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:3.1.3.1 (alkaline phosphatase)
47,916 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

1. Homogenates of the mucosa of the small intestine of the guinea pig were separated by fractional sedimentation into seven different fractions. The enzymic properties of some of these subcellular fractions were compared with those obtained from the mucosa of the small intestine of the rabbit and cat. 2. The enzymic properties of the low-speed sediment (15000g-min.) were investigated and it was shown that invertase and alkaline ribonuclease were predominantly located in this subcellular fraction, whereas alkaline phosphatase, aryl-amidase, acid phosphatase, acid ribonuclease and phosphoprotein phosphatase, though true constituents of this fraction, occurred to varying degrees in other subcellular structures also. 3. It was shown that the most probable source of the enzymic activities observed in the low-speed sediment was the brush border. Electron micrographs of the purified brush-border fraction indicated vesicles derived from the brush-border membrane. 4. A method is described for the fractionation of mucosal homogenates into a brush border-plus-nuclei fraction, a mitochondrial fraction, a microsomal fraction and a particle-free supernatant. The fractions were shown to be relatively pure, as indicated by the distribution of invertase, DNA, succinate dehydrogenase, glucose 6-phosphatase and 6-phosphogluconate dehydrogenase. 5. Most of the activity of four lysosomal enzymes present in the nuclei-free homogenate was sedimented at 375000g-min., suggesting the occurrence of lysosomal particles in mucosal homogenates. 6. Further fractionation of the microsomal membranes into three fractions is described. The enzymic composition of the membrane fractions is given and discussed in relation to their structure as seen in electron micrographs.
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PMID:Studies on the fractionation of mucosal homogenates from the small intestine. 428 74

The role of phosphatidylinositol-specific phospholipase C (PIase C) in a) the enigmatic phosphatidylinositol (PI) turnover and b) in our understanding of membrane enzyme-PI interactions is the subject matter of this article. PIase C is present in both procaryotes and eukaryotes. This enzyme is considered to be involved in the cells PI breakdown which occurs in response to several external stimuli. Recent information on the physical properties, Ca2+ requirement, cellular localization and modulation of the activity of PIase C of mammalian systems can help to evaluate the PI turnover from a new angle. Existing evidence suggests that Ca2+-dependent PI breakdown is probably mediated through the cytosolic and particulate PIase C while a Ca2+ independent pathway is catalyzed by a lysosomal enzyme. Apparently PI turnover may be operating through more than one mechanism. The association of this phenomenon with a membrane receptor event linked with "Ca2+ gating" may have to be reconsidered. Modulation of the PIase C activity by unsaturated amphiphiles or the presence of this enzyme in different physico-chemical forms could be a potential regulatory feature. Hydrolysis of membrane PI of a number of cells and tissues by the bacterial PIase C has been shown to cause substantial release of acetylcholinesterase, alkaline phosphatase and 5'-nucleotidase in free, soluble form. Other membrane enzymes, e.g., alkaline phosphodiesterase I, L-leucyl-beta naphthyl amidase and Ca2+ or Mg2+ ATPase are not affected. These results indicate a specific interaction between PI and certain enzymes in membranes. The chemical nature of this linkage, whether it is covalent or non-covalent, has also been explored and has provided intriguing insight into this phenomenon. New findings also indicate that hydrolysis of PI by PIase C also can cause modifications in membrane-enzyme activities, e.g., adenylate cyclase.
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PMID:Minireview. Phosphatidylinositol specific phospholipases C. 708 67

A soil incubation experiment was carried out in a complex experimental system to simulate the effect of environmental factors and 3 trace elements on the phosphomonoesterase and amidase activity of 2 soils. DISITOBI model assures information about experimental object characterized by multidimensional response-function. Plant residue+mineral nitrogen addition increased phosphomonoesterase activity of investigated soils through its effect on enzyme synthesis. A significant part of the increase in phosphomonoesterase activity is the result of the exogenous enzyme input of plant residues. Mineral phosphorus addition reduced the activity after 16 days incubation under our experimental conditions. Plant residue+mineral nitrogen addition reduced amidase activity of chernozem soil presumably due to ammonium and nitrate inhibiting soil amidases.
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PMID:Effects of environmental factors and Mn, Zn, Cu trace elements on the soil phosphomonoesterase and amidase activity. Application of DISITOBI model. 774 Aug 99

Major histocompatibility (MHC) class II antigens are heterodimeric cell surface glycoproteins consisting of an alpha and a beta chain. Although one-dimensional SDS-polyacrylamide gel electrophoresis analysis of purified MHC class II antigens shows a single diffuse band for each chain, multiple spots of identical molecular size were observed for each chain when analyzed by two-dimensional electrophoresis. The basis of this heterogeneity has not been clearly defined and has been predicted partially to be due to glycosylation and/or phosphorylation of the mature protein. To investigate the role of the three N-linked oligosaccharides of the alpha and beta chains in determining the isoelectric point of each chain, affinity-purified MHC class II antigens from human and rat sources were deglycosylated using asparagine amidase. The complete enzymatic removal of all three N-linked oligosaccharides was confirmed by SDS-polyacrylamide gel electrophoresis as well as by four different lectin-linked Western blot analyses. Two-dimensional gel analysis of the deglycosylated molecules shows no significant difference from the fully glycosylated chains. We have expressed truncated forms of the HLA DR2 chains which lack the transmembrane and cytoplasmically exposed regions in Escherichia coli. Two-dimensional electrophoresis of these single chains also reveal multiple banding patterns. The two-dimensional banding patterns described are unaffected by exposure to acidic or basic conditions, increased gel running time in the first dimension, treatment of the proteins with alkaline phosphatase to remove any potential phosphorylation, or preincubation in the presence of iodoacetamide. Multiple forms of recombinant alpha and beta chains were also observed in Tris-glycine-urea gels which merged into a single band in the presence of SDS. In addition, partially fractionated bands from preparative isoelectric focusing gels, when refocused, showed an identical number of multiple spots spanning the same range of isoelectric points. These results together suggest that each polypeptide chain of MHC class II antigens may exist in multiconformational forms, and the observed charge heterogeneity is independent of glycosylation and phosphorylation of the proteins.
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PMID:Intramolecular charge heterogeneity in purified major histocompatibility class II alpha and beta polypeptide chains. 814 5

The syntheses and cytotoxic activities of substituted N-phenylacetamido derivatives of doxorubicin and melphalan are described. The derivatives were designed as prodrugs which could be activated in a site-specific manner by monoclonal antibody-penicillin-G amidase (mAb-PGA) conjugates. N-(Phenylacetamido)doxorubicin (2) and N-(phenylacetyl)melphalan (6) were found to be 10- and 20-fold less cytotoxic against H2981 lung adenocarcinoma cells than doxorubicin and melphalan, respectively. When incubated with PGA, the cytotoxicity of 2 and 6 increased and became equivalent to that of the corresponding drugs from which they were made. The poor solubility characteristics of 2 in aqueous solutions provided the basis for the development of the more soluble doxorubicin derivatives, N-(4-aminophenylacetyl)doxorubicin (3) and N-(4-phosphonooxy)phenylacetyl)-doxorubicin (4). In vitro cytotoxicity assays indicated that 3 and 4 were at least 1000-fold less toxic than doxorubicin against H2981 cells. PGA and the mAb conjugate L6-PGA were able to effect the activation of 3 and 6 on H2981 cells (L6-antigen positive). Hydrolysis of the phosphate group of 4 was required prior to activation with PGA or L6-PGA. This was achieved using alkaline phosphatase, or by exposing 4 to phosphatases present in cell culture medium. The activation of 3, 4, and 6 on H2981 cells by L6-PGA occurred in an immunologically specific manner, since activation could be blocked by saturating cell surface antigens with L6 prior to treatment with L6-PGA. These results demonstrate that 3, 4, and 6 are prodrugs that can be specifically activated to release clinically approved anticancer agents by a mAb-PGA conjugate.
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PMID:Prodrugs of doxorubicin and melphalan and their activation by a monoclonal antibody-penicillin-G amidase conjugate. 846 46

We have developed a method for monitoring the N-glycosylation of recombinant glycoproteins directly from conditioned medium samples. Proteins in the conditioned medium are separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and electroblotted onto polyvinylidene fluoride membranes. After staining the membranes with Coomassie blue, the protein(s) of interest is excised. Oligosaccharides are released from the membrane-bound glycoprotein by digesting with peptide N4-(acetyl-beta-glucosaminyl) asparagine amidase and labeled with the fluorophore 8-aminonaphthalene-1,3,6-trisulfonate (ANTS). Labeled oligosaccharides are then separated on polyacrylamide gels which allow for the direct comparison of samples. We have shown that recombinant human lysosomal hydrolase alpha-galactosidase A is N-glycosylated with both sialylated and phosphorylated oligosaccharides. ANTS-labeled oligosaccharide bands from alpha-galactosidase A were isolated from polyacrylamide gels. Sialylated and phosphorylated bands were identified by shifts in their electrophoretic mobility after digesting with neuraminidase or alkaline phosphatase to remove sialic acid or phosphate groups, respectively. Using the ANTS-labeled oligosaccharides from alpha-galactosidase A, we have shown that polyacrylamide gels can be used to resolve sialylated and phosphorylated oligosaccharide structures.
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PMID:A method for monitoring the glycosylation of recombinant glycoproteins from conditioned medium, using fluorophore-assisted carbohydrate electrophoresis. 857 98

The N-linked glycans assembled in Pichia pastoris on the recombinant kringle 2 domain of human tissue-type plasminogen activator (r-[K2tPA]) are composed of approx. 80% neutral and 20% charged species. After peptide:N4-(N-acetyl-beta-glucosaminyl)asparaginyl amidase-catalysed liberation of the oligosaccharides from the purified glycopeptide, the glycan mixture was resolved by HPLC on amino-silica-based resin. Oligosaccharide mapping of the resulting mixture by HPLC, gel filtration and time-of-flight matrix-assisted laser-desorption-ionization-with-delayed-extraction mass spectrometry (TOF-MALDI DE-MS) revealed that > 90% of the charged species consisted of a series of oligosaccharides possessing molecular masses that were consistent with a range of saccharides comprising phospho-Man10GlcNAc2-phospho-Man14GlcNAc2, with phospho-Man11GlcNAc2 representing the major species. The remaining material in the charged fraction contained identifiable phosphorylated glycans that were one or two mannose units shorter, and one to four mannose units longer, than those present in the above range of oligosaccharides. Treatment of the native charged glycan pool with alkaline phosphatase did not result in molecular-size alterations, showing that phosphomonoesters are not present. Mild acid hydrolysis of the glycans led to a decrease in the size of all charged glycans by one mannose residue, providing phospho-Man9GlcNAc2-phospho-Man13GlcNAc2. Following this procedure, treatment with alkaline phosphatase resulted in size decreases that were equivalent to the loss of one phosphate group from each glycan. This demonstrates that all charged glycans isolated contained phosphate in phosphodiester bonds to two mannose units. The present study shows that P. pastoris cells possess the capability of assembling phosphorylated glycans having the phosphate moiety present in phosphodiester linkages with two mannose units. These saccharides, like the neutral oligosaccharides, contain considerably smaller amounts of mannose than glycans present in other strains of yeast.
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PMID:Characterization of the acidic oligosaccharides assembled on the Pichia pastoris-expressed recombinant kringle 2 domain of human tissue-type plasminogen activator. 935 3

The present study was made to isolate and assess some physiological characteristics of root nodule-colonizing fungi. During this study, 17 fungal species were isolated from root nodule samples taken from faba bean plants (Vicia faba L.) collected from different sites at Assiut area (Egypt). The growth of faba bean plants in pots was significantly promoted by soil inoculation with most fungi. Growth was checked in pots with inocula of Cladosporium cladosporioides, Fusarium moniliforme, F: oxysporium, F solani, Macrophominia phaseolina and Rhizoctonia solani which were added separately. All growth-promoting fungi were capable of producing cellulase, pectin lyase, polygalacturonase, protease, urease, amidase, acid phosphatase, alkaline phosphatase and arylsulfatase in growth medium supplemented with the corresponding substrates. Four fungal species, Aspergillus awamori, A. flavus, Penicillium chrysogenum and Trichoderma koningii showed the highest rates of enzyme formation. The effect of the addition of six trace elements to the growth media at 30 micromol/ml on enzyme production revealed some dependency on species, enzyme and metal ion. Cd2+, Hg2+ and Zn2+ generally inhibited enzyme activity. Cu(1+), Fe3+ and Al3+ showed a stimulatory effect. Fungicides (afugan and tilt) and herbicides (brominal and fusilade) at 50 ppm generally promoted enzyme activity, but insecticides (kelthane and fenvalerate) caused some inhibition to enzyme activities. Salinization of the growth media with NaCl strongly inhibited the enzymatic activity of all fungi at concentrations between 0.5 and 1.5%.
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PMID:Physiological aspects of fungi isolated from root nodules of faba bean (Vicia faba L.). 1077 56

Light's criteria have classically been used to differentiate exudates from transudates. Nevertheless, a number of studies have attempted to identify more efficient parameters. The objective of our study was to determine the usefulness of biochemical parameters to differentiate transudates from exudates, and to compare them with the so far best studied criteria: the Light's criteria. We prospectively analysed 850 non selected cases of pleural effusion, with closed final diagnosis after its confirmation, therapeutic response and follow-up, collected consecutively at the Pleura Unit of our hospital. The parameters evaluated as potentially discriminatory between transudates and exudates included: glucose, proteins, albumin, lactate-dehydrogenase (LDH), cholesterol, triglycerides, bilirubin, alkaline phosphatase and adenosin-deaminase (ADA), both separately and in combination to obtain the highest yield. The highest diagnostic yield was observed with the combination of pleural cholesterol, pleural LDH, and the pleural fluid/serum protein ratio, but without significant differences between combinations of pleural cholesterol and LDH, pleaural LDH and pleural proteins, Light's criteria or modified Light's criteria. We recommend the use of pleural cholesterol higher than 47 mg/dl and pleural LDH higher than 222 IU/l to offer the same yield as the combination of three parameters, due to its lower cost and because the necessity of serum determinations is avoided.
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PMID:[Comparative analysis of Light's criteria and other biochemical parameters to distinguish exudates from transudates]. 1194 Apr 25

Extracts of Aspergillus niger NRRL3 catalyzed dephosphorylation of AMP, GMP, CMP and UMP over a wide range of pH values from pH 1.5 to pH 10. They also catalyzed hydrolytic deamination of only cytidine out of the tested ribonucleotides, ribonucleosides and bases. Neither cleavage of the N-glycosidic linkages of these nucleotides nor those of the corresponding nucleosides could be effected by the extracts. Phosphate liberation from the four RNA monomers seemed to be effected by two phosphate-non repressible phosphatases, acid and alkaline. Optimum activity of the acid phosphatase with all the substrates was at pH2 and 40 degrees C while that of the alkaline phosphatase was at pH8 and 40 degrees-70 degrees C. Affinities of both phosphatases for the different ribonucleotides were in the order of magnitude AMP, CMP and phph > GMP > UMP. Freezing and thawing of the extracts had no effect either on the activities of two phosphatases or on that of the aminohydrolase. However, heating the extracts at 55 degrees for 25 min, in absence of the substrate, inactivated the phosphatases and had no effect on the deaminase. No evidence for the involvement of specific nucleotidases in ribonucleotides dephosphorylation was recorded.
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PMID:Hydrolysis of RNA monomers by extracts of Aspergillus niger NRRL3. 1518 88


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