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 effects were assessed of high hydrostatic pressure on the activity and structure of rabbit skeletal muscle proteasome. The pressure effects on the activity were measured by the amount of fluorometric products released from synthetic substrates under pressure and from fluorescein isothiocyanate (FITC)-labeled casein after releasing the pressure. The effects on the structure were measured by fluorescene spectroscopy under pressure, and by circular dichroism (CD) spectroscopy and surface hydrophobicity after releasing the pressure. The optimal pressure for the hydrolyzing activity of synthetic peptides was 50 MPa. The degradation of FITC-labeled casein increased linearly with increasing pressure applied up to 200 MPa, and then markedly decreased up to at 400 MPa. The changes in the tertiary structure detected by fluorometric measurement were irreversible, whereas the changes in the secondary structure were small compared with those by heat treatment. The pressure-induced activation of proteasome therefore seems to have been due to a little unfolding of the active sites of proteasome.
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PMID:Effects of a high-pressure treatment on the activity and structure of rabbit muscle proteasome. 1604 Nov 25

An alkaline protease was purified to apparent homogeneity from culture supernatants of Bacillus sp. PS719, a novel alkaliphilic, thermophilic bacterium isolated from a thermal spring soil sample, by ammonium sulfate precipitation followed by DEAE-cellulose and alpha-casein agarose column chromatographies. The purified enzyme migrated as a single protein band of 42 kDa during both denaturing and nondenaturing gel electrophoresis, suggesting that it consists of a single polypeptide chain. Its isoelectric point was approximately 4.8. The protease exhibited maximum activity towards azocasein at pH 9.0 and at 75 degrees C. The enzyme activity was stimulated by Ca2+, but was inhibited in the presence of Fe2+ or Cu2+. The enzyme was stable in the pH range 8.0 to 10.0 and up to 80 degrees C in the absence of Ca2+. Since phenylmethylsulfonyl fluoride (PMSF) and 3,4-dichloroisocoumarin (DCI) in addition to N-alpha-p-tosyl-L-lysine chloromethyl ketone (TLCK) completely inhibited the activity, this enzyme appears to be a trypsin-like serine protease. Among the various oligopeptidyl-p-nitroanilides tested, the protease showed a preference for cleavage at arginine residues on the carboxylic side of the scissile bond of the substrate, liberating p-nitroaniline from N-carbobenzoxy (CBZ)-L-arginine-p-nitroanilide with the K(m) and V(max) values of 0.6 mM and 1.0 micromol.min(-1).mg protein(-1), respectively.
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PMID:Purification and characterization of an extracellular protease from alkaliphilic and thermophilic Bacillus sp. PS719. 1623 22

Pyrococcus furiosus was shown to grow on casein or peptides as the sole carbon, energy, and nitrogen sources, while maltose could be used as a carbon and energy source only if peptides were present in the medium. A mixture of all 20 single amino acids could not replace the peptide requirement. Specific intracellular proteolytic activity was induced under low casein or tryptone levels and was decreased by the addition of maltose to both peptide-limiting and peptide-rich media in batch and continuous cultures. In a peptide-limited chemostat, activity towards azocasein and MeO-Suc-Arg-Pro-Tyr-p-nitroanilide reached a maximum at a dilution rate of 0.28 h, while activity toward l-lysine-p-nitroanilide reached a maximum at 0.50 h. Under peptide-limiting conditions, levels of the 66-kDa protease (S66) were enhanced relative to those of other cell proteins. Preliminary evidence suggests that this protease is immunologically related to the eukaryotic multicatalytic proteinase complex (proteosome).
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PMID:Regulation of Proteolytic Activity in the Hyperthermophile Pyrococcus furiosus. 1634 84

An extracellular alkaline protease from an alkalophilic bacterium, Bacillus cereus, was produced in a large amount by the method of extractive fermentation. The protease is thermostable, pH tolerant, and compatible with commercial laundry detergents. The protease purified and characterized in this study was found to be superior to endogenous protease already present in commercial laundry detergents. The enzyme was purified to homogeneity by ammonium sulfate precipitation, concentration by ultrafiltration, anion-exchange chromatography, and gel filtration. The purified enzyme had a specific activity of 3256.05 U/mg and was found to be a monomeric protein with a molecular mass of 28 and 31 kDa, as estimated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and nondenaturing PAGE, respectively. Its maximum protease activity against casein was found to be at pH 10.5 and 50 degrees C. Proteolytic activity of the enzyme was detected by casein and gelatin zymography, which gave a very clear protease activity zone on gel that corresponded to the band obtained on SDS-PAGE and nondenaturing PAGE with a molecular mass of nearly 31 kDa. The purified enzyme was analyzed through matrix-assisted laser desorption ionization-time-of-flight-mass spectrometry (MALDI-TOF-MS) and identified as a subtilisin class of protease. Specific serine protease inhibitors, suggesting the presence of serine residues at the active site, inhibited the enzyme significantly.
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PMID:Purification and characterization of Bacillus cereus protease suitable for detergent industry. 1637 46

An alkaline serine protease that hydrolyzes soybean protein into strong angiotensin I-converting enzyme inhibitory hydrolysates was isolated from alkalophilic Bacillus sp. SS103 and purified. The enzyme was purified by ammonium sulfate precipitation followed by gel filtration, cationic exchange column chromatography, and anionic exchange column chromatography. When run on sodium dodecyl sulfate-polyacrylamide gel electrophoresis and isoelectric focusing gel, the purified enzyme gave a 36-kDa band and pI 5.5, respectively. The enzyme showed maximum activity at pH 11.0 and 50 degrees C. This enzyme activity was highly inhibited by aprotinin, suggesting it belongs to the serine protease class of enzymes. The K (m) and V (max) of the enzyme, when casein was used for the substrate, were 9.7 x 10-4 mM and 244 microg/minute, respectively. From the results of this study, it is concluded that the purified alkaline protease isolated from Bacillus sp. SS103 should be further studied for production of biofunctional hydrolysates.
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PMID:Purification and characterization of an alkaline serine protease producing angiotensin I-converting enzyme inhibitory peptide from Bacillus sp. SS103. 1637 56

Optimization of the fermentation medium for maximum alkaline protease production was carried out with a new strain of Pseudomonas aeruginosa (B-2). Replacing the protein source/inducer (albumin in place of casein) brought about significant increase in yield after 48 hr of inoculation. Three most effective medium constituents identified by initial screening method of Plackett-Burman were albumin, (NH4)2SO4 and glucose. Central Composite Design (CCD) and Response Surface Methodology (RSM) were used in the design of the experiment and in the analysis of the results. Optimum levels of the effective medium constituents were albumin (6.586%); (NH4)2SO4, 0.164%; and glucose, 6.72%. The alkaline protease production increased from 533460 to 793492 Ul(-1).
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PMID:Response surface optimization of effective medium constituents for the production of alkaline protease from a newly isolated strain of Pseudomonas aeruginosa. 1648 Jan 83

We have cloned the proteasome and the proteasome activating nucleotidase (PAN) genes from the mesophilic archaeon Methanosarcina mazei and produced the respective proteins in Escherichia coli cultures. The recombinant complexes were purified to homogeneity and characterized biochemically, structurally, and by mass spectrometry. We found that the degradation of Bodipy-casein by Methanosarcina proteasomes was activated by Methanosarcina PAN. Notably, the Methanosarcina PAN unfolded GFP-SsrA only in the presence of Methanosarcina proteasomes. Structural analysis by 2D averaging electron microscopy of negatively stained complexes displayed the typical structure for the proteasome, namely four-striped side-views and sevenfold-symmetric top-views, with 15 nm height and 11 nm diameter. The structural analysis of the PAN preparation revealed also four-striped side-views, albeit with a height of 18 nm and sixfold-symmetric top-views with a diameter of 15 nm, which corresponds most likely to a dimer of two hexameric complexes. Mass spectrometric analysis of both the Methanosarcina and the Methanocaldococcus PAN proteins indicated hexameric complexes. In summary, we performed a functional and structural characterization of the PAN and proteasome complexes from the archaeon M. mazei and described unique new structural and functional features.
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PMID:Functional and structural characterization of the Methanosarcina mazei proteasome and PAN complexes. 1669 Mar 22

An alkalophilic bacterial isolate identified as Bacillus pantotheneticus, isolated from saline-alkali soils of Avadh region of UP, India, was studied for the production of alkaline protease. The mutant of the isolated species showed 44% improved production over the parent strain. Organic nitrogen sources supported better protease production than the inorganic sources. The production of alkaline protease was (242 U/ml) in the medium containing molasses, which was comparable with molasses and wheat bran (285 U/ml) as carbon and nitrogen sources, respectively. Protease production was best at pH 10 and temperature 30 degrees C. The Km (for casein) was 11 mg/ml and Vmax was 380-microg tyrosine/ml/min. The enzyme was stable between pH 7 and 10.7 and temperature between 30 and 60 degrees C with a pH and temperature optimum at 8.4 and 40 degrees C, respectively. The results indicated that molasses was an optimal substrate for alkaline protease production.
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PMID:Improved production of alkaline protease from a mutant of alkalophilic Bacillus pantotheneticus using molasses as a substrate. 1676 94

Hyperparathyroidism (HPT) can be associated with muscle atrophy and weakness. Muscle atrophy is typically caused by increased muscle protein breakdown. The influence of HPT on calpains and the ubiquitin-proteasome pathway, which are important regulators of muscle proteolysis, is not yet known. We examined the expression in skeletal muscle of mu- and m-calpain and the ubiquitin ligases, atrogin-1 and MuRF1, in patients with primary HPT. A biopsy was obtained from the sternohyoid muscle in patients undergoing surgery for primary HPT (n=8) and in normocalcemic control patients undergoing thyroid surgery (n=11). mRNA levels for atrogin-1, MuRF1 and the calcium-regulated proteases, mu- and m-calpain, were determined by real-time PCR. Calpain activity was measured using the calpain-specific substrate, BODIPY-FL-casein, and by zymography. Serum calcium was 11.4+/-0.46 and 9.5+/-0.10 mg/dl in HPT and control patients, respectively (p<0.01). The corresponding phosphate levels were 2.7+/-0.2 and 3.6+/-0.1 mg/dl (p<0.05). Parathyroid hormone serum concentration was 286+/-103 pg/ml (range, 77-946 pg/ml) in patients with HPT and was not measured in control patients. There were no significant differences in mRNA levels for atrogin-1, MuRF1, mu- or m-calpain and in calpain activity between HPT and control patients. The results suggest that the ubiquitin-proteasome and calpain systems are not activated in skeletal muscle in patients with primary HPT, at least not in patients with moderate hypercalcemia.
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PMID:The gene expression and activity of calpains and the muscle wasting-associated ubiquitin ligases, atrogin-1 and MuRF1, are not altered in patients with primary hyperparathyroidism. 1686 32

The proteasome plays a fundamental role in processes essential for cell viability. A loss in proteasome function has been associated with aging, as well as a number of age-related diseases. Defining the mechanism(s) behind this loss in function will add important information regarding the molecular basis for aging. In the current study, we performed an age-based comparison of proteasome function and composition of subunits and regulatory proteins in the neural retina and retinal pigment epithelium (RPE) in Fischer 344 rats. In the RPE, there was no age-dependent difference in activity, subunit composition, or content of proteasome regulators, PA28 and PA700. In contrast, the aged neural retina demonstrated a significant reduction in the chymotrypsin-like activity and decreased degradation of both casein and casein modified by 4-hydroxynonenal. This loss in function could not be explained by differences in subunit composition, content of PA28 and PA700, or reversible modification of cysteine residues. To begin investigating the molecular basis for the age-associated decrement in proteasome function, we modified the cysteine residues in proteasome from young rats with the sulfhydryl-reactive chemical N-ethylmaleimide. We observed inhibition of the chymotrypsin-like activity and decreased degradation of casein that was comparable to that seen in aged retinas. Thus, chemical modification of cysteine provides an in vitro method that partially recapitulates aging proteasome. Further studies are required to confirm irreversible modification of functionally significant cysteine as a potential mechanism behind the age-related loss in proteasome function.
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PMID:Age-dependent inhibition of proteasome chymotrypsin-like activity in the retina. 1725 1


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