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)

Activation of neutrophils was recently reported to be accompanied by large changes in their Cl- content [J. B. Myers, H. F. Cantiello, J. H. Schwartz, and A. I. Tauber. Am. J. Physiol. 259 (Cell Physiol. 28): C531-C540, 1990]. The significance of these ionic changes to the immune response has not been studied. To evaluate the role of intracellular [Cl-], the anionic composition of the cytosol was varied in human neutrophils permeabilized by electroporation or by treatment with streptolysin O. In Cl(-)-rich media, permeabilized but otherwise untreated cells remained quiescent, resembling unstimulated intact cells. In contrast, suspension of permeabilized cells in Cl(-)-depleted media elicited protein phosphorylation, actin polymerization, secretion of lysozyme, and a respiratory burst. The latter was demonstrated by several criteria to be mediated by the NADPH oxidase. The responses observed in Cl(-)-depleted media were insensitive to pretreatment of the cells with pertussis toxin but were inhibited by addition of GDP or by omission of ATP. The data suggest that an early event in signal transduction, common to several effectors, is sensitive to the ionic composition of the cytosol. This component, possibly a GTP-binding protein, may be affected by the anion concentration changes reported to occur during physiological stimulation of neutrophils.
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PMID:Activation of permeabilized neutrophils: role of anions. 163 84

The role of proteasomes in ubiquitin (Ub)-dependent protein degradation was studied by analyzing lysates of human promyelocytic leukemia HL-60 cells by glycerol density gradient centrifugation. High succinyl-Leu-Leu-Val-Tyr-4-methylcoumaryl-7-amide hydrolyzing activity was found in the 26S fraction, whereas the 20S fraction containing proteaomes had no activity. Addition of 0.05% sodium dodecylsulfate to the latter fraction, however, induced marked activity. The 26S, but not the 20S fraction catalyzed ATP-dependent degradation of [125I]lysozyme-Ub conjugate. Depletion from the lysate of ATP caused complete shift of the active 26S complex to the latent 20S form, whereas in the lysate prepared from ATP-depleted cells, ATP converted 20S proteasomes to 26S complexes. The immunoprecipitated 26S complexes were found to consist of proteasomes and 13-15 other proteins ranging in size from 35 to 110 kDa. We conclude that in the lysate, latent proteasomes undergo reversible, ATP-dependent association with multiple protein components to form 26S complexes that catalyze ATP-dependent degradation of Ub-protein conjugates.
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PMID:ATP-dependent reversible association of proteasomes with multiple protein components to form 26S complexes that degrade ubiquitinated proteins in human HL-60 cells. 164 82

A serine protein kinase that phosphorylates the beta-subunit of the insulin receptor has been partially purified 5,000-fold from HeLa cell membranes. The enzyme has been purified by ion-exchange and hydroxylapatite chromatography and sucrose gradient centrifugation; it has an apparent molecular weight of 36,000-43,000 daltons. It exhibits the following properties: (a) it catalyzes the phosphorylation of the autophosphorylated insulin receptor more efficiently than the nonautophosphorylated insulin receptor, (b) it decreases insulin receptor phosphorylation of tubulin but has no effect on insulin receptor phosphorylation of microtubule-associated proteins or reduced and carboxyamidomethylated lysozyme. The enzyme also phosphorylates casein and ribosomal protein S6 and shares many properties with casein kinase I: (a) similar molecular weight, (b) utilization of ATP but not GTP as phosphoryl donor, and (c) sensitivity to inhibition by heparin. Based on several criteria the receptor serine kinase is neither protein kinase C nor the cAMP-dependent protein kinase.
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PMID:Phosphorylation of the insulin receptor by a casein kinase I-like enzyme. 164 67

A simple method was developed for preparation of proteins conjugated with ubiquitin. Heat-denatured 125I-labeled lysozyme was highly ubiquitinated by incubation at pH 9.0 with a ubiquitin-protein ligase system consisting of E1, E2 and E3 that had been partially purified from rabbit reticulocytes by affinity chromatography with ubiquitin as a ligand. The resulting conjugates were separated from free lysozyme and other proteins by successive chromatographies on anion and cation ion-exchange resins. The ubiquitinated 125I-lysozymes recovered in the fraction not adsorbed to either resin served as an efficient substrate for ATP-dependent proteolysis in a reticulocyte lysate or with a purified 26 S protease complex. By the present method, 125I-lysozyme-Ub conjugates can be prepared in 3 h with a high yield of 15-20%.
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PMID:Improved method for preparation of ubiquitin-ligated lysozyme as substrate of ATP-dependent proteolysis. 165 94

Rabbit red blood cells of various ages were separated on Percoll gradients and the activities of two large cytosolic proteases were measured. Both the multicatalytic protease (MCP), assayed by hydrolysis of fluorigenic peptides, and the 26 S ubiquitin/ATP-stimulated protease, assayed by degradation of ubiquitin-lysozyme conjugates, declined 3-fold or less during maturation of rabbit reticulocytes to erythrocytes. The ability of MCP to hydrolyze three classes of peptides decreased in parallel indicating that the 20 S protease is not significantly remodeled during red blood cell maturation.
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PMID:Multicatalytic and 26 S ubiquitin/ATP-stimulated proteases in maturing rabbit red blood cells. 184 90

To investigate the existence of a ubiquitin-dependent protein degradation system in the brain, the proteolytic activity of the cerebral cortex was examined. The soluble extract of rat cerebral cortex degraded 125I-radiolabeled lysozyme in an ATP-dependent manner. The ATP-dependent proteolysis was suppressed with iodoacetamide, which inhibits ubiquitin conjugation, and was abolished by blocking of the amino residues of lysozyme. These results suggest the participation of ubiquitination in the proteolytic activity. An ATP-dependent 125I-ubiquitin-conjugating activity was detected in fraction II from the cerebral cortex. The presence of ATP-dependent proteolytic activity which acted preferentially on ubiquitinated lysozyme was demonstrated, using ubiquitin-125I-lysozyme conjugates as a substrate. The proteinase had a molecular mass of 1500 kDa and displayed nucleotide dependence and sensitivity to various proteinase inhibitors similar to those of the 26S proteinase complex found in reticulocytes. Dialysis of the soluble fraction caused a decrease in the proteolytic activity of ATP-dependent and preferential for ubiquitin-lysozyme conjugates and a reciprocal increase in the ATP-independent free 125I-lysozyme-degrading activity which was scarcely detected before dialysis. The former ATP-dependent proteolytic activity may play a physiological role in the brain.
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PMID:Identification of a ubiquitin- and ATP-dependent protein degradation pathway in rat cerebral cortex. 184 44

Haemoglobin damaged by exposure of red blood cells to oxidants is rapidly degraded by a proteolytic pathway which does not require ATP [Fagan, Waxman & Goldberg (1986) J. Biol. Chem. 261, 5705-5713]. By fractionating erythrocyte lysates, we have purified two proteases which hydrolyse oxidatively damaged haemoglobin (Ox-Hb). One protease hydrolysed small fluorogenic substrates in addition to Ox-Hb. Its molecular mass was approximately 700 kDa and it consisted of several subunits ranging in size from 22 to 30 kDa. This enzyme may be related to the high-molecular-mass multicatalytic proteinase previously isolated from a variety of tissue and cell types. The other Ox-Hb-degrading activity had an apparent molecular mass of 400 kDa on gel filtration, a subunit size of 110 kDa and an isoelectric point between 4.5 and 5.0. This protease also hydrolysed the small polypeptides insulin and glucagon, as well as other large proteins such as lysozyme. Insulin blocked the degradation of Ox-Hb and Ox-Hb blocked the hydrolysis of insulin by the purified protease. Thiol reagents and metal chelators strongly inhibited the hydrolysis of both Ox-Hb and insulin, whereas inhibitors of serine, aspartic and thiol proteases had little effect. These properties suggest that the Ox-Hb-degrading activity purified from rabbit erythrocytes is the cytosolic insulin-degrading enzyme that is believed to play a role in the metabolism of insulin in several tissues. We propose that this enzyme may also function as a key component in a cytoplasmic degradative pathway responsible for removing proteins damaged by oxidants.
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PMID:Purification of a protease in red blood cells that degrades oxidatively damaged haemoglobin. 187 13

The lipopeptide antibiotic surfactin is a potent extracellular biosurfactant produced by various Bacillus subtilis strains. Biosynthesis of surfactin was studied in a cell-free system prepared from B. subtilis ATCC 21332 and OKB 105, which is a transformant producing surfactin in high yield [Nakano, M. M., Marahiel, M. A., & Zuber, P. (1988) J. Bacteriol. 170, 5662-5668]. Cell material was disintegrated by treatment with lysozyme and French press. A cell-free extract was prepared by ammonium sulfate fractionation, which catalyzed the formation of surfactin at the expense of ATP. Lipopeptide biosynthesis required the L-amino acid components of surfactin and D-3-hydroxytetradecanoyl-coenzyme A thioester. D-Leucine which is present in surfactin was not utilized but inhibited the biosynthetic process. The structure of surfactin, synthesized enzymatically in vitro, was confirmed by chromatographic comparison with the authentic compound and by amino acid analyses. An enzyme fraction was prepared by gel permeation chromatography which catalyzed ATP/pyrophosphate exchange reactions dependent on the component amino acids of surfactin. This enzyme fraction was capable of binding substrate amino acids covalently, probably via thioester linkages. The formation of these intermediates was inhibited by various thiol blocking reagents and phenylmethanesulfonyl fluoride. De novo synthesis of the lipopeptide was not observed with this partially purified enzyme fraction most likely due to the lack of an acyltransferase activity required for linking the beta-hydroxy fatty acid to the peptide moiety.
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PMID:Cell-free biosynthesis of surfactin, a cyclic lipopeptide produced by Bacillus subtilis. 190 54

The mechanism involved in the decreased numbers of several trans-membrane proteins such as sodium pump sites, sodium-lithium countertransport, sodium potassium cotransport proteins, proteins mediating the passive efflux of sodium and insulin receptors in erythrocytes from patients with hyperthyroidism is not known. The ATP-dependent proteolytic system which is involved in the loss of trans-membrane proteins during the maturation of the reticulocyte may be involved in the accelerated loss of these membrane proteins. Therefore, the effect of thyroid hormones on the ATP-dependent proteolytic activity of reticulocyte lysates was examined in this study. Reticulocytosis was induced in 14 guinea pigs by phenylhydrazine hydrochloride injections for 5 consecutive days followed by 2 days of rest. T3 (10 micrograms/100 g body weight) was injected into 7 animals on day 4 and day 6. Reticulocyte-rich blood was withdrawn on day 8. Oxygen consumption determined 24 hours after injection of T3 was 25% higher (p less than 0.01) and T3 treated animals had a 2.5 fold higher (p less than 0.01) weight loss than control animals. The ATP-dependent proteolytic activity measured in reticulocyte lysates using 125I labelled lysozyme was 3.6 fold higher in the T3 than in the control group of guinea pigs (p less than 0.01). We conclude that thyroid hormones induce the ATP-dependent proteolytic activity of reticulocyte lysates which may be responsible for the reduced number of several trans-membrane proteins found in erythrocytes from patients with hyperthyroidism.
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PMID:Induction of the ATP-dependent proteolytic system in guinea pig reticulocyte lysates by triiodothyronine. 194 42

The predicted amino acid sequence of the alpha subunit of the rat liver mitochondrial ATP synthase has been obtained by sequencing a cDNA for the alpha subunit. Analysis of the sequence shows that it contains the A and B consensus sequences found in many nucleotide-binding proteins. Twelve amino acids of the rat liver alpha subunit differ from the sequence of the bovine heart alpha subunit; four of these involve differences in charge. The rat liver alpha subunit, from arginine 15 to the C-terminal proline 510, has been overexpressed in Escherichia coli using the alkaline phosphatase promoter (phoA) and leader peptide to direct the export of the expressed protein to the bacterial periplasm. By treating the cells with lysozyme, osmotic shock, and alkaline pH washes, the alpha subunit can be extracted in high yield (greater than 25 mg/liter) and in a high state of purity. The expressed alpha subunit remains soluble at pH 9.5 or greater and precipitates when treated with Mg2+ ions at low millimolar concentration. The bacterially expressed alpha subunit interacts with 2'(3')-O-(2,4,6-trinitrophenyl)adenosine 5'-triphosphate (TNP-ATP), resulting in a marked fluorescence enhancement upon binding. An enhancement of fluorescence is also observed upon the interaction of the alpha subunit with TNP-ADP. Preincubating the alpha subunit with 1.5 mM ATP significantly reduces the fluorescence enhancement seen with TNP-ATP. The alpha subunit binds TNP-ATP with an apparent Kd in the low micromolar range (1-5 microM) and binds TNP-ADP with an affinity at least 10-fold lower. This work shows that the rat liver alpha subunit can be overexpressed in E. coli to yield a large amount of functional protein. With the acquisition of the overexpressed alpha subunit, it is now possible to test the reconstitution of ATPase activity from a mixture of recombinant and rat liver-derived subunits and to test the formation of complexes by the overexpressed alpha and beta subunits of the rat liver F1-ATPase.
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PMID:Mitochondrial ATP synthase. cDNA cloning, amino acid sequence, overexpression, and properties of the rat liver alpha subunit. 213 25


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