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 steady-state kinetics of the alkaline mesentericopeptidase-catalysed hydrolysis of esters of the general formula Ac-X-OMe(OEt) has been studied, "X" being an amino acid residue (Ala, Val, Leu, Ile, Phe, Tyr, Trp, Lys). The values of the specificity ratio kcat/Km indicate that the bonds involving the carboxyl group of amino acids with aromatic and bulky aliphatic side chain are hydrolysed most effectively. On account of this, alkaline mesentericopeptidase is classified as a chymotrypisn-like alkaline protease. The primary specificity of mesentericopeptidase reveals the similarity of this enzyme to the group of subtilisins, as well as the distinctive characteristic feature of the enzyme to hydrolyse Ac-Leu-OMe with an efficiency practically equal to that of aromatic amino acid derivatives. Suggestions are made about the nature of the substrate-binding centre, taking into consideration Schechter's and Berger's concept of the secondary specificity.
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PMID:Primary specificity of alakaline mesentericopeptidase. Kinetic parameters for the hydrolysis of alpha-N-acetyl-L-amino acid methyl esters. 63 84

We have purified proteasomes to apparent homogeneity from the archaebacterium Thermoplasma acidophilum. This proteinase has a molecular mass of about 650 kDa and an isoelectric point of 5.6. The proteasome hydrolyses peptide substrates containing an aromatic residue adjacent to the reporter group, as well as [14C]methylated casein optimally at pH 8.5 and 90 degrees C. The enzyme activity is enhanced severalfold by Mg2+ and Ca2+ at 25-500 mM. This increase in activity results primarily from a change in Km. The serine-proteinase inhibitors diisopropylfluorophosphate and 3,4-dichloroisocoumarin irreversibly inhibit the enzyme, obviously by modification of both the alpha and beta subunits in the proteasome. The inhibition of proteasomal activity by the peptidylchloromethanes, Cbz-Leu-Leu-CH2Cl and Cbz-Ala-Ala-Phe-CH2Cl (Cbz, benzyloxycarbonyl), is reversible and predominantly of a competitive type. The enzyme is not activated by any of the compounds that typically stimulate the activities of the eukaryotic proteasome.
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PMID:Biochemical properties of the proteasome from Thermoplasma acidophilum. 139 84

The multicatalytic proteinase (MCP) complex catalyses cleavage of bonds on the carboxy-group side of basic, hydrophobic or acidic amino acid residues. Originally, it was proposed that the complex contained three distinct types of catalytic component. MCP from rat liver has been assayed for so-called trypsin-like activity with Boc-Leu-Ser-Thr-Arg-NH-Mec (Mec, 4-methylcoumarin; Boc, t-butoxycarbonyl), for chymotrypsin-like activity with Ala-Ala-Phe-NH-Mec and Suc-Leu-Leu-Val-Tyr-NH-MEc (Suc, succinyl), and peptidyl-glutamylpeptide hydrolase activity with Cbz-Leu-Leu-Glu-Nap (Nap, naphthylamide; Cbz, benzyloxycarbonyl). Results of these studies suggest that as many as five distinct components can be distinguished, one for the trypsin-like activity and two for each of the others. The activities were tested with a variety of serine-protease inhibitors, and other novel effectors have also been identified. The two most effective inhibitors were 4-(2-amino-ethyl)benzenesulphonyl fluoride, which selectivity inactivates the trypsin-like activity, and 3,4-dichloroisocoumarin which inhibits chymotrypsin-like activity and the second, cooperative component [Djaballah, H. & Rivett, A. J. (1992) Biochemistry 31, 4133-4141] of peptidylglutamylpeptide hydrolase activity. The three activities inhibited by 3,4-dichloroisocoumarin can easily be distinguished by the effects of chymostatin analogues, diisopropylfluorophosphate, guanidine/HCl and casein. The results support the view that the enzyme is a novel type of serine protease and suggest that it may contain at least five distinct catalytic components. Marked differences in the reactivities of the different catalytic sites with different reagents can be used to distinguish between them.
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PMID:Use of serine-protease inhibitors as probes for the different proteolytic activities of the rat liver multicatalytic proteinase complex. 142 69

The multicatalytic proteinase (MCP) complex or proteasome is a major nonlysosomal proteinase of eukaryotic cells. The proteinase can cleave peptide bonds on the carboxyl side of hydrophobic, basic, or acidic amino acid residues. These activities have been referred to as "chymotrypsin-like", "trypsin-like", and "peptidylglutamyl-peptide hydrolase" activities, respectively, and have been shown to be catalyzed at distinct sites. The latter activity is often assayed with the synthetic peptide substrate Z-Leu-Leu-Glu-beta-naphthylamide (LLE-NA). N-tBoc-Ala-Ala-Asp-SBzl is also a substrate for the rat liver MCP, suggesting a broader specificity for cleavage on the carboxyl side of acidic residues than the peptidylglutamyl-peptide hydrolase activity previously reported. The pH optimum is in the range of pH 7.0-7.5. Studies of the dependence of velocity on LLE-NA concentration show (a) that there is a high-affinity site (LLE1) which obeys Michaelis-Menten kinetics with a Km value of approximately 100 microM and (b) that at higher substrate concentrations (LLE2) the curve is sigmoidal, suggesting either allosteric activation of the proteinase at a second site or the involvement of multiple catalytic sites which display positive cooperativity. Activity at the high-affinity site (LLE1) can be distinguished from that of the activity of the LLE2 component by the effect of inhibitors, divalent metal ions, and KCl, as well as by its response to heat treatment. The addition of 1 mM MnCl2 stimulates both LLE1 and LLE2 activities and also permits saturation of MCP with substrate at concentrations of LLE-NA below the solubility limit of this peptide.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Peptidylglutamyl-peptide hydrolase activity of the multicatalytic proteinase complex: evidence for a new high-affinity site, analysis of cooperative kinetics, and the effect of manganese ions. 156 59

Two metalloendopeptidases, designated as Streptomyces griseus metalloendopeptidases I and II (SGMPI and SGMPII), were isolated from a commercial Pronase P by a method including affinity chromatography on carbobenzoxy-L-alaninyl-triethylenetetraminyl-Sepharose (Z-Ala-T-Sepharose). The two enzymes differed from each other in behavior on ion-exchange chromatography but showed the same amino-terminal sequence at least up to the 20th residue. Their molecular weights were both estimated to be 37,000 by SDS-polyacrylamide gel electrophoresis. Elemental and amino acid composition analyses indicated that both of them contained about 1 g atom of zinc and one cystine residue per mol of protein. Cleavage specificities of the two enzymes toward synthetic peptide-substrates were very similar to those observed with thermolysin. EDTA, o-phenanthroline, and phosphoramidon strongly inhibited these enzymes, while typical serine-protease inhibitors and cysteine-protease inhibitors had no effect. The findings clearly indicate that SGMPI and SGMPII can be classified into the family of zinc-endopeptidases. It was unexpectedly found, however, that these metalloendopeptidases were strongly inhibited by protein serine-protease inhibitors produced by Streptomycetes, such as Streptomyces subtilisin inhibitor (SSI), alkaline protease inhibitor-2c' (API-2c'), and plasminostreptin (PS).
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PMID:Purification and characterization of Streptomyces griseus metalloendopeptidases I and II. 176 59

The proteolytic activities of the 20 S proteasome were found to change in their levels during the development of chick embryonic muscle. The peptide-cleaving activities against N-succinyl-Leu-Leu-Val-Tyr-7-amido-4-methylcoumarin and N-benzyloxycarbonyl-Ala-Arg-Arg-4-methoxy-beta-naphthylamide gradually decreased with the time of development. On the other hand, the casein-degrading activity in the presence of poly-L-lysine markedly increased from embryonic day 11 and reached a maximal level by day 17. These changes appeared to be tissue-specific because little or no change in any of the proteolytic activities was observed with developing embryonic brain, while dramatic alterations occurred in the extents of the peptide hydrolyses in liver. Furthermore, a number, but not all, of the proteasome subunits in embryonic muscle were changed in their amounts during the development. These results suggest that the alterations in the proteasome activities and subunit pattern are developmentally regulated and may be correlated.
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PMID:Developmental regulation of proteolytic activities and subunit pattern of 20 S proteasome in chick embryonic muscle. 187 33

The egg jelly-induced acrosome reaction of the sea urchin, Strongylocentrotus intermedius, was inhibited by succinyl-Leu-Leu-Val-Tyr-4-methyl-coumaryl-7-amide (Suc-Leu-Leu-Val-Tyr-MCA), but not by Suc-Ala-Ala-Pro-Phe-MCA. The proteases with hydrolytic activity toward the former were purified from sperm extract by DEAE-Sephacel and hydroxylapatite chromatographies, Sephacryl S-300 gel filtration, and heparin-Sepharose CL-6B chromatography. Two types of protease were separated, and the molecular weights were estimated to be 65 and 700 kDa, respectively, by gel filtration. The former was accompanied by hydrolytic activity toward Suc-Ala-Ala-Pro-Phe-MCA, which was not hydrolyzed by the latter. Polyacrylamide gel electrophoresis of 700 kDa protease gave a single protein band under nondenaturing conditions and at least eight bands in the range of 22-33 kDa in the presence of sodium dodecyl sulfate (SDS). The substrate specificity and the inhibitor sensitivity of 700 kDa protease indicate that it contains two types of the activity, one is chymotrypsin-type and the other trypsin-type. The former activity was enhanced by poly-L-lysine or SDS. These properties of 700 kDa protease are similar to those of proteasomes (multicatalytic proteinases) isolated from various eukaryotic sources. We had previously shown that inhibitors of chymotrypsin-like proteases inhibit the increase of intracellular Ca2+ concentration by egg jelly, resulting in the inhibition of the acrosome reaction of St. intermedius (Matsumura and Aketa, Gamete Res 23:255-266, 1989). Bringing these findings together, we suggest that the chymotrypsin-like activity of sperm proteasome participates in the onset of the acrosome reaction of St. intermedius.
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PMID:Proteasome (multicatalytic proteinase) of sea urchin sperm and its possible participation in the acrosome reaction. 187 26

A novel endogenous inhibitor of the proteasome (high molecular weight multicatalytic protease) has been isolated and characterized from human erythrocytes. After purification by ion-exchange and sizing chromatography, the inhibitor displayed a native molecular mass of approximately 200 kDa and contained a single subunit of 50 kDa with an isoelectric point of 6.9. Although the inhibitor noncompetitively blocks proteolysis of [methyl-14C]-alpha-casein (Ki = 7.1 x 10(-8) M) and inhibits hydrolysis of Suc-Leu-Leu-Val-Tyr-AMC, it did not affect hydrolysis of other peptide substrates, such as MeOSuc-Phe-Leu-Phe-MNA and Z-Ala-Arg-Arg-MNA. To further characterize the 50-kDa inhibitor, a monoclonal antibody (MI-8) was generated that showed specific binding upon Western blot analysis of both native PAGE and SDS-PAGE. Immunoprecipitation with MI-8 specifically removed inhibitor activity against the proteasome. The 50-kDa inhibitor is distinct from a previously described 40-kDa inhibitor of the proteasome (Murakami, K., & Etlinger, J.D. (1986) Proc. Natl. Acad. Sci. U.S.A. 83, 7588-7592) on the basis of lack of cross-reactivity with MI-8 and dissimilar peptide digest patterns. It is suggested that these endogenous inhibitors may have a role in ATP/ubiquitin-dependent proteolysis and/or other cellular functions involving this protease.
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PMID:Isolation and characterization of a novel endogenous inhibitor of the proteasome. 191 59

Of the two proteases produced by Pseudomonas aeruginosa, one whose optimal pH is neutral, exhibits elastolytic activity. This elastase is produced as a prepropeptide, subsequently modified by proteolysis, and finally secreted as an active enzyme. Both proteases act mainly on hydrophobic aminoacids. The most important site for the elastase seems to be the P'1 site where Phe, Tyr and Leu residues enhance hydrolysis. The alkaline protease is less specific of this site. At least four aminoacids are needed to obtain measurable rates of hydrolysis. A synthetic substrate, Ala-Ala-Phe-Ala, is proposed for conductimetric measurements of Pseudomonas aeruginosa elastase activities in the nanomolar range. A review of recent studies shows that a very wide range of proteins can be hydrolyzed by the two Pseudomonas aeruginosa proteases, a fact which may explain why these enzymes are major determinants of the bacteria's infectivity.
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PMID:[Mechanism of action of proteinases of Pseudomonas aeruginosa]. 212 91

Pseudomonas aeruginosa elastase is a zinc metalloproteinase which is released during P. aeruginosa infections. Pseudomonas keratitis, which occurs following contact lens-induced corneal trauma, can lead to rapid, liquefactive necrosis of the cornea. This destruction has been attributed to the release of both host-derived enzymes and the bacterial products P. aeruginosa elastase, alkaline protease, exotoxin A, and lipopolysaccharide endotoxin. A synthetic metalloproteinase inhibitor, HSCH2 (DL)CH[CH2CH(CH3)2]CO-Phe-Ala-NH2, which we previously showed to be a potent inhibitor of corneal collagenase and alkali-induced corneal ulceration, was tested as a potential inhibitor of P. aeruginosa elastase. Inhibition constants (Kis) for the resolved diastereomers were determined with the chromogenic substrate furylacryloyl-glycyl-L-leucyl-L-alanine. One isomer had a Ki of 0.3 microM, while the other had a Ki of 0.4 microM. The more potent diastereomer was evaluated in vivo in experimentally induced Pseudomonas keratitis in rabbits. Following inoculation of one cornea of each rabbit, topical treatment with a 1 mM solution of the inhibitor significantly delayed the onset of corneal melting and perforation, as compared with the results for the control and gentamicin-treated groups. This protective effect suggests that the inhibitor may have a therapeutic application by delaying the progression of corneal destruction in Pseudomonas keratitis.
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PMID:Inhibition of Pseudomonas aeruginosa elastase and Pseudomonas keratitis using a thiol-based peptide. 212 41


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