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Query: UNIPROT:P08758 (annexin V)
 9,383 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Annexin I homology models were built from the annexin V crystal structure. Three methods for side-chain prediction were tested based on molecular mechanics conformational search, the use of a rotamer database, or a combination of these two methods. We showed that rotamer-based methods were more efficient and that molecular mechanics energy minimizations, prior to rotamer selection, did not afford clearly improved predictions. Models built in vacuo and with an implicit solvation term were compared with the annexin I crystal structure which became available during the course of this study. The analysis of solvation energies, root mean square deviations, chi 1 angles and hydrogen bonds showed that models built with implicit solvation were of better quality. In annexin V, repeat III displays A-B and D-E loop conformations quite different from other repeats. Since the sequence differences suggest that repeat III in annexin I might present a conformation similar to other repeats, two annexin I models with different repeat III conformations were built and compared to determine whether the correct conformation could have been predicted. We show that using a combination of evaluation criteria, it is possible to discriminate unequivocally between the native and the incorrect fold, stressing that only one criterion should not be used to evaluate protein structures.
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PMID:Homology modelling of annexin I: implicit solvation improves side-chain prediction and combination of evaluation criteria allows recognition of different types of conformational error. 770 Aug 65

Oxidant-induced damage has been proposed to be the underlying mechanism for loss of membrane phospholipid asymmetry in the erythrocyte membrane. In sickle cell disease, thalassemia, and diabetes as well as in senescent erythrocytes, an apparent correlation between oxidative damage and loss of phosphatidylserine asymmetry has been reported. In the present study, erythrocytes were subjected to various levels of oxidative stress and/or sulfhydryl modifying agents. The transmembrane location of phosphatidylserine (PS) was assessed by FITC-conjugated annexin V labeling and the PS-dependent prothrombinase assay. Transbilayer movement of spin-labeled PS was used to determine aminophospholipid translocase activity. Our data show that cells did not expose PS as the result of oxidative stress induced by phenylhydrazine, hydrogen peroxide, tert-butyl hydroperoxide, cumene hydroperoxide, or sulfhydryl modification by N-ethylmaleimide (NEM) and diamide, even under conditions that led to severe cellular damage and impairment of aminophospholipid translocase activity. In contrast, the increase of intracellular calcium induced by treatment with calcium and ionophore A23187 leads to a rapid scrambling of the lipid bilayer and the exposure of PS, which can be exacerbated by the inhibition of aminophospholipid translocase activity. Oxidation of the cells with hydrogen peroxide or phenylhydrazine did not affect A23187-induced uptake of calcium, but partly inhibited calcium-induced membrane scrambling. In conclusion, oxidative damage of erythrocytes does not induce exposure of phosphatidylserine on the membrane surface, but can interfere with both aminophospholipid translocase activity and calcium-induced randomization of membrane phospholipids.
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PMID:Oxidative damage does not alter membrane phospholipid asymmetry in human erythrocytes. 918 59

Oxygen-derived free radical injury has been associated with several cytopathic conditions. Oxygen radicals produced by chondrocytes is an important mechanism by which chondrocytes induce matrix degradation. In the present study, we extend these observations by studying oxidative processes against osteoblasts. Osteoblasts were mixed in in vitro culture with 200 microM menadione. The cytotoxic effect of menadione-induced oxidative stress was monitored by lucigenin- or luminol-amplified chemiluminescence, tetrazolium assay and immunocytochemical study. Results showed that adding menadione induces an oxidative stress on osteoblasts, via superoxide and hydrogen peroxide production, that can be eradicated by superoxide dismutase (SOD) and catalase in a dose-dependent manner. Catalase and the appropriate concentration of dimethyl sulfoxide have a protective effect on cytotoxicity induced by menadione, whereas SOD does not. Menadione-treated osteoblasts have a strong affinity for annexin V, and the nuclei are strongly stained by TUNEL (TdT-mediated dUTP nick-end labelling). The results suggest that menadione-triggered production of reactive oxygen species leads to apoptosis of osteoblasts.
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PMID:Menadione-induced cytotoxicity to rat osteoblasts. 944 50

A model of domain II of annexin I has been built by homology modelling using an annexin V crystal structure as a template. The method used is based on that of Summers and Karplus (J Mol Biol (1989) 210, 785-811) and involves the calculation of torsion-angle rotational energy maps to position side chains. The RMS deviation of the backbone heavy atoms between the model and a crystal structure of annexin I is 1.1 A. Similarities and differences in the experimental and model-derived side-chain rotameric conformations and hydrogen-bonding interactions are examined. It is found that whereas many of the side chains are well positioned some of those placed using the 'entropy argument' in which the broadest of the available minima are preferred, are erroneous. The domain is subjected to molecular dynamics simulation in explicit solvent. The simulations are found to 'correct' some of the side-chain rotamer positions that were poorly placed in the homology modelling. Considerable helix instability is seen in the simulations, consistent with the requirement of domain interactions for the structural integrity of the protein.
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PMID:Structure of human annexin I: comparison of homology modelling and crystallographic experiment. 947 53

Annexins are ubiquitous cellular proteins of unknown primary function that bind to anionic phospholipid membranes in a calcium-dependent manner. Correlative studies involving X-ray crystallography and electron microscopy suggest that annexins undergo a structural change upon binding to supported lipid monolayer membranes. In this investigation, novel spectroscopic and analytical techniques have been applied to verify and characterize this change. Soluble annexin V was examined with ordinary transmission infrared spectroscopy, while membrane-bound annexin V was examined with both transmission and internal reflection infrared spectroscopy. Spectra were processed by linked analysis, whereby multiple spectra are fit simultaneously with component bands that are constrained to share common fitting parameters. This approach is shown to enhance the sensitivity and accuracy of the bandfitting procedure. Our results are consistent with the general mode of membrane binding inferred from electron microscopy studies, and they provide independent support for the conclusion that annexin V undergoes a conformational change upon binding to lipid monolayer membranes. Most likely, this change involves the formation of new beta structure in which interstrand hydrogen bonds orient parallel to the membrane surface.
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PMID:Fourier transform infrared linked analysis of conformational changes in annexin V upon membrane binding. 989 Aug 89

The single tryptophan residue (Trp187) of human recombinant annexin V, containing 320 residues and 5328 atoms, was replaced with three different isosteric analogues where hydrogen atoms at positions 4, 5, and 6 in the indole ring were exchanged with fluorine. Such single atom exchanges of H --> F represent atomic mutations that result in slightly increased covalent bond lengths and inverted polarities in the residue side-chain structure. These minimal changes in the local geometry do not affect the secondary and tertiary structures of the mutants, which were identical to those of wild-type protein in the crystal form. But the mutants exhibit significant differences in stability, folding cooperativity, biological activity, and fluorescence properties if compared to the wild-type protein. These rather large global effects, resulting from the minimal local changes, have to be attributed either to the relatively strong changes in polar interactions of the indole ring or to differences in the van der Waals radii or to a combination of both facts. The changes in local geometry that are below resolution of protein X-ray crystallographic studies are probably of secondary importance in comparison to the strong electronegativity introduced by the fluorine atom. Correspondingly, these types of mutations provide an interesting approach to study cooperative functions of integrated residues and modulation of particular physicochemical properties, in the present case of electronegativity, in a uniquely structured and hierarchically organized protein molecule.
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PMID:Atomic mutations at the single tryptophan residue of human recombinant annexin V: effects on structure, stability, and activity. 1045 59

Effects of hydrogen peroxide (H(2)O(2)) on rat thymocytes were examined, using a flow cytometer and three fluorescent probes, annexin V-fluorescein isothiocyanate (annexin V-FITC) for detecting phosphatidylserine expressed on the membrane surface, ethidium bromide for estimating dead cells, and fluo-3-acetoxymethyl ester (fluo-3-AM) for monitoring changes in intracellular Ca(2+) concentration ([Ca(2+)](i)), to characterize H(2)O(2)-induced cytotoxicity. Exposure to H(2)O(2) (30 microM or more) increased the number of annexin V-positive live cells dose- and time-dependently while the number of dead cells increased at concentrations of 1 mM or more. H(2)O(2) (30 microM or more) increased [Ca(2+)](i) in a dose-dependent manner. Threshold concentration of H(2)O(2) to increase [Ca(2+)](i) was similar to that to increase annexin V binding to membranes. The H(2)O(2)-induced change in cell membranes was attenuated under Ca(2+)-free conditions. Therefore, it is likely that Ca(2+) is involved in the H(2)O(2)-induced cytotoxicity. Deferoxamine was effective to protect the cells suffering from H(2)O(2)-induced oxidative stress, suggesting a contribution of hydroxyl radicals generated by the Fenton reaction. Quercetin also exerted a potent protective action on cells suffering from H(2)O(2)-induced oxidative stress. The results indicate that the exposure of rat thymocytes to H(2)O(2) at micromolar concentrations increases annexin V binding to cell membranes in a Ca(2+)-dependent manner, suggesting the possibility that the oxidative stress caused by H(2)O(2) (and/or hydroxyl radicals) induces apoptosis via increasing [Ca(2+)](i).
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PMID:Exposure of rat thymocytes to hydrogen peroxide increases annexin V binding to membranes: inhibitory actions of deferoxamine and quercetin. 1061 19

We investigated cellular injury and death induced by ultrapure human Hb (HbA(0)) and its diaspirin cross-linked derivative DBBF-Hb in normal and glutathione (GSH)-depleted bovine aortic endothelial cells subjected to hydrogen peroxide (H(2)O(2)). HbA(0) underwent extensive degradation and heme loss, whereas DBBF-Hb persisted longer in its ferryl (Fe(4+)) form. The formation of ferryl HbA(0) or ferryl DBBF-Hb was associated with a significant decrease in endothelial cell GSH compared with the addition of H(2)O(2) or Hbs alone. This effect was inhibited by catalase, but not by superoxide dismutase or deferoxamine mesylate. The presence of HbA(0) and DBBF-Hb reduced H(2)O(2)-induced apoptosis, as measured by cell morphology, annexin V binding assay, and caspase inhibition, consistent with the ability to consume H(2)O(2) in an enzyme-like fashion. However, the pattern of cell death and injury produced by HbA(0) and DBBF-Hb appeared to be distinctly different among proteins as well as among cells with and without GSH. These findings may have important implications for the use of cell-free Hb as oxygen therapeutics in patients with coexisting pathologies who may lack antioxidant protective mechanisms.
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PMID:Interactions of hemoglobin with hydrogen peroxide alters thiol levels and course of endothelial cell death. 1100 76

Apoptosis followed by macrophage phagocytosis is the principal mechanism by which neutrophil granulocytes (PMN) are removed from the site of inflammation. To investigate whether Streptococcus pneumoniae causes apoptosis of PMN, we exposed PMN to viable and heat-killed pneumococci and purified pneumococcal cell walls (PCW). The occurrence of PMN cell death was quantified by flow cytometry using annexin V/propidium iodide labelling of the cells. Intracellular histone-associated DNA fragments were quantified by ELISA. The presence of apoptosis was confirmed by in situ tailing. Exposure of PMN to viable pneumococci caused necrosis of the cells. The pneumococcal cytotoxin pneumolysin, the bacterial production of hydrogen peroxide, and PCW contributed to necrosis. Heat-killed pneumococci accelerated the process of apoptosis observed in cultivated non-stimulated PMN in vitro. These results demonstrated that pneumococci induce PMN cell death. Depending on the intensity of the stimulus, PMN necrosis and apoptosis were observed.
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PMID:Induction of necrosis and apoptosis of neutrophil granulocytes by Streptococcus pneumoniae. 1101 19

Crystallographic studies have shown that the binding of calcium to domain III of annexin V is accompanied by a large conformational change involving surface exposure of Trp187. Here we examine this conformational transition using computer simulation. It is found that the burial of Trp187 is accompanied by a large increase in conformational strain, compensated by improved protein-protein interaction energies. A low energy pathway for the conformational change is determined using the conjugate peak refinement method [Fischer, S., and Karplus, M. (1992) Chem. Phys. Lett. 194, 252-261] with solvent effects taken into account using nonuniform charge scaling. The pathway obtained is complex, involving >300 dihedral angle transitions and the complete unwinding of one helix. Acidic residues play a key role in the conformational pathway, via a succession of direct hydrogen bonds with the indole ring of Trp187. This finding is discussed in the light of experimentally determined pH, calcium ion and mutational effects on the conformational transition.
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PMID:Pathway for large-scale conformational change in annexin V. 1108 53


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