Crystal and molecular structure of human annexin V after refinement

Huber, Robert; Berendes, Robert; Burger, Alexander; Schneider, Monika; Karshikov, Andrej; Luecke, Hartmut; Römisch, Jürgen; Pâques, Eric-P.

doi:10.1016/0022-2836(92)90984-r

Cited by 243 publications

(162 citation statements)

References 78 publications

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“…1 µg (28 pmol) of annexin V. Moreover, the stoichiometry of the inhibition curve suggests a substrate depletion mechanism involving a coating of the phospholipid vesicle surface by annexin V. Thus it would be expected that one annexin V molecule would cover approx. 30-60 molecules of phospholipid [7,[19][20][21] and hence 2.5 nmol of phospholipid (which should equate to at least 1.25 nmol of the phospholipid in the outer monolayer of the SUV) would require between 42 and 84 pmol (1.5-3 µg) of annexin V. The results shown in Figure 1 are clearly consistent with this stoichiometric relationship. The lack of inhibition seen when identical assays were performed with excess substrate (50 µg\ml) is consistent with this mechanism of substrate depletion (results not shown).…”

Section: Figure 1 Effect Of Annexin V On Sapc and Dopg Hydrolysis By supporting

confidence: 63%

Inhibition of human cytosolic phospholipase A2 by human annexin V

Buckland

Wilton

1998

Biochemical Journal

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The ability of annexins, particularly annexin 1 (lipocortin 1), to inhibit phospholipase A2 (PLA2) is well known and a substrate depletion mechanism is now widely accepted as the explanation for most inhibitory studies. However, there are only a very limited number of reported studies involving annexins and the high-molecular-mass cytosolic PLA2 (cPLA2). In this study we have examined the effect of human recombinant annexin V, a potentially abundant cytosolic protein, on the ability of human recombinant cPLA2 to hydrolyse a variety of phospholipid substrates. The results show clearly that, under the conditions of our study, annexin V can inhibit cPLA2 activity by a mechanism of substrate depletion and that this inhibition is dependent on the nature of the phospholipids and the concentration of Ca2+ ions in the assay. The hydrolysis of 1-stearoyl 2-arachidonyl phosphatidylcholine by cPLA2 was not significantly affected by annexin V over a range of Ca2+ concentrations (1 μM-2.5 mM), a result that presumably reflects the zwitterionic nature of the phospholipid and the known inability of annexins to bind to such interfaces. In contrast, the hydrolysis of dioleoyl phosphatidylglycerol, which is an effective anionic phospholipid substrate for this enzyme, and more significantly that of 1-stearoyl 2-arachidonyl phosphatidic acid, were readily inhibited by annexin V, although these effects were Ca2+-dependent. The Ca2+ concentrations required for inhibition in the assay system in vitro are greater than those associated with Ca2+-stimulated events within the cell, suggesting that a role for annexin V in regulating cPLA2 activity might not involve a substrate depletion mechanism in vivo unless factors in addition to Ca2+ and phospholipids contribute to the binding of annexin V to cell membranes.

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Section: Figure 1 Effect Of Annexin V On Sapc and Dopg Hydrolysis By supporting

confidence: 63%

Inhibition of human cytosolic phospholipase A2 by human annexin V

Buckland

Wilton

1998

Biochemical Journal

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“…The cationic functional groups of Lys 27 , Arg 23 , Arg 61 , and Arg 149 Ј form a closely spaced row, each separated by 4 -5 Å (Fig. 4a) (15,17). In the crystal structures of two ternary complexes formed between annexin V, Ca 2ϩ , and phospholipid head group analogs, this sulfate ion site is occupied by a phosphoryl group (21).…”

Section: Solution Liposomementioning

confidence: 99%

Interfacial Basic Cluster in Annexin V Couples Phospholipid Binding and Trimer Formation on Membrane Surfaces

Mo¹,

Campos²,

Mealy³

et al. 2003

Journal of Biological Chemistry

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Annexin V is an abundant eukaryotic protein that binds phospholipid membranes in a Ca2؉ -dependent manner. In the present studies, site-directed mutagenesis was combined with x-ray crystallography and solution liposome binding assays to probe the functional role of a cluster of interfacial basic residues in annexin V. Four mutants were investigated: R23E, K27E, R61E, and R149E. All four mutants exhibited a significant reduction in adsorption to phospholipid membranes relative to the wild-type protein, and the R23E mutation was the most deleterious. Crystal structures of wild-type and mutant proteins were similar except for local changes in salt bridges involving basic cluster residues. The combined data indicate that Arg 23 is a major determinant for interfacial phospholipid binding and participates in an intermolecular salt bridge that is key for trimer formation on the membrane surface. Together, crystallographic and solution data provide evidence that the interfacial basic cluster is a locus where trimerization is synergistically coupled to membrane phospholipid binding.Annexins comprise a large family of homologous eukaryotic proteins with a common core region that promotes Ca 2ϩ -dependent binding to phospholipid membrane surfaces (for a review, see Refs. 1-5). This property underlies many proposed in vivo annexin functions, including membrane trafficking, cell signaling, and roles in inflammatory and coagulation processes. Annexin V is an abundant protein that binds preferentially to acidic phospholipid membranes in the presence of Ca 2ϩ . The annexin-phospholipid association is of high affinity but is reversible with the removal of Ca 2ϩ . When bound to the membrane surface, annexin V molecules assemble laterally into well organized trimers and higher order arrays of trimers (6). This lateral assembly promotes spontaneous two-dimensional crystallization of annexin V on membrane surfaces, a process that is inhibited by extreme membrane surface curvature (7,8). Annexin-coated membrane surfaces undergo changes in their properties, becoming markedly rigid as the crystallization domains increase in size (9 -11). This membrane-bound layer of annexin V, which has anticoagulant properties, is believed to play a functional role in processes associated with the protein. In placenta, reduced levels of annexin V occur with anti-phospholipid syndrome, which causes hypercoagulation and recurrent miscarriage. Rand and co-workers (12, 13) have proposed a mechanism for anti-phospholipid syndrome in which the disease process strips off the protective layer of annexin V at the maternal-fetal interface, permitting excessive thrombosis to occur.The mechanism of membrane adsorption of annexin V has generated considerable interest from many disciplines, from clinical to structural biology. The protein provides an excellent model for elucidating complex interfacial behavior by peripheral membrane proteins. The molecular structure of the 35-kDa protein has been well characterized in both soluble and membrane-bound forms by x-ray ...

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“…tI values are correlation times satisfying the condition o2 . z f P 1 (o is the Larmor frequency) and defined by: l/zA = 6Rz (2) l/rB = RI + 5R2…”

Section: Theoretical Estimation Of the T2 Value Of Residual Annexin Vmentioning

confidence: 99%

“…The three-dimensional structure of human annexin V has been solved [2] and shows that the sequence domains correspond to structural domains, with each consisting of five a helices (A-E). The overall shape of the molecule is somewhat flat, with a concave surface with the N-and C-termini on one side and the calcium sites on the opposite, convex, surface.…”

mentioning

confidence: 99%

Calcium‐Induced Changes in Annexin V Behaviour in Solution as Seen by Proton NMR Spectroscopy

Neumann

Sanson

Lewit-Bentley

1994

European Journal of Biochemistry

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The behaviour of human annexin V in the presence of calcium was studied by NMR. We observe the formation of well defined dimers, as well as a change in the local dynamics of one His side chain. We assign the observed changes to either His98 or His267 residues and conclude that they could be related either to the hinge‐bending motion reported from crystal structures, or to a local side chain rearrangement within the calcium‐binding loops concerned. Dimerization was also confirmed by a small‐angle neutron‐scattering experiment. Under the experimental conditions used, we do not observe the conformational change involving Trp187 seen in previous studies, which occurs at higher relative calcium concentrations.

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Crystal and molecular structure of human annexin V after refinement

Cited by 243 publications

References 78 publications

Inhibition of human cytosolic phospholipase A2 by human annexin V

Inhibition of human cytosolic phospholipase A2 by human annexin V

Interfacial Basic Cluster in Annexin V Couples Phospholipid Binding and Trimer Formation on Membrane Surfaces

Calcium‐Induced Changes in Annexin V Behaviour in Solution as Seen by Proton NMR Spectroscopy

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