, ClpP, and the proteasome.
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PMID:Functional interactions of HslV (ClpQ) with the ATPase HslU (ClpY). 1203 94

The potential of a protein-engineered His tag to immobilize macromolecules in a predictable orientation at metal-chelating lipid interfaces was investigated using recombinant 20 S proteasomes His-tagged in various positions. Electron micrographs demonstrated that the orientation of proteasomes bound to chelating lipid films could be controlled via the location of their His tags: proteasomes His-tagged at their sides displayed exclusively side-on views, while proteasomes His-tagged at their ends displayed exclusively end-on views. The activity of proteasomes immobilized at chelating lipid interfaces was well preserved. In solution, His-tagged proteasomes hydrolyzed casein at rates comparable with wild-type proteasomes, unless the His tags were located in the vicinity of the N termini of alpha-subunits. The N termini of alpha-subunits might partly occlude the entrance channel in alpha-rings through which substrates enter the proteasome for subsequent degradation. A combination of electron micrographs and atomic force microscope topographs revealed a propensity of vertically oriented proteasomes to crystallize in two dimensions on fluid lipid films. The oriented immobilization of His-tagged proteins at biocompatible lipid interfaces will assist structural studies as well as the investigation of biomolecular interaction via a wide variety of surface-sensitive techniques including single-molecule analysis.
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PMID:Specific orientation and two-dimensional crystallization of the proteasome at metal-chelating lipid interfaces. 1211 6

Under microgravity conditions similar to those in space, it is known that various nutritional and physiological changes in the body are induced. Especially in the aspect of nutrition, muscle atrophy is a characteristic phenomenon accompanying weightlessness. This study was conducted to investigate the ameliorated effect of muscle atrophy caused by suspension hypokinesia by using the soy protein isolate (SPI) as the protein source in comparison with casein. Male Wistar strain rats (8 wk old) were divided into two groups, each suspended with a suspension harness, and fed on a 20% SPI diet or a 20% casein diet for 10 d. The body weights of the suspended rats fed casein or SPI decreased similarly. The weight of the gastrocnemius and soleus muscle were decreased by suspension hypokinesia; however, the degree of the decrease of the muscle weights, especially soleus muscles, of rats fed the SPI diet was smaller than that of rats fed the casein diet. Serum Ntau-methylhistidine concentration was significantly lower in rats fed the SPI diet than in rats fed the casein diet. Similarly, the activities of muscle protein-degrading enzymes such as calpain and proteasome were significantly lower in rats fed the SPI diet than in rats fed the casein diet. Cathepsin B+L activities were not affected by the SPI or the casein diet. Therefore it is suggested that SPI caused a reduction of the proteolysis of myofibrillar protein in skeletal muscles through a reduction of calpain and proteasome activities, in consequence to ameliorate the muscle atrophy.
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PMID:Effect of different dietary protein composition on skeletal muscle atrophy by suspension hypokinesia/hypodynamia in rats. 1217 31

The effect of oxidative stress induced by neurotoxic metal ions on the properties of the brain 20S proteasome or multicatalytic proteinase complex (MPC) has been studied. Exposure of the 20S proteasome to increasing amounts of Fe(III), Fe(II), Cu(II) or Zn(II) affects its main hydrolytic activities: trypsin-like (T-L), chymotrypsin-like (ChT-L), peptidylglutamyl-peptide hydrolase (PGPH), branched-chain amino acid preferring (BrAAP) and caseinolytic activities, although in different ways. T-L activity showed gradual activation by both iron ions but inhibition by Cu(II) and Zn(II). ChT-L and PGPH activities were inhibited whereas BrAAP activity was widely activated by all the tested metal salts except for zinc ions. Moreover, the exposure to ferrous salt increased the degradation rate of casein. The functional effects appear to be linked to oxidation-induced modifications, as demonstrated by an increase of carbonyl groups following the exposure to metal ions. In addition, modifications induced by ferrous salt on the catalytic subunits were also supported by western blot analyses performed using anti-X, anti-Y and anti-Z antibodies. The results obtained clearly indicate that metal-catalyzed oxidation strongly affects the functions of the brain 20S proteasome, even though the catalytic subunits seem to be differently influenced by oxidative phenomena.
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PMID:Effect of neurotoxic metal ions on the proteolytic activities of the 20S proteasome from bovine brain. 1220 11

A digestive protease from Spilosoma obliqua (Lepidoptera: Arctiidae) fifth instar larval guts was purified and characterized. The protease was purified using ammonium sulfate fractionation, ion-exchange chromatography, and hemoglobin-sepharose affinity chromatography. The purification procedure resulted in a 37-fold increase in the specific activity of the protease. Protease thus obtained was found to be electrophoretically pure under native and denaturing conditions. The purified protease had a molecular mass of 90 kDa as determined by gel filtration, and a pH optimum of 11.0. The purified protease optimally hydrolyzed casein at 50 degrees C. A Km of 2 x10(-6) M was obtained using BApNA as a substrate for the purified alkaline protease. The ability of S. obliqua protease and bovine trypsin to hydrolyze various synthetic substrates (BApNA, BAEE, and BAME), and the inhibition patterns of S. obliqua and bovine trypsin with "classical" trypsin inhibitors are also reported.
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PMID:Purification and characterization of a digestive alkaline protease from the larvae of Spilosoma obliqua. 1221 Sep 56

The proteasomal pathway is responsible for processes essential for cell viability, including the selective degradation of oxidized proteins. An age-dependent loss in proteasome function has been reported in many tissues, but has not been examined in the retina. In this study, we evaluated proteasome function and protein oxidation in retinal homogenates from young adult and old F344BN rats. For retinal proteasome from old rats, we observed an 80% decrease in the rate of casein degradation and a 75% loss in chymotrypsin-like activity. This loss in activity could be partially accounted for by a 50% reduction in expression of the 20S proteasome. The regulatory complex PA700 and the inducible beta-subunit, LMP7, which is associated with the chymotrypsin-like activity, were expressed in equivalent concentrations relative to the 20S catalytic core in both young and old rats. Immunochemical analysis using antibodies that recognize the protein oxidative modifications, nitrotyrosine and 4-hydroxy-2-nonenal, showed that retinal proteins from old rats exhibited the greatest immunoreactivity. These results suggest that the age-related loss in proteasome function contributes to the accumulation of oxidized retinal proteins. Thus, the combined effect of an increase in oxidized proteins and inactivation of the protease responsible for ridding the cell of oxidized proteins places the aged retina at greater risk for irreversible damage caused by oxidative stress.
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PMID:Proteasome function and protein oxidation in the aged retina. 1238 90

Protein phosphorylation has a key role in modulating the stabilities of circadian clock proteins in a manner specific to the time of day. A conserved feature of animal clocks is that Period (Per) proteins undergo daily rhythms in phosphorylation and levels, events that are crucial for normal clock progression. Casein kinase Iepsilon (CKIepsilon) has a prominent role in regulating the phosphorylation and abundance of Per proteins in animals. This was first shown in Drosophila with the characterization of Doubletime (Dbt), a homologue of vertebrate casein kinase Iepsilon. However, it is not clear how Dbt regulates the levels of Per. Here we show, using a cell culture system, that Dbt promotes the progressive phosphorylation of Per, leading to the rapid degradation of hyperphosphorylated isoforms by the ubiquitin-proteasome pathway. Slimb, an F-box/WD40-repeat protein functioning in the ubiquitin-proteasome pathway interacts preferentially with phosphorylated Per and stimulates its degradation. Overexpression of slimb or expression in clock cells of a dominant-negative version of slimb disrupts normal rhythmic activity in flies. Our findings suggest that hyperphosphorylated Per is targeted to the proteasome by interactions with Slimb.
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PMID:Role for Slimb in the degradation of Drosophila Period protein phosphorylated by Doubletime. 1244 74

The production of alkaline protease of Aspergillus oryzae U1521 was examined in liquid culture. In a culture of defatted soybean only, it gave satisfactory enzyme yields at 584,000 U/g defatted soybean. When various carbohydrates were supplemented, enzyme production was significantly increased. An increase in production by lactose was the most marked. Enrichment with casitone or casein increased productivity, but not cornsteep solid. Media formulation (g/L) of defatted soybean 10, lactose 5, casitone 1, and KH(2)PO(4) 5 enhanced alkaline protease production by A. oryzae U1521 to a maximum of 1,410,000 U/g defatted soybean. Scaling-up experiments indicated the flask-scale results could be reproduced at 40 g of substrate in 5-L fermenter. The enzyme activity was maximum between pH 8-9 and at a temperature of 45 degrees C.
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PMID:Production of alkaline protease by a genetically engineered Aspergillus oryzae U1521. 1250 79


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