Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0272170 (SDS)
 50,377 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The multicatalytic endopeptidase complex (20S proteasome) is a latent high-molecular-mass multisubunit proteinase. In many investigations, SDS has been used as a proteasome activator at some fixed concentration that was apparently optimal. This study examined the effects of various divalent cations on the SDS-dependent peptidase and casein degradation activities of 20S proteasome purified from Xenopus laevis oocytes at a series of SDS concentrations and the correlation between these effects and the critical micelle concentration (CMC) of SDS. Surprisingly, it was found that divalent cations such as Mg2+ markedly shifted the SDS-dependent activation profiles to a lower concentration range. Ca2+, Mn2+, Co2+, and Zn2+ also markedly reduced the optimum SDS concentration in the Suc-Leu-Leu-Val-Tyr-MCA hydrolysis reaction: for example, 5 mM Co2+ reduced the optimum SDS concentration from 0.065 to 0.005%. However, in all cases examined the optimum concentrations were below the CMC. Cu2+, Hg2+, and Cd2+ strongly inhibited the SDS-dependent maximum activity without remarkably shifting the optimum SDS concentration. No correlation between the shift and the inhibition was recognized. Most interestingly, remarkable activation of casein degradation by SDS was observed only by addition of the divalent cations Mg2+, Ca2+, and Mn2+. These cations might be essential for casein degradation. The activation and inactivation ranges of SDS concentration varied with the species of substrate.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Reaction of 20S proteasome: shift of SDS-dependent activation profile by divalent cations. 749 Feb 55

The aim of this study was to purify epidermal cathepsin B from rat skin and investigate its proteolytic activities on filaggrin and several synthetic substrates. The molecular weight of purified monomeric cathepsin B was estimated to be 30 kDa by SDS-polyacrylamide gel electrophoresis. The amino acid composition, similar to that of liver cathepsin B, indicated the enzyme to be an acidic protease. The enzyme had strong hydrolytic activity toward N-benzyloxy-carbonyl-L-arginyl-L-arginine-7-amido-4-methylcoumarin (Z-Arg-Arg-MCA) (152 mU/mg) and N-benzyloxycarbonyl-L-phenylalanyl-L-arginine-7-amido-4-methylcoumarin (424 mU/mg), but had no proteolytic activity toward L-arginine-7-amido-4-methylcoumarin. The Km value for Z-Arg-Arg MCA was 0.34 mM and pH optimum was 5.5. Cathepsin B degraded rat epidermal filaggrin into small fragments at pH 4.0 and 5.5., and was inhibited by a specific cysteine proteinase inhibitor, N-[N-(L-3-trans-carboxyoxirane-2-carbonyl)L-leucyl]- agmatin. This study demonstrated that filaggrin was susceptible to degradation by cathepsin B. Such an action may have relevance to skin differentiation in which acid proteases are thought to participate.
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PMID:Rat epidermal cathepsin B: purification and characterization of proteolytic properties toward filaggrin and synthetic substrates. 776 85

A novel serine proteinase, designated as prostasin, has been purified from human seminal fluid to apparent homogeneity by DEAE-Sepharose CL-6B and aprotinin-affinity chromatography. The purified protein migrates as two close bands with an apparent molecular mass of 40 kDa on SDS-polyacrylamide gel electrophoresis under reducing conditions. It can be labeled with [14C]diisopropyl fluorophosphate and has a pI ranging from 4.5 to 4.8. Sequence analysis reveals that the two protein bands have an identical NH2-terminal amino acid sequence which is different from any known protein sequence in the SwissPro or GenBank data base. The NH2-terminal 20-amino acid sequence shares 50-55% identity with human alpha-tryptase, elastase 2A and 2B, chymotrypsin, acrosin, and the catalytic chains of hepsin, plasma kallikrein, and coagulation factor XI. Prostasin has trypsin-like activity with a pH optimum of 9.0, hydrolyzing peptidyl fluorogenic substrates: D-Pro-Phe-Arg-MCA, D-Phe-Phe-Arg-MCA, D-Val-Leu-Arg-MCA, and Z-Gly-Pro-Arg-AFC. It is inhibited by aprotinin, antipain, leupeptin, and benzamidine. The tissue distribution of prostasin was determined by a newly developed radioimmunoassay. Linear displacement curves for immunoreactive prostasin in body fluids and tissues were parallel with the standard curve of purified prostasin, indicating their immunological identity. Immunoreactive prostatin levels were 8.61 +/- 0.42 microgram/ml in the seminal fluid and 0.201 +/- 0.029 microgram/ml in urine. Prostasin is present at high levels in the prostate gland (143.7 +/- 15.9 ng/mg protein), moderate levels (2-6 ng/mg protein) in colon, lung, kidney, pancreas, salivary gland, liver, and bronchi, but it is not detected in the brain, muscle, testis, ventricle, atrium, and aorta. Immunohistochemical localization reveals that prostasin is present in epithelial cells and ducts of the prostate gland. These studies indicate that prostasin purified from seminal fluid is a novel serine proteinase and originates from the prostate gland.
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PMID:Prostasin is a novel human serine proteinase from seminal fluid. Purification, tissue distribution, and localization in prostate gland. 803 38

The effect of phospholipids on the trypsin-like, chymotrypsin-like and peptidylglutamyl-peptide-hydrolysing activities of the so-called latent form of the rat liver multicatalytic proteinase was studied, assaying them with the following substrates: N-Cbz-ARR-4MNA (N-Cbz, N-benzyloxycarbonyl; 4MNA, 4-methoxy-beta-naphthylamide), N-Suc-LLVY-MCA (N-Suc, N-succinyl; MCA, methylcoumarin) and N-Cbz-LLE-beta-NA (beta-NA, beta-naphthylamide) respectively (amino acids are shown as their one-letter symbol). For the most part neither lysophospholipids nor phospholipids at 20 micrograms/ml have any effect on the activity of the enzyme (assayed at 50 microM peptide), except for phosphatidylserine, which activates 2-fold the hydrolysis of N-Suc-LLVY-MCA, and phosphatidylinositol, which inhibits by 20% the hydrolysis of N-Cbz-LLE-beta-NA. By contrast, cardiolipin (diphosphatidylglycerol) is a strong activator of the hydrolysis of N-Suc-LLVY-MCA (60-fold) and N-Cbz-LLE-beta-NA (30-fold), with half-maximal activation at concentrations of 0.15 micrograms/ml and 1.5 micrograms/ml respectively. The activation of N-Suc-LLVY-MCA hydrolysis is due to an increase of the affinity of the enzyme for the peptide and to an increase in the Vmax. (30-fold). The activation of N-Cbz-LLE-beta-NA hydrolysis is explained by suppressing the co-operativity for this substrate, producing hyperbolic kinetics with a Km of 60 microM and a 15-fold increase in the Vmax. of the enzyme. This activation by cardiolipin was completely suppressed by micromolar concentrations of fluophenazine, a drug known to inhibit other phospholipid-regulated process. Cardiolipin activation and the known activation by SDS are additive, either at suboptimal or optimal concentrations of both activators. Cardiolipin also activates the in vitro degradation of some proteins from metabolically labelled total cellular extracts by the latent multicatalytic proteinase. These results clearly show that cardiolipin is a natural positive modulator of the peptidase and proteolytic activities of the multicatalytic proteinase, probably acting through a binding site different from that of SDS.
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PMID:Kinetic mechanism of activation by cardiolipin (diphosphatidylglycerol) of the rat liver multicatalytic proteinase. 825 Aug 60

An endogenous activator of 20S proteasome was purified from human platelets and its effect on three peptidase activities of proteasome was studied. This activator had a molecular weight of 170 kDa, and was composed of 32 kDa polypeptides as determined by SDS-PAGE. It was highly labile upon heat treatment (56 degrees C, 20 s) and proteinase (pronase CB) digestion. Suc-LLVY-MCA degrading activity of the platelet proteasome showed positive cooperativity between two or more catalytic sites because the coefficient was 1.54 when analyzed by use of the Hill plot. The endogenous activator increased Vmax and caused a loss of cooperativity. The plot of reaction velocity as a function of activator concentration yielded a saturation curve, implying the binding of the activator to proteasome. Boc-LTR-MCA degrading activity followed Michaelis-Menten kinetics. The activator enhanced the activity by increasing Vmax and decreasing Km. In contrast, CBz-LLE-2NA degrading activity could not be analyzed according to any kinetic scheme reported so far. The activator stimulated this activity at lower substrate concentrations (below 200 microM), while it inhibited the activity at higher substrate concentrations (400-800 microM). It is concluded from these findings that the endogenous protein activator may regulate the intracellular proteasome activity by functioning as a positive allosteric effector.
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PMID:Purification and characterization of endogenous protein activator of human platelet proteasome. 828 19

Metalloproteinases, myosinase I and myosinase II, that hydrolyze the heavy chain of myosin, were purified from squid mantle muscle. Myosinase I does not hydrolyze other muscle proteins, casein, haemoglobin, or MCA-substrates, while II hydrolyzes tropomyosin. Both myosinase I and myosinase II gave a single protein band on SDS-PAGE with a molecular mass of 16 and 20 kDa, respectively. Their activities were inhibited by EDTA and 1,10-phenanthroline, and II was also inhibited by EGTA. They could be reactivated with some divalent cations, I was especially reactivated with Co2+ and II especially with Zn2+. The optimum pH of both activities was 7.0; the optimum temperature for both was 40 degrees C. Myosinase I hydrolyzes myosin heavy chains to produce 130 and 90 kDa fragments. The N-terminal amino-acid sequence of the 90 kDa fragment indicates that myosinase I splits the myosin heavy chain between Ala-1161 and Thr-1162 in subfragment 2. Myosinase II hydrolyzes myosin heavy chain to produce 158 and 65 kDa fragments, and it splits between Glu-1381 and Thr-1382 in LMM. Myosinases I and II are most likely related to the metabolism of myosin in vivo.
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PMID:Purification and characterization of two metalloproteinases from squid mantle muscle, myosinase I and myosinase II. 842 25

A ubiquitin (Ub)/ATP-dependent proteolytic complex (26S proteasome) purified from rabbit skeletal muscle was dissociated into two subcomplexes, a 20S proteasome and a regulatory subunit complex, by preparative non-denaturing polyacrylamide gel electrophoresis (PAGE). The isolated regulatory subunit complex preparation gave a single broad band on analytical non-denaturing PAGE, and several bands ranging between 33 and 110 kDa on SDS-PAGE. This complex was found to consist of about 20 subunits on the basis of two-dimensional PAGE, the pattern of which appeared identical or very similar to that of the 33-110 kDa 26S proteasome subunits. The apparent molecular mass of the complex was estimated to be 1100 kDa by Ferguson plot analysis and also by Superose 6 gel filtration. Unlike the 26S proteasome, neither ATPase activity nor protease activities toward Suc-Leu-Leu-Val-Tyr-MCA, Boc-Phe-Ser-Arg-MCA, Z-Leu-Leu-Glu-beta NA, [14C]-casein, [125I]-lysozyme and Ub-[125I]-lysozyme were significantly detectable in the regulatory subunit complex. This complex was found to be capable of associating with itself in MgATP-dependent manner. These results suggest that a regulatory subunit complex dissociated from the 26S proteasome comprises all the higher molecular mass subunits of the 26S proteasome, and has no detectable ATPase and protease activities, although the homo-oligomerization occurs in an ATP-dependent fashion.
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PMID:Regulatory subunit complex dissociated from 26S proteasome: isolation and characterization. 854 9

Dipeptidyl peptidase II (DPP II) was purified to homogeneity from porcine seminal plasma by polyacrylamide gel electrophoresis (PAGE). The molecular weight of the purified enzyme was calculated to be approx. 185,000 and 200,000 on Superdex 200 column chromatography and non-denatured PAGE, respectively, and to be 58,000 and 61,000 on SDS-PAGE in the absence and presence of beta-mercaptoethanol (beta-ME), respectively. These findings suggested that the enzyme is composed of three identical subunits. The enzyme rapidly hydrolyzed the substrates Lys-Ala-MCA and Gly-Pro-MCA at acidic pH. The Km and V(max) values of DPP II at optimal pH (pH 6.0) were 1330 microM and 2.9 mumol/mg per min for Gly-Pro-MCA, and 360 microM and 1.43 mumol/mg per min for Lys-Ala-MCA, respectively. It was strongly inhibited by diisopropylphosphofluoride (DFP), and moderately by 4-(2-aminoethyl)benzenesulfonyl fluoride (AEBSF). These findings suggest that DPP II is a serine peptidase. Furthermore, the enzyme activity was also strongly inhibited by copper ions. The amino-acid sequence of the first 41 residues of the enzyme was determined as Ala1-Ser-Pro-Pro-Glu-Pro-Gly-Phe-Arg- Glu10-Val-Tyr-Phe-Glu-Gln-Leu-Leu-Asp-His-Phe20-Asn-Phe-Glu- Arg-Phe- Gly-Lys-Lys-Thr-Phe30-Arg-Gln-Arg-Phe-Leu-Val-Ser-Asp-Lys-Phe40 -Trp. This sequence showed homology (11.6-30.2%) to the N-terminal amino-acid sequences of cytotoxic cell proteinases (CCP 1-4), granzymes. Other properties of DPP II including pH optimum, pH stability, and heat stability were characterized.
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PMID:Dipeptidyl peptidase II from porcine seminal plasma: purification, characterization, and its homology to granzymes, cytotoxic cell proteinases (CCP 1-4). 864 18

In order to clarify the mechanism of activation plant 20S proteasomes by fatty acids, we examined the effects of oleic, linoleic and linolenic acids on the three of peptidase activities of purified 20S proteasomes from spinach leaves and compared them with the effects of SDS, a previously characterized activator of 20S proteasomes. The three fatty acids all activated the hydrolysis of succinyl-Leu-Leu-Val-Tyr-4-methylcoumaryl-7-amide (Suc-LLVY-MCA) and benzyloxycarbonyl-Leu-Leu-Glu-2-naphthyl-amide (Cbz-LLE-2NA) at low concentrations (one-third to one-sixth of that required for activation by SDS). The range of concentrations of linolenic acid for the activation of Suc-LLVY-MCA hydrolysis was very narrow. All the fatty acids inhibited the hydrolysis of tert-butoxycarbonyl-Leu-Arg -Arg-4-methylcoumaryl-7-amide (Boc-LRR-MCA) at extremely low concentrations (one-fifth to one-fifteenth of that required for the activation of the hydrolysis of Suc-LLVY-MCA and Cbz-LLE-2NA). In the case of hydrolysis of Suc-LLVY-MCA, SDS and the three fatty acids increased the Vmax value and decreased the apparent Km value to similar relative extents. In the case of hydrolysis of Boc-LLE-MCA, SDS and the three fatty acids also decreased the Km and increased the Vmax. However, SDS markedly increased Vmax. the curves representing the SDS-dependent activation were shifted to a lower range by the addition of linoleic acid, but the maximum activity at the optimum concentration of SDS was essentially unchanged. These results suggest that the activation by SDS and that by the fatty acids has an additive effect. The results imply that fatty acids, such as linolenic acid, might act as physiological regulators in plant cells.
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PMID:Activation of 20S proteasomes from spinach leaves by fatty acids. 866 92

Cytochrome P450-dependent 7-ethoxyresorufin O-deethylase (EROD), 7-pentoxyresorufin O-dealkylase (PROD) and 7-ethoxycoumarin O-deethylase (ECOD) activities in 14-day-old chick embryo livers were determined 24 h after pretreatment with selected widespread aromatic environmental contaminants, including polychlorinated biphenyls (PCBs), polycyclic aromatic hydrocarbons (PAHs), hexachlorobenzene, and dialkylesters of phthalic acid, and compared with the inducing potencies of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and the coplanar and mono-o-chlorinated PCBs. The effects of other model inducers, i.e. phenobarbital and pyrazole, were also examined. Specificity of EROD induction was estimated with regard to contaminants frequently present in environmental samples and dose-response curves for EROD induction were determined. A strong induction (comparable with that by mono-o-chlorinated biphenyl treatment) by dibenzo[a,h]anthracene, benzo[k]fluoranthene or benzo[b]fluoranthene was found, but the maximal level of EROD activity inducible by TCDD was not achieved, partly due to the high toxicity of the tested PAHs. 3-Methylcholanthrene showed moderate inducing potencies; benz[a]anthracene, benzo[a]pyrene, chrysene and 2,2',3,4,4',5'-hexachlorobiphenyl appeared to be weak inducers. Other PAHs and PCBs tested, as well as hexachlorobenzene, dialkyl phthalates, phenobarbital and pyrazole had no marked effects on the EROD level. ECOD activities were increased non-specifically by TCDD, 3-methylcholanthrene, hexachlorobenzene and phenobarbital. A significant enhancement of PROD activity by TCDD and related inducers was observed, while phenobarbital induced the PROD activity only weakly; SDS-PAGE analysis showed that the chicken phenobarbital-inducible cytochromes P4502H with apparent molecular weights 50 kDa were not markedly induced by the TCDD- or 3-methylcholanthrene treatments. Inhibition of EROD and PROD by 9-hydroxyellipticine, a specific inhibitor of rat hepatic cytochrome P4501A1, revealed that PROD induction by TCDD and other P4501A-inducers was probably a result of a broader substrate specificity of chick embryo P4501A. Measurement of EROD activities in chick embryo liver is highly sensitive, specific and suitable for the determination of TCDD-type toxicity of new drugs, agrochemicals, and industrial pollutants.
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PMID:Induction effects of polychlorinated biphenyls, polycyclic aromatic hydrocarbons and other widespread aromatic environmental pollutants on microsomal monooxygenase activities in chick embryo liver. 897 35


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