Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P08758 (annexin V)
 9,383 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Two calcium-dependent phospholipid- and membrane-binding proteins have been purified from bovine brain. These are termed CaBP33 and CaBP37. Complete sequence analysis has revealed that these two proteins are isoforms of annexin V. Despite an apparent difference of 4 kDa between the two proteins on SDS-PAGE, only two amino-acid substitutions were found. These are, in CaBP33, Ser-36 and Lys-125 and in CaBP37, Thr-36 and Glu-125. This corresponds to a mass difference of 15 Da. This was confirmed by electrospray mass spectrometric analysis. Both isoforms can be phosphorylated substoichiometrically in vitro by protein kinase C at residue Thr-22.
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PMID:Novel isoforms of CaBP 33/37 (annexin V) from mammalian brain: structural and phosphorylation differences that suggest distinct biological roles. 142 Mar 35

Annexins represent a widespread family of Ca(++)-dependent phospholipid binding proteins. Although their precise functions are still unknown, they probably play an important role in cell regulation because they are major substrates for various growth factor receptor kinases. We characterized annexins in human skin using three different antisera raised against annexin II, annexin V, and a synthetic peptide that resembles the consensus sequence of all annexins. In normal human skin, using SDS-PAGE, two-dimensional gel electrophoresis and immunoblot analysis, we identified two major 34-kDa proteins and one 36-kDa protein, with respective isoelectric points of 6.5, 5.2, and 7.2-7.9. According to these criteria they were identified as annexins, I, V, and II, respectively. Minor 45-51 kDa and 68-kDa proteins with 6.1-6.7 and 6.8-7.1 isoelectric points were also present, and likely corresponded to annexins VII and VI, respectively. We investigated the ability of these proteins to bind phospholipids in the presence of calcium using liposomes formed from a mixture of phosphatidylserine and phosphatidylcholine. The cellular distribution of annexins in normal human skin was determined by immunofluorescence with antiannexin II and anti-annexin V antibodies. Labeling with both antibodies was observed predominantly at the cell membrane with some cytoplasmic staining also being apparent. Specificity was confirmed by the absence of staining using pre-immune sera or after the absorption of the antibodies with their corresponding antigens. These proteins were also characterized in vitro in a reconstituted human skin model. All were present in this system except annexin VI and VII, which were lost after phospholipid purification. Further experiments should now be carried out using this system to clarify the role and regulation of these proteins within the epidermis.
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PMID:Characterization and subcellular localization of calcium-dependent phospholipid binding proteins (annexins) in normal human skin and reconstituted epidermis. 153 82

Annexin V belongs to a large family of calcium-binding and phospholipid-binding proteins and may act as an endogenous regulator of the protein kinase C (PKC) activity. This study examines the effect of annexin V on the in vitro PKC activity in cultured mesangial cells using histone H1, the peptide [Ser25]PKC-(19-31), or endogenous proteins as substrates. The SDS/PAGE pattern of 32P-labeled mesangial proteins showed that the calcium-independent PKC [(n+a)PKC] phosphorylated several proteins from 70 kDa to 40 kDa and 22 kDa to 15 kDa. Three additional proteins from 34 kDa to 29 kDa, including annexin I and its proteolytic forms, were detected after activation of calcium-dependent PKC (cPKC). Increasing concentrations of annexin V did not alter the phosphorylation of (n+a)PKC substrates. By contrast, specific phosphorylation of proteins and annexin I by cPKC, was reduced in a dose-dependent manner. Addition of high concentration of calcium and phosphatidylserine did not reverse the inhibitory effect of annexin V. Annexin V also inhibited the phosphorylation of histone H1 or peptide [Ser25]PKC-(19-31) by cPKC. Moreover, removal of annexin V from cytosols increased the annexin I phosphorylation by these isoforms. From these results, we propose that annexin V may regulate the signal-transduction pathway involving the activation of cPKC, as they act in vitro as an inhibitor of these kinases.
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PMID:Inhibitory effect of annexin V on protein kinase C activity in mesangial cell lysates. 758 28

The cloning, purification and characterization of full-length annexin V, expressed intracellularly in Saccharomyces cerevisiae is detailed. Following homogenization in a glass bead mill, clarification by ultracentrifugation and fractional ammonium sulfate precipitation, the 319 amino acid protein was purified by column chromatography on phenyl-Sepharose and heparin-Sepharose. Annexin V elutes on reverse phase C4 silica as a single peak with greater than 97% homogeneity and is further characterized by a molecular mass of 34 kDa from electrophoresis under reducing conditions on SDS gels. Dynamic light scattering experiments reveal annexin V exists as a monomer in solution. Amino terminal Edman degradation afforded no sequence, therefore the carbamidomethylated protein was chemically cleaved with cyanogen bromide. Separation of the resulting peptide fragments on reverse phase HPLC followed by N-terminal sequencing and electrospray mass spectrometry supported the correct sequence as well as the existence of an acetyl blocking group on the N-terminus. The protein exhibits an isoelectric point of 4.73 by column chromatofocusing. Secondary structure predictions from CD spectroscopy indicate that the molecule is correctly folded. In anticoagulant assays, the purified protein exhibits dose-response effects in activated partial thromboplastin time (APTT) prolongation and doubles the clotting time of control human plasma at 70 micrograms ml-1. More specifically, in a factor Xa inhibition assay in which the activation of factor X via the tissue factor-factor VIIa complex is monitored by the cleavage of a factor Xa chromogenic substrate, recombinant annexin V exhibits a 50% inhibitory concentration (IC50) in the low nanomolar range.
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PMID:Isolation and characterization of recombinant annexin V expressed in Saccharomyces cerevisiae. 776 33

We previously reported a new type of lectin, p33/41 (annexin IV), which was isolated from a bovine tissue extract [Kojima, K. et al. (1992) J. Biol. Chem. 267, 20536-20539]. When the expression of p33/41 (annexin IV) was surveyed in the lysates of 39 human tumor cell lines by SDS-PAGE, followed by Western blot analysis with polyclonal anti-bovine p33/41 and monoclonal anti-annexin IV (Z016, Zymed) antibodies, 21 cell lines were found to be reactive with the polyclonal antibody, whereas all 39 cell lines were stained with Z016. These results together with those obtained with standard proteins, annexins IV and V, suggested that the monoclonal antibody, Z016, recognizes annexin V, but not p33/41 (annexin IV). Therefore, we performed cDNA cloning of human p33/41 (annexin IV) to prepare a recombinant protein and raised monoclonal antibodies against the protein. Northern blot analysis with the cDNA as a probe showed that a human colon cancer cell line, HT29, contains p33/41 (annexin IV) mRNA of two sizes, 2.0 and 3.0 kb. The two monoclonal antibodies, AS11 and AS17, against the recombinant protein generated were useful for flow cytometric analysis, ELISA, Western blot analysis and immunoprecipitation. Flow cytometric analysis with AS17 showed that p33/41 (annexin IV) is located in the cytoplasm of HT29 cells, but not on the cell surface. However, one of the cell surface proteins first labeled with biotin and then solubilized with a detergent was immunoprecipitated with AS17. The results suggest the existence of a membrane spanning form of p33/41 (annexin IV).
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PMID:Expression of carbohydrate-binding protein p33/41 in human tumor cell lines. 888 29

While previous studies revealed that matrix vesicles (MV) contain a nucleational core (NC) that converts to apatite when incubated with synthetic cartilage lymph, the initial mineral phase present in MV is not well characterized. This study explored the physicochemical nature of this Ca2+ and Pi-rich NC. MV, isolated from growth plate cartilage, were analyzed directly by solid-state 31P NMR, or incubated with hydrazine or NaOCl to remove organic constituents. Other samples of MV were subjected to sequential treatments with enzymes, salt solutions, and detergents to expose the NC. We examined the NC using transmission electron microscopy, energy-dispersive analysis with x-rays, and electron and x-ray diffraction, Fourier transform-infrared spectroscopy, high performance thin-layer chromatographic analysis, and SDS-polyacrylamide gel electrophoresis. We found that most of the MV proteins and lipids could be removed without destroying the NC; however, NaOCl treatment annihilated its activity. SDS-polyacrylamide gel electrophoresis showed that annexin V, a phosphatidylserine (PS)-dependent Ca2+-binding protein, was the major protein in the NC; high performance thin-layer chromatographic analysis revealed that the detergents removed the majority of the polar lipids, but left significant free cholesterol and fatty acids, and small but critical amounts of PS. Transmission electron microscopy showed that the NC was composed of clusters of approximately 1.0 nm subunits, which energy-dispersive analysis with x-rays revealed contained Ca and Pi with a Ca/P ratio of 1.06 +/- 0. 01. Electron diffraction, x-ray diffraction, and Fourier transform-infrared analysis all indicated that the NC was noncrystalline. 1H-Cross-polarization 31P NMR indicated that the solid phase of MV was an HPO42--rich mixture of amorphous calcium phosphate and a complex of PS, Ca2+, and Pi. Taken together, our findings indicate that the NC of MV is composed of an acid-phosphate-rich amorphous calcium phosphate intermixed with PS-Ca2+-Pi, annexin V, and other proteins and lipids.
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PMID:Physicochemical characterization of the nucleational core of matrix vesicles. 902 Jan 63

Previous studies revealed that matrix vesicles (MV) have an acid-labile nucleationally active core (ALNAC) essential for mineral formation; current studies were aimed at characterizing and reconstituting ALNAC. SDS-PAGE and FTIR analyses revealed the presence of lipids, proteins and amorphous calcium phosphate (ACP) in ALNAC. Extraction with chloroform-methanol reduced, but did not destroy MV calcification; treatment with chloroform-methanol-HCl destroyed all activity. This acidic solvent extracted the annexins, (phosphatidylserine (PS)-dependent Ca(2+)-binding proteins), and dissociated PS-Ca(2+)-Pi complexes present in the MV. Attempts to reconstitute ALNAC, centered on the Ca(2+)-PS-Pi complex. Various pure lipids, electrolytes and proteins were combined to form a synthetic nucleationally active complex (SNAC), analyzing the rate of Ca2+ uptake. Inclusion of phosphatidylethanolamine (PE) or sphingomyelin (SM) with PS, or Mg2+ or Zn2+ with Ca2+, strongly inhibited activity; incorporation of annexin V increased SNAC activity. Thus, approaching from either deconstruction or reconstruction, it appears that ALNAC is composed of ACP complexed with PS and the annexins. Other lipids, proteins and electrolytes modulate its activity. These findings also indicate how ALNAC must be formed in vivo.
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PMID:Characterization and reconstitution of the nucleational complex responsible for mineral formation by growth plate cartilage matrix vesicles. 908 69

Annexin V has been characterized as a major collagen type II binding cell-surface component of normal chondrocytes and is also called anchorin CII in chondrogenic populations. Herein we present evidence that in vitro cultured Swarm rat chondrosarcoma cells are not capable of binding collagen type II in significant quantities to their surfaces, as compared to normal rat chondrocytes. This finding coincides with a deficiency of annexin V on the surface of these cells. A small quantity of an intracellular polypeptide could be detected which is immunologically cross-reactive with annexin V but displayed a mobility in SDS-PAGE of less than 34 kD compared to the M(r) 36 kD of intact rat annexin V. By immunohistochemistry the protein could be localized in the cytoplasm of in vitro and in vivo grown tumor cells. By reverse transcription-polymerase chain reaction and Northern blot analysis, a regular-sized mRNA for annexin V could be detected in the chondrosarcoma cells that is expressed in only slightly lower quantities than in normal chondrocytes. Taken together, the data suggest a modified processing or turnover for annexin V in the chondrosarcoma excluding it from being a functionally active collagen type II binding protein. The findings support the hypothesis of cell-surface annexin V as a key component for the formation of the pericellular matrix of chondrocytes.
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PMID:Absence of cell-surface annexin V is accompanied by defective collagen matrix binding in the Swarm rat chondrosarcoma. 913 73

We have previously developed a human macrophage hybridoma model system to study the effect of HIV-1 infection on monocytic function. Upon coculture of one chronically (35 days postinfection) HIV-1-infected human macrophage hybridoma cell line, 43HIV, there was a dose-dependent decrease in the viability of cocultured Ag-stimulated T cells associated with an increase in DNA strand breaks. Enhanced apoptosis was determined by labeling with biotinylated dUTP and propidium iodide, increased staining with annexin V, increased side light scatter and expression of CD95, and decreased forward light scatter and expression of Bcl-2. There was also increased DNA strand breaks as determined by propidium iodide staining in unstimulated T cells cocultured with 43HIV and in T cells stimulated with anti-CD3 mAb and PHA. Pretreatment with 5145, a human polyclonal anti-gp120 Ab that recognizes the CD4 binding region, as well as with an anti-Fas ligand mAb blocked apoptosis in CD4+ T cells but not in CD8+ T cells. A soluble factor with a Mr below 10,000 Da was defined that induced apoptosis in CD4+ and CD8+ T cells and B cells. SDS-PAGE analysis of the active fractions revealed a band of 6000 Da that, after electroelution, had proapoptotic activity. The pI of the activity was estimated to be between 6.5 and 7.0. In conclusion, chronically HIV-1-infected monocytic cells induce apoptosis in bystander-, Ag-, anti-CD3-, and mitogen-stimulated T cells by multiple factors, which may contribute to the depletion of lymphocytes induced by HIV-1.
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PMID:Chronically HIV-1-infected monocytic cells induce apoptosis in cocultured T cells. 1705 88

Activated blood platelets shed microparticles with procoagulant activity that probably participate in normal hemostasis. We have isolated spontaneously formed microparticles from human blood and analysed them for ultrastructure, antigenic profile, and biochemical composition. In transmission electron microscopy microparticles appeared as regular vesicles with a mean diameter of 300 nm (50-600 nm). In flow cytometry almost all microparticles reacted with fluorescein isothiocyanate (FITC) labeled antibody to platelet glycoprotein complex IIb-IIIa (GpIIb-IIIa) and with FITC-annexin V but only 40-50% of microparticles reacted with FITC-antibody to platelet glycoprotein Ib (GpIb). The latter result was confirmed by double labeling of microparticles with FITC-antibody to GpIIb-IIIa and phycoerythrin (PE) labeled antibody to GpIb. Large microparticles reacted better with anti-GpIb than the small ones. A decreased level of GpIb was also demonstrated by SDS/polyacrylamide gel electrophoresis of microparticles. Compositional studies indicated, that in terms of cholesterol and protein contents, microparticles resembled platelets rather than platelet membranes as previously thought. They are, however, deficient in certain components. Thus, in comparison to platelets, microparticles had reduced contents of sialic acid (by 56.4%), galactosamine (by 48.2%), glucosamine (by 22.4%), galactose by (11.8%) and fucose (by 21.6%). Mannose content was increased by 11.8%. Total phospholipids in microplatelets were lower by 17.8%. Glycerophospholipids only were affected with phosphatidylserine being decreased as much as by 43.2%. Neutral glycosphingolipids, gangliosides and ceramides in microparticles were reduced by half.
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PMID:In comparison to progenitor platelets, microparticles are deficient in GpIb, GpIb-derived carbohydrates, glycerophospholipids, glycosphingolipids, and ceramides. 982 72


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