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:3.2.1.17 (lysozyme)
21,489 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Though DNase does not contain any cysteine residues, incubation of the enzyme with 2-nitro-5-thiocyanobenzoic acid in the presence of Ca2+ at pH values above 7.5 results in an irreversible inactivation of the enzyme. The inactivation also occurs when Ca2+ is replaced by Mg2+, but not in their absence. Amino acid analyses after acid hydrolyses of the completely inactivated ant the native enzymes show no significant differences in composition, including tryptophan and half-cystine residues. However, sodium dodecyl sulfate gel electrophoresis indicates enzyme cleavage by the treatment with 2-nitro-5-thiocyanobenzoic acid. This reagent does not inactivate chymotrypsin and lysozyme, and under conditions where bovine DNase is inactivated, does not inactivate other nucleases such as ribonuclease, snake venom phosphodiesterase, and spleen acid DNase. However, it inactivates malt DNase and can, therefore, be considered a specific inhibitor of DNase I. The inactivation kinetics is pseudo-first order, resembling Michaelis-Menten, with an affinity constant of 16.7 mM. It is the cyano group, not the thionitrobenzoic acid of 2-nitro-5-thiocyanobenzoic acid that reacts to form cyano-DNase.
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PMID:Inactivation of bovine pancreatic DNase by 2-nitro-5-thiocyanobenzoic acid. I. A novel inhibitor for DNase I. 48 54

In the chromatin domain of the chicken lysozyme gene of myeloid and oviduct cells, which both have the potential to activate the gene, a developmentally stable DNase I-hypersensitive site is formed around 6.1 kb upstream of the gene. This implies that this DNA region, which has previously been demonstrated to function as a transcriptional enhancer element in myeloid cells, is intimately involved in the cell-type-specific activation of the lysozyme gene locus. Deletion analysis identifies a 157-bp minimal fragment that confers the same promacrophage-specific enhancer activity as the originally described 562-bp -6.1-kb enhancer fragment. By introducing specific point mutations, we demonstrate in transient gene transfer experiments that the minimal fragment consists of at least six adjacent elements, each substantially contributing to enhancer function. The compact multifactorial enhancer complex includes a nuclear factor I (NF-I)/TGGCA binding site, homologies to AP1, and octanucleotide or enhancer core consensus motifs. Point mutation of the NF-I binding site results in the loss of NF-I binding in vitro and enhancer activity in vivo after gene transfer. Surprisingly, four overlapping oligonucleotides, each consisting of at least two elements of the -6.1-kb enhancer, confer myeloid-cell-specific enhancer activity. We found several myeloid-cell-specific DNA-binding proteins interacting with the -6.1-kb enhancer, a result consistent with that described above. Therefore, we suggest that more than a single trans-acting factor mediates the cell type specificity of the -6.1-kb enhancer.
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PMID:The -6.1-kilobase chicken lysozyme enhancer is a multifactorial complex containing several cell-type-specific elements. 156 54

Lysozyme gene expression is a specific marker for the macrophage/granulocyte lineage of hematopoietic differentiation in mammals, its expression being gradually increased during maturation. Analysis of the mechanisms regulating mouse M lysozyme gene expression during myeloid differentiation revealed a complicated pattern of DNase I hypersensitive sites (HS sites) within the flanking regions of the gene. The HS-3 site, located in the 3'-flanking region of the gene, overlapped with an enhancer element, which is the only strong enhancer identified in the vicinity of the gene. We demonstrate a positive correlation between undermethylation of the entire 3'-flanking region, the appearance of the HS-3 site, and M lysozyme gene expression during in vitro differentiation of hematopoietic stem cells. We furthermore show that methylation of a single CpG site within the enhancer core element, only observed in immature macrophage cells in vivo, is sufficient to inhibit nuclear factor binding to this element in vitro and to inhibit its transactivation potential in DNA transfection experiments.
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PMID:The involvement of demethylation in the myeloid-specific function of the mouse M lysozyme gene downstream enhancer. 157 94

Quantitative analysis of distribution of chromosomal proteins on single copy DNA sequences has been further developed. Our approach consists of DNA-protein crosslinking within whole cells or isolated nuclei, specific immunoaffinity isolation of crosslinked complexes via protein and identification of crosslinked DNA by hybridisation with single-stranded DNA probes. The present study shows that transcribed chromatin of chicken embryonic erythrocyte beta globin gene is characterized by about 1.5-2.5-fold higher density of HMG 14/17 and 2-fold lower density of H1 and H5 as compared with non-transcribed chromatin of ovalbumin and lysozyme genes, whereas HMG 1/2, E proteins were equally distributed between DNA of both transcribed and non-transcribed genes. The depletion of H1/H5 in beta globin sequences was verified by the 'protein image' hybridisation technique (1). The DNase I hypersensitive site located 5' upstream from beta globin gene is deficient in all the proteins assayed, what implies a drastic disruption in the nucleosomal array. Minor quantitative changes of protein pattern suggest transient local perturbation of the chromatin on transcription.
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PMID:Distribution of high mobility group proteins 1/2, E and 14/17 and linker histones H1 and H5 on transcribed and non-transcribed regions of chicken erythrocyte chromatin. 201 59

We identified the earliest events in autophosphorylation of the insulin receptor after insulin addition. Insulin-stimulated autophosphorylation at specific sites in the tyrosine kinase domain of the receptor's beta-subunit is correlated kinetically with activation of kinase-catalyzed phosphorylation of a model substrate (reduced and carboxyamidomethylated lysozyme; RCAM-lysozyme). To identify these sites, the deduced amino acid sequence of the 3T3-L1 adipocyte insulin receptor of the mouse was determined. Insulin-induced activation of substrate phosphorylation was shown to require autophosphorylation of three neighboring tyrosines (Tyr1148, Tyr1152, and Tyr1153) in the mouse receptor. A search for cellular substrates of the receptor kinase revealed that insulin causes accumulation of a 15,000-Mr phosphorylated (on tyrosine) cytosolic protein (pp15) in 3T3-L1 adipocytes treated with oxophenylarsine (PAO). PAO blocks turnover of the phosphoryl group of pp15, causing its accumulation, and thereby appears to interrupt signal transmission from the receptor to the glucose-transport system. Two membrane-bound protein phosphotyrosine phosphatases that are inhibited by PAO and are apparently responsible for the turnover of the pp15 phosphoryl group have been purified from 3T3-L1 adipocytes and characterized. These and other results support the hypothesis that turnover of the phosphoryl group of pp15, a product of insulin-receptor tyrosine kinase action, couples signal transmission to the glucose-transport system. [32P]pp15 was purified to homogeneity from 3T3-L1 adipocytes. Amino acid and radiochemical sequence analysis of the purified tryptic [32P]phosphopeptide revealed that pp15 is the phosphorylation product of 422(aP2) protein, a 15,000-Mr adipocyte protein whose cDNA we previously cloned and sequenced. 422(aP2) protein was found to bind fatty acids. When exposed to a free fatty acid, notably oleic acid, 422(aP2) protein becomes an excellent substrate of the isolated insulin-receptor tyrosine kinase. Compelling evidence indicates that on binding fatty acid, 422(aP2) protein undergoes a conformational change whereby Tyr19 becomes accessible to the receptor tyrosine kinase and undergoes O-phosphorylation. Adipose tissue and skeletal and heart muscle, which exhibit insulin-stimulated glucose uptake, express a specific insulin-responsive glucose transporter. A cDNA (GT2) that encodes this protein was isolated from a mouse 3T3-L1 adipocyte library and sequenced. We also isolated and characterized the corresponding mouse gene GLUT4. DNase I footprinting with nuclear extracts from 3T3-L1 cells revealed that a differentiation-specific nuclear factor binds to the GLUT4 promoter. The purified transcription factor C/EBP binds at the same position.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Insulin-receptor tyrosine kinase and glucose transport. 216 54

Vimentin is one member of the intermediate filament multigene family which exhibits both tissue- and developmental stage-specific expression. In vivo, vimentin is expressed in cells of mesenchymal origin. Previously, we identified both enhancer and promoter elements in the chicken vimentin gene which regulate gene expression in a positive manner. In this report, we have identified a 40-base-pair region at -568 base pairs between the proximal and distal enhancer elements which represses transcriptional activity. This silencer region can also repress the heterologous herpes simplex virus thymidine kinase promoter, which is comparable to the vimentin promoter. In addition, the element is able to function in a position- and orientation-independent manner, and the amount of repression is increased by multiple copies. Here we show by gel retardation assays and DNase I footprinting that this region binds a protein in nuclear extracts from HeLa cells. Southwestern (DNA-protein) blot analysis indicates this protein is approximately 95 kilodaltons in size. Moreover, protein distribution and activity mimic the expression pattern of vimentin during myogenesis, i.e., protein binding increases as vimentin gene expression decreases. The silencer region shares strong sequence similarity with 5'-flanking sequences found in both the human and hamster vimentin genes and with other characterized silencer elements, including the human immunodeficiency virus long terminal repeat, rat growth hormone, chicken lysozyme, and rat insulin genes. Thus, a negative element appears to bind a 95-kilodalton protein involved in regulating the tissue-specific expression of the chicken vimentin gene.
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PMID:A negative element involved in vimentin gene expression. 232 56

A 21.5 kb DNA fragment carrying the entire chicken lysozyme gene locus was introduced into the germ line of mice. The fragment contains the transcribed region plus 11.5 kb 5'-flanking and 5.5 kb 3'-flanking sequences including all known cis-regulatory elements and the 5' and 3' attachment elements (A-elements) which define the borders of the DNase I sensitive chromatin domain. All sequences which adopt a DNase I hypersensitive chromatin conformation in vivo are present on the construct. Seven founder mice were analysed. All of these expressed chicken lysozyme RNA at high levels specifically in macrophages, as is the case in the donor species. Expression levels are dependent on the copy number of integrated genes indicating that a complete gene locus, as defined by its chromatin structure, functions as an independent regulatory unit when introduced into a heterologous genome.
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PMID:Tissue specific and position independent expression of the complete gene domain for chicken lysozyme in transgenic mice. 239 Sep 72

We have investigated the influence of the 5'-flanking region of the chicken lysozyme gene on steroid dependent gene expression. By transient transfection of lysozyme-CAT fusion genes into the human breast cancer cell line T-47D, a DNA element was identified which stimulates CAT expression when transfected cells are treated with progesterone. This element is distinct from a second hormone responsive element (HRE) located in the lysozyme promoter region; it activates the lysozyme and the TK promoter, irrespective of orientation and distance, and is therefore referred to as hormone responsive element on its own. The location of this newly discovered HRE between -2250 and -1815 relative to the transcriptional start site, corresponds to the position of a steroid inducible DNase I-hypersensitive site in chromatin of oviduct cells. This observation suggests a physiological role for the upstream element. In vitro DNase I protection experiments revealed six binding sites for both progesterone and glucocorticoid receptors within the sequences of the upstream HRE. The three distal binding sites are not required for hormonal stimulation of the TK promoter, while the three proximal binding sites, which are contiguously arranged, work in a cooperative manner.
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PMID:A progesterone responsive element maps to the far upstream steroid dependent DNase hypersensitive site of chicken lysozyme chromatin. 341 33

The chicken lysozyme gene can be induced in oviduct cells by four classes of steroid hormones, including glucocorticosteroids and progestins. The glucocorticosteroid receptor of rat liver and the progesterone receptor of rabbit uterus both bind, although with different relative affinities, to two sites in the promoter region of the chicken lysozyme gene located, respectively, between 50 and 80 and between 160 and 200 base pairs upstream of the transcription start point. Now we show that the purified progesterone binding unit of the chicken oviduct progesterone receptor (Mr 110,000, or so-called B subunit) generates a DNase I protection pattern ("footprint") in the promoter-distal site that is longer than the footprint generated by the glucocorticosteroid receptor. Methylation protection studies within the promoter-distal binding site identify four contact points for the chicken progesterone receptor and three contact points for the glucocorticosteroid receptor, of which only one is shared by both receptors. Computer graphics models allow one to envisage a different interaction of each receptor with the B form of the DNA double helix.
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PMID:Receptors for glucocorticosteroid and progesterone recognize distinct features of a DNA regulatory element. 345 41

The pattern of DNase I-hypersensitive sites in the chromatin domain of the lysozyme gene was investigated in several organs and cell-types of the chicken. In the cluster of hypersensitive chromatin sites framing the gene, different classes of sites could be discerned: A subset was common to essentially all cells examined except for erythrocytes. Thus several highly nuclease susceptible structures exist around the gene even in its repressed state. Beside the promoter region a second site 6.1 kb upstream of the transcriptional start site of the gene strictly correlates with the transcriptionally active or potentially active state of the gene in both, oviduct cells and macrophages. A final class of sites is specific for the particular lysozyme expressing tissue and the presence of its members distinguish whether the gene is steroid regulated or in a steroid independent active mode.
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PMID:DNase I-hypersensitive sites in the chromatin structure of the lysozyme gene in steroid hormone target and non-target cells. 356 13


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