Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.2.1.17 (lysozyme)
 21,489 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The substrate specificity of the catalytic subunit of rabbit skeletal muscle 3': 5'-cyclic AMP-dependent protein kinase (EC 2.7.1.37; ATP: protein phosphotransferase) has been studied using the synthetic peptide Arg-Gly-Tyr-Ser-Leu-Gly corresponding to the sequence around serine 24, a phosphorylation site in reduced, carboxymethylated, maleylated (RCMM) chicken egg white lysozyme. This peptide served as a substrate for the enzyme and exhibited a 6-fold higher Vmax and a 100-fold higher Km than RCMM-lysozyme. Replacement of the arginine with glycine, histidine, or lysine resulted in a dramatic reduction in the Vmax. These results support the concept that arginine is an important residue in determining the substrate specificity of the protein kinase, predominantly influencing the Vmax of the phosphorylation reaction. Two synthetic peptides in which serine was replaced by an alanine acted as competitive inhibitors of phosphorylation of the synthetic peptide substrate and RCMM-lysozyme.
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PMID:Synthetic hexapeptide substrates and inhibitors of 3':5'-cyclic AMP-dependent protein kinase. 17 70

The phosphate content of rat thymus histones was determined. As expected for a replicating tissue, histones 1 and 2B were more phosphorylated and had higher 32P uptakes than did histones from resting liver nuclei; the other histones all showed 32P uptake, but the phosphate content and uptake of histone 2A was about half that for liver histone 2A. When thymus nuclei were incubated in a slightly hypo-osmotic medium, non-histone proteins and phosphorylated histones were released into solution; this was enhanced if ATP was present in the medium. [gamma-32P]ATP was incorporated into non-histone proteins, including protein P1, and into the ADP-ribosylated form of histone 1; negligible 32P was incprporated into the other, bound, histones. Histones 1 and 2B added to the incubation medium were extensively, and histones 2A and 4 slightly, phosphorylated. Histones released by increasing the ionic strength of the medium were phosphorylated. Added lysozyme and cytochrome c were neither bound nor phosphorylated, but added non-histone protein P1 was phosphorylated, causing other histones to be released from the nuclei, especially histones 2A and 3. The released histones were phosphorylated. gamma-Irradiation decreased 32P uptake into the non-ADP-ribosylated histones 1 and 4; phosphorylation of histone 1 in vitro was unaffected. The importance of non-histone proteins, ATP availability and nuclear protein kinases to the control of histone phosphorylation in vivo is discussed.
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PMID:Phosphorylation of rat thymus histones, its control and the effects thereon of gamma-irradiation. 19 8

Protein kinase, which was isolated from cells infected with T7, is indeed a viral gene product. This is shown by DNA-dependent synthesis in vitro. The protein kinase transfers phosphate from ATP to seryl or threonyl residues in protein. The enzyme has only a relative requirement for magnesium ions, but is only active at low ionic strength. The best substrate is lysozyme. T7 protein kinase activity is not stimulated by cyclic 3':5'-AMP and/or cyclic 3':5'-GMP. The T7 protein kinase carries -- SH groups essential for activity. There is indication that the enzyme phosphorylates itself and causes self inactivation, which may explain the fast disappearance of enzyme activity in vivo. Bacteriophage T3 also induces a protein kinase which is similar to the T7-induced enzyme in all respects tested.
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PMID:Protein kinase of bacteriophage T7. 2. Properties, enzyme synthesis in vitro and regulation of enzyme synthesis and activity in vivo. 24 Jun 95

Genetically differerent clones of myeloid leukemic cells have been used to study the activation of normal genes in these malignant cells by the normal physiological inducer of myeloid cell differentiation, the protein MGI. In appropriate clones, MGI induced the normal differentiation-associated property of chemotaxis to a variety of compounds including the steroid hormone dexamethasone. The induced cells could also distinguish among different steroids by chemotaxis, suggesting that there are specific membrane interaction sites for steroids. The sequence of differentiation in these cells was the formation of C3 and Fc rosettes leads to phagocytosis of these rosettes and chemotaxis leads to synthesis and secretion of lysozyme leads to mature macrophages or granulocytes. The use of appropriate mutants and the comparison of induction by MGI and dexamethasone has shown that chemotaxis to casein can be dissociated from: chemotaxis to dexamethasone, ATP, and bacterial factor; formation of C3 or Fc rosettes; phagocytosis of these rosettes; synthesis of lysozyme; and the formation of mature cells. It is suggested from this dissection of normal differentiation that there are different membrane changes for specific chemotaxis, formation of these rosettes, and their phagocytosis, and that induction of each of these properties requires activation of different genes.
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PMID:Activation of normal genes in malignant cells: activation of chemotaxis in relation to other stages of normal differentiation in myeloid leukemia. 29 88

Initiation of new DNA synthesis was observed in B. subtilis cells upon gamma-ray irradiation followed by toluene treatment and incubation in the presence of the four deoxynucleotide triphosphates and Mg2+. This DNA synthesis took place in the absence of ATP and was refractory to 6-(p-hydroxyphenylazo)-uracil which is a specific inhibitor for the type III polymerase of Bacillus subtilis. This repair-type DNA synthesis was greatly reduced in mutant cells deficient in DNA polymerase I. Restoration of transforming activity of cellular DNA was found to occur in parellel with the above repair type DNA synthesis. A protein factor which enhances the priming activity of gamma-irradiated DNA for DNA polymerase I was detected in DNA-free extracts prepared from B. subtilis cells by means of lysis with a buffer containing lysozyme, Brij-58 and EDTA.
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PMID:Studies on DNA repair in Bacillus subtilis. I. A cellular factor acting on gamma-irradiated DNA and promoting its priming activity for DNA polymerase I. 80 54

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.
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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.
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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.
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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

We have identified and purified a protein complex from human red blood cells that activates the multicatalytic protease (MCP). The complex, which we call the regulator, sediments at 11 S and is composed of 30-kDa subunits. The regulator does not hydrolyze fluorogenic peptides, but when multicatalytic protease and regulator are combined, MCP cleaves succinyl-Leu-Leu-Val-Tyr-7-amido-4-methylcoumarin and Leu-Leu-Glu-p-nitroanilide as much as 60-fold faster. Hydrolysis of several other fluorogenic peptides is stimulated to a lesser extent, and activated MCP does not degrade ubiquitin-lysozyme conjugates, bovine serum albumin, or lysozyme. Latent and activated forms of MCP display similar sensitivity to protease inhibitors, suggesting that activation does not generate new kinds of catalytic sites. In addition, ATP suppresses peptide hydrolysis by activated and latent MCPs to the same extent. Activation involves binding of regulator to MCP, and activated MCP migrates slower on native acrylamide gels. Dissociation of the MCP regulator complex during prolonged sedimentation on glycerol gradients releases active regulator and MCP molecules capable of being reactivated. Moreover, two-dimensional electrophoresis does not reveal changes in MCP or regulator subunits following activation. Thus, activation appears to result from reversible association of regulator subunits with MCP.
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PMID:Purification of an 11 S regulator of the multicatalytic protease. 142 90

Ascorbic acid is believed to protect cells from oxidative damage by reacting with oxygen-derived free radicals. We investigated whether ascorbic acid would affect the rate of breakdown of skeletal muscle proteins in extracts exposed to hydrogen peroxide. Ascorbic acid (20 mmol/L) alone had little or no effect on the rate of ATP-independent or ATP-dependent breakdown of proteins in chicken skeletal muscle. Pretreatment of chicken skeletal muscle extracts with 10 mmol/L H2O2 resulted in a complete loss of ATP-dependent proteolysis and a significant increase (14- to 15-fold) in the rate of ATP-independent protein breakdown. Ascorbic acid (20 mmol/L) did not prevent H2O2 (10 mmol/L) from inactivating the ATP-dependent proteolytic pathway in skeletal muscle. However, ascorbic acid (20 mmol/L) prevented the H2O2-induced increase in the ATP-independent proteolysis of endogenous muscle proteins. Ascorbic acid also slowed the rate of hydrolysis of exogenously added [3H]superoxide dismutase exposed to H2O2 and inhibited the enhanced degradation of [3H]lysozyme and H2O2-treated [3H]superoxide dismutase by the proteolytic systems exposed to H2O2. Thus ascorbic acid seems to inhibit the H2O2-induced increase in ATP-independent proteolysis 1) by preventing damage to proteins by H2O2 resulting in a decreased supply of substrates for the ATP-independent degradative system and 2) by preventing activation of the proteolytic enzymes that participate in the energy-independent degradation of H2O2-treated proteins.
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PMID:Protective effect of ascorbic acid on the breakdown of proteins exposed to hydrogen peroxide in chicken skeletal muscle. 143 49


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