Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.4.3 (phospholipase C)
 18,461 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In Pseudomonas aeruginosa, the genes pilB, pilC, and pilD encode proteins necessary for posttranslational modification and assembly of pilin monomers into pilus organelles (D. Nunn, S. Bergman, and S. Lory, J. Bacteriol. 172:2911-2919, 1990). We show that PilD, encoding a putative pilin-specific leader peptidase, also controls export of alkaline phosphatase, phospholipase C, elastase, and exotoxin A. pilD mutants accumulate these proteins in the periplasmic space, while secretion of periplasmic and outer membrane proteins appears to be normal. The periplasmic form of exotoxin A was fully mature in size, contained all cysteines in disulfide bonds, and was toxic in a tissue culture cytotoxicity assay, suggesting that in pilD mutants, exotoxin A was folded into its native conformation. The function of the other two accessory proteins, PilB and PilC, appears to be restricted to pilus biogenesis, and strains carrying mutations in their respective genes do not show an export defect. These studies show that in addition to cleaving the leader sequence from prepilin, PilD has an additional role in secretion of proteins that are released from P. aeruginosa into the surrounding media. PilD most likely functions as a protease that is involved in processing and assembly of one or more components of the membrane machinery necessary for the later stages of protein extracellular localization.
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PMID:Multiple roles of the pilus biogenesis protein pilD: involvement of pilD in excretion of enzymes from Pseudomonas aeruginosa. 167 84

Surface charge of Leishmania mexicana amazonensis was investigated by direct zeta-potential determination and ultrastructural cytochemistry, and its surface tension was studied by measurements of the advancing contact angle formed by the parasite monolayers with drops of liquids of different polarities. Both virulent and avirulent promastigotes exhibited negatively charged surfaces with a zeta-potential of about -15 mV. Treatment of these cells with trypsin, alkaline phosphatase, or phospholipase C rendered their surfaces less negatively charged, whereas neuraminidase did not alter the parasite negativeness. Cytochemically, we could observe a reduction in the cationized ferritin binding after the parasite treatment with each of the former enzymes, but not with neuraminidase. The surface free energy of parasites was calculated by taken to account the London dispersion, the Keeson dipole-dipole, and the Debye dipole-induced forces, as well as the surface polarity of the parasites and their zeta-potentials, by considering their adhesion to polystyrene surfaces. The delta G values of -6.4 and -18.1 mJ.m-2 were obtained for avirulent and virulent promstigotes, respectively.
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PMID:The surface free energy of Leishmania mexicana amazonensis. 170 80

The number of identifiable stages and expression of differentiation markers in cells of the osteoblast lineage are not well understood. In the present study, a mAb, designated rat bone marrow (RBM) 211.13, was prepared that stained selectively the osteogenic and preosteoblastic cells along the surfaces of bone in calvariae, femurs, and metatarsals. The staining was cell surface associated and coincided with that for alkaline phosphatase (APase) detected histochemically. Only cells positive for APase activity by biochemical assay and not those without APase activity (e.g., fetal rat skin) stained with RBM 211.13. By immunoblotting, RBM 211.13 recognized a band coinciding with APase activity on nonreducing/nondenaturing gels, and RBM 211.13 precipitated a protein which on reduced gels migrated with an apparent molecular mass of approximately 80 kD. RBM 211.13 labeling was abolished by phosphatidylinosital-specific phospholipase C, known to release APase from the cell surface. All of these data support the concept that RBM 211.13 recognizes the bone isoenzyme of APase. RBM 211.13 was used to sort by flow cytometry the APase-positive and APase-negative cells from mixed fetal rat calvaria (RC) cell populations. The osteoprogenitors we identified earlier that form bone nodules in vitro (Bellows, C. G., J. E. Aubin, J. N. M. Heersche, and M. E. Antosz. 1986. Calcif. Tissue Int. 36:143-154; Bellows, C. J., J. N. M. Heersche, and J. E. Aubin. 1990. Dev. Biol. 140:132-138) were found within the APase-positive pool. By immunopanning, RC cells were separated into APase-enriched (APase-positive, adherent) and APase-depleted (APase-negative, nonadherent) populations. The APase-positive fraction was enriched two-to-threefold for bone-forming osteoprogenitors compared to unfractionated cells, while the APase-negative population formed very few nodules under the same conditions. Both populations responded to the glucocorticoid dexamethasone (DEX) with an increase in bone nodule formation. However, the fold stimulation in bone formation in the APase-negative population was approximately 30-fold, while the fold stimulation in the APase-positive population was only approximately 5-fold. These data suggest that APase expression can be used for immunoselection to fractionate osteoblastic populations into an APase-positive population and a population initially APase-negative, that virtually all osteoprogenitors forming bone in vitro in the absence of added glucocorticoids reside in the APase-positive pool, and that the only osteoprogenitors present in the APase-negative pool are those requiring DEX to differentiate.
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PMID:Positive and negative immunoselection for enrichment of two classes of osteoprogenitor cells. 171 92

The antigen detected by monoclonal antibodies reacting with human osteosarcoma-associated antigen was shown to be a phosphatidyl-inositol (PI)-glycan-anchored protein, which can be released from the cell surface by PI-specific phospholipase C-treatment. The antigen detected by 2D3 and 2H10 antibodies exhibited alkaline phosphatase activity. Both antibodies strongly reacted with bone-type alkaline phosphatase. However, importantly, immunohistochemical analysis demonstrated that 2D3 and 2H10 did not react with alkaline phosphatase present in kidney or liver. In addition, neither placental nor intestinal alkaline phosphatase was recognized by 2D3 and 2H10 antibodies. These results indicated that two monoclonal antibodies, 2D3 and 2H10, are highly specific for bone-type alkaline phosphatase and can distinguish bone alkaline phosphatase from liver alkaline phosphatase in spite of the fact that liver and bone alkaline phosphatase are encoded by the same gene.
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PMID:Detection of bone-type alkaline phosphatase by monoclonal antibodies reacting with human osteosarcoma-associated antigen. 171 40

We have isolated and characterized four toxin A excretion-deficient mutants of Pseudomonas aeruginosa PAO1. Similar to previously described mutants (B. Wretlind and O. R. Pavlovskis, J. Bacteriol. 158:801-808, 1984), the mutants appear to have a pleiotropic defect in the excretion of several extracellular products, including toxin A, elastase, alkaline phosphatase, and phospholipase C. However, the mutants are not defective in the excretion of either alkaline protease or exoenzyme S. We also examined the localization and processing of toxin A in these mutants by using pulse-labeling experiments. Mature toxin A was found to be localized to the membranes only. Our results suggest that toxin A is localized to the outer membrane but is not exposed to the extracellular surfaces of the outer membranes. The results also suggest that toxin A obtained from the excretion-deficient mutants has intact disulfide bonds.
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PMID:Isolation and characterization of toxin A excretion-deficient mutants of Pseudomonas aeruginosa PAO1. 173 Apr 83

Subcellular fractionation of pig kidney cortex revealed that aminoacylase I (EC 3.5.1.14, N-acyl-L-amino-acid aminohydrolase) is predominantly a soluble enzyme with only 0.5% of the total activity being recovered in the membrane fraction. The aminoacylase I activity associated with the membrane preparations displayed neither rapid release following incubation with phosphatidylinositol-specific phospholipase C from Bacillus thuringiensis nor the distinctive differential pattern of detergent solubilization which was seen with glycosyl-phosphatidylinositol-anchored proteins (renal dipeptidase, alkaline phosphatase). When fractionated by phase separation in Triton X-114, integral membrane proteins of kidney microvillar membranes partitioned predominantly (greater than 90%) into the detergent-rich phase. In contrast, only 3.7% of aminoacylase I activity associated with microvillar membranes partitioned into the detergent-rich phase. Aminoacylase I activity of pig kidney would therefore appear to be a hydrophilic protein in nature and is not, as suggested previously, a G-PI-anchored integral membrane protein.
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PMID:Aminoacylase I is not a glycolipid-anchored ectoenzyme in pig kidney. 182 88

The molecular species of 1,2-diacyl-sn-glycerol (DAG), phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylserine (PS), phosphatidylinositol (PI), phosphatidylinositol 4-phosphate (PIP), and phosphatidylinositol 4,5-bisphosphate (PIP2) from brains of adult rats (weighing 150 g) were determined. The DAG, isolated from brain lipid extracts by TLC, was benzoylated, and the molecular species of the purified benzoylated derivatives were separated from each other by reverse-phase HPLC. The total amount and the concentration of each species were quantified by using 1,2-distearoyl-sn-glycerol (18:0-18:0) as an internal standard. About 30 different molecular species containing different fatty acids at the sn-1 and sn-2 positions of DAG were identified in rat brains (1 min postmortem), and the predominant ones were 18:0-20:4 (35%), 16:0-18:1 (15%), 16:0-16:0 (9%), and 16:0-20:4 (8%). The molecular species of PC, PE, PS, and PI were determined by hydrolyzing the lipids with phospholipase C to DAG, which was then benzoylated and subjected to reverse-phase HPLC. PIP and PIP2 were first dephosphorylated to PI with alkaline phosphatase before hydrolysis by phospholipase C. The molecular species composition of phosphoinositides showed predominantly the 18:0-20:4 species (50% in PI and approximately 65% in PIP and PIP2). PS contained mainly the 18:0-22:6 (42%) and 18:0-18:1 (24%) species. PE was mainly composed of the 18:0-20:4 (22%), 18:0-22:6 (18%), 16:0-18:1 (15%), and 18:0-18:1 (15%) species. In PC the main molecular species were 16:0-18:1 (36%), 16:0-16:0 (19%), and 18:0-18:1 (14%). Studies on postmortem brains (30 s to 30 min) showed a rapid increase in the total amount (from 40-50 nmol/g in 0 min to 210-290 nmol/g in 30 min) and in all the molecular species of DAG. Comparatively larger increases (seven- to 10-fold) were found for the 18:0-20:4 and 16:0-20:4 species. Comparison of DAG species with the molecular species of different glycerolipids indicated that the rapid postmortem increase in content of DAG was mainly due to the breakdown of phosphoinositides. However, a slow but continuous breakdown of PC to DAG was also observed.
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PMID:Molecular species of diacylglycerols and phosphoglycerides and the postmortem changes in the molecular species of diacylglycerols in rat brains. 184 96

We have previously shown that two ectoenzymes, acetylcholinesterase (AChE) and alkaline phosphatase, are released from the surface and from particulate fractions of the parasite Schistosoma mansoni, by a phosphatidylinositol-specific phospholipase C (PtdIns-PLC) of bacterial origin. Exposure to PtdIns-PLC not only removes large amounts of AChE from the surface of intact, viable Schistosoma in culture, but is accompanied by a concomitant increase in overall levels of AChE in the parasite. The same phenomenon is observed with PtdIns-PLC from two different bacterial sources; Staphylococcus aureus and Bacillus thuringiensis. The increase in AChE levels may be ascribed to de novo synthesis since exposure to PtdIns-PLC, in the presence of the protein-synthesis inhibitor cycloheximide, totally blocked the increase in AChE activity. Furthermore, PtdIns-PLC induced an increased incorporation of [35S]methionine into the AChE immunoprecipitated by a specific anti-AChE serum. This increase is selective for AChE, since total protein synthesis remained almost unchanged after PtdIns-PLC addition, and little or no effect was observed on the enzymatic activity of alkaline phosphatase, which is also glycophosphatidylinositol anchored. Since cleavage of the phosphatidylinositol anchor by PtdIns-PLC should liberate diacylglycerol, which may act as second messenger, we investigated the effect of exogenous diacylglycerols on the synthesis of AChE in S. mansoni. Three different diacylglycerols were tested as possible inducers of AChE activity in the parasite. Both 1-oleoyl-2-acetyl-sn-glycerol and 1,2-dimyristoyl-sn-glycerol were able to increase AChE activity by 35-40% at concentrations of 25 micrograms/ml. A higher concentration of 1,2-dioctanoyl-sn-glycerol (70 micrograms/ml) was needed to produce an equivalent effect. Moreover, addition of phorbol-12-myristate-13-acetate, together with the calcium ionophore A23187, produced a similar increase in AChE activity. Finally, polymixin B, a specific inhibitor of protein kinase C, partially blocked the increase in AChE activity induced by PtdIns-PLC. Our results suggest the involvement of glycophosphatidyl membrane-anchor breakdown products as putative second messengers in the parasite S. mansoni.
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PMID:Phosphatidylinositol-specific phospholipase C induces biosynthesis of acetylcholinesterase via diacylglycerol in Schistosoma mansoni. 184 73

Plasmids encoding mercury resistance carried by Pseudomonas aeruginosa PAO1161 and PA103 were found to be involved in regulating the secretion of protease, phospholipase C, and alkaline phosphatase. Previously, mutations in Pseudomonas strains that caused pleiotropic effects on the production of extracellular enzymes were mapped to the bacterial chromosome. We show that pleiotropic changes in extracellular enzyme production can also be regulated by plasmids. In this study, the effects on secretion of exoenzymes by two mercury resistance plasmids, FP2 from PAO1161 and pRLW103 from PA103, were assayed in P. aeruginosa PAO1 and PAO18. The introduction of either plasmid into PAO1 resulted in a significant decrease in exoprotease production. Additionally, pRLW103 significantly increased the production of alkaline phosphatase by both strains. Phospholipase C was produced only in strain PAO18 containing the pRLW103 plasmid. FP2 had no effect on alkaline phosphatase or phospholipase C production in either strain and was found to decrease exoprotease secretion only in strain PAO1. The results indicate the P. aeruginosa mercury resistance plasmids vary in their ability to modify exoenzyme expression, and this ability is influenced by the host strain.
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PMID:Effects of FP2 and a mercury resistance plasmid from Pseudomonas aeruginosa PA103 on exoenzyme production. 190 22

The molecular nature and possible presence of a glycan-phosphatidylinositol anchor (GPI-anchor) in CA125 molecules was investigated. Serial lectin affinity chromatography and N- or O-glycanase treatment to reduce antigenicity showed that CA125 contained certain N- and O-glycosylated sugar chains in the molecule, like a glycoprotein. CA125 released from ovarian cancer tissues increased time-dependently following phosphatidylinositol-specific phospholipase C (PI-PLC) treatment, concomitant with the release of tissue-unspecific alkaline phosphatase. Western blotting of CA125 treated by PI-PLC showed a single band of 90 kD instead of the 162- and 76-kD bands of the native antigen. Further, ovarian cancer tissues subjected to PI-PLC treatment lost the immunohistochemical localization of CA125 with OC125 antibody. Consequently, it is strongly suggested that CA125 is a glycoprotein that has both N- and O-linked sugar chains and a membranous GPI-anchoring moiety, and further, that its 90-kD form is the antigen without the GPI-anchor.
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PMID:Molecular nature and possible presence of a membranous glycan-phosphatidylinositol anchor of CA125 antigen. 196 50


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