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

Mo, Youde; Campos, Begoña; Mealy, Tanya R.; Commodore, Lois; Head, James F.; Dedman, John R.; Seaton, Barbara A.

doi:10.1074/jbc.m210286200

Cited by 39 publications

(39 citation statements)

References 41 publications

(27 reference statements)

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“…Data obtained from crystallography provide evidence that the interfacial basic cluster is the place for dimerization of molecules of ANX V, which is synergistically coupled to membrane phospholipid binding [65]. The lipid bilayer shows a bent shape and contains a concave region in the annexin-membrane interaction interface, which supports the idea that ANX V could disturb the stability of lipids and bend membranes [66].…”

Section: Understanding the Use Of Annexin V In Sperm Selectionsupporting

confidence: 60%

Use of Annexin V based Sperm Selection in Assisted Reproduction

Teijeiro¹,

Munuce²,

Caille³

et al. 2017

Andrology

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Innovative methods to select sperm subpopulations with the best fertilizing ability are needed in assisted reproductive techniques (ART) in order to improve fertilization and pregnancy rates, while also considering possible epigenetic effects on the offspring. Molecular based selection methods are searched for, under the premise that they could be an improvement over classical selection by morphology and movement. One of these methods sustains the elimination of sperm that can bind to annexin A5 (ANX V), coupled to paramagnetic beads, through the phosphatidyl-serine exposed on their membranes upon apoptosis. Although reports accumulate about the use of this method, controversy persists as to the benefits of ANX V based sperm selection in ART. In this review we consider the arguments in favour and against this method and conclude that to the moment the evidence does not support MACS regular use in ART.

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Section: Understanding the Use Of Annexin V In Sperm Selectionsupporting

confidence: 60%

Use of Annexin V based Sperm Selection in Assisted Reproduction

Teijeiro¹,

Munuce²,

Caille³

et al. 2017

Andrology

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“…Arg-25 and Lys-29 are part of a basic cluster at a key trimer interface, and mutation of these residues disrupts salt bridges between monomers in the trimer unit (37). However, mutation of these residues to neutral glutamine had no effect on the curve of FRET as a function of occupancy, which was identical to that of the wild-type protein (Fig.…”

Section: Constant Free Energy Of Binding To Membranes With Different mentioning

confidence: 56%

Entropic and Enthalpic Contributions to Annexin V-Membrane Binding

Jeppesen

Smith

Gibson

et al. 2008

Journal of Biological Chemistry

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Annexin V binds to membranes with very high affinity, but the factors responsible remain to be quantitatively elucidated. Analysis by isothermal microcalorimetry and calcium titration under conditions of low membrane occupancy showed that there was a strongly positive entropy change upon binding. For vesicles containing 25% phosphatidylserine at 0.15 M ionic strength, the free energy of binding was ؊53 kcal/mol protein, whereas the enthalpy of binding was ؊38 kcal/mol. Addition of 4 M urea decreased the free energy of binding by about 30% without denaturing the protein, suggesting that hydrophobic forces make a significant contribution to binding affinity. This was confirmed by mutagenesis studies that showed that binding affinity was modulated by the hydrophobicity of surface residues that are likely to enter the interfacial region upon protein-membrane binding. The change in free energy was quantitatively consistent with predictions from the WimleyWhite scale of interfacial hydrophobicity. In contrast, binding affinity was not increased by making the protein surface more positively charged, nor decreased by making it more negatively charged, ruling out general ionic interactions as major contributors to binding affinity. The affinity of annexin V was the same regardless of the head group present on the anionic phospholipids tested (phosphatidylserine, phosphatidylglycerol, phosphatidylmethanol, and cardiolipin), ruling out specific interactions between the protein and non-phosphate moieties of the head group as a significant contributor to binding affinity. Analysis by fluorescence resonance energy transfer showed that multimers did not form on phosphatidylserine membranes at low occupancy, indicating that annexin-annexin interactions did not contribute to binding affinity. In summary, binding of annexin V to membranes is driven by both enthalpic and entropic forces. Dehydration of hydrophobic regions of the protein surface as they enter the interfacial region makes an important contribution to overall binding affinity, supplementing the role of protein-calcium-phosphate chelates.

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“…Deletion of AB calcium binding sites in all four domains created a molecule with minimal binding activity that could only be detected under conditions of very low ionic strength and very high membrane PS content. The residual activity detected under these conditions could be due to the remaining calcium binding sites (in the DE helices or elsewhere), or perhaps due to ionic interactions with certain positively charged residues that may be involved in regulating binding affinity and/or trimer formation (18,37).…”

Section: Role Of Individual Sites In Membranementioning

confidence: 99%

Essential Role of B-helix Calcium Binding Sites in Annexin V-Membrane Binding

Jin¹,

Smith²,

Hsieh³

et al. 2004

Journal of Biological Chemistry

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Crystal structures of annexin V have shown up to 10 bound calcium ions in three different types of binding sites, but previous work concluded that only one of these sites accounted for nearly all of the membrane binding affinity of the molecule. In this study we mutated residues contributing to potential calcium binding sites in the AB and B helices in each of the four domains (eight sites in total) and in DE helices in the first, second, and third domains (three sites in total). We measured the affinity of each protein for phospholipid vesicles and cell membranes by quantitative calcium titration under low occupancy conditions (<1% saturation of available membrane binding sites). Affinity was calculated from the midpoint and slope of the calcium titration curve and the concentration of membrane binding sites. The results showed that all four AB sites were essential for high affinity binding, as were three of the four B sites (in domains 1, 2, and 3); the DE site in the first domain made a slight contribution to affinity. Multisite mutants showed that each domain contributed additively and independently to binding affinity; in contrast, AB and B sites within the same domain were interdependent. The number of functionally important sites identified was consistent with the Hill coefficient observed in calcium titrations. This study shows an essential and previously unappreciated role for B-helix calcium binding sites in the membrane binding of annexins and indicates that all four domains of the molecule are required for maximum membrane binding affinity.The annexins are a family of calcium-dependent membranebinding proteins with diverse functions (1-4). Annexin V is a typical member of this protein family, which is becoming important as a means to measure cell death in vivo in cancer chemotherapy, organ transplant rejection, and myocardial infarction (5-10), creating a need to better understand structural and functional properties relevant to its clinical use. Novel engineered forms of annexin V are being developed to improve its utility for imaging (11,12), and there has been a recent report of a single-domain version of annexin V that may be useful for imaging cell death in vivo (13).Calcium binding sites in annexins are of three structural types: AB (or Type II), DE (or Type III), and B or ABЈ (14). In these binding sites, calcium ligands consist of one to three carbonyl oxygens from the protein backbone, and one carboxyl group from an Asp or Glu residue; sites are often named based on the domain (1 through 4) and ␣-helices (A through E), which contribute the backbone carbonyl ligands. However, the number of calcium ions observed in different structures varies widely; for example, anywhere from 2 or 3 (15, 16) to 10 (17) calcium ions have been observed in different crystal forms of annexin V. Thus, it has been difficult to determine which calcium binding sites are functionally important based on structural data alone. Previous work with annexin V mutants indicated that the AB site in domain 1 was the most impor...

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

Cited by 39 publications

References 41 publications

Use of Annexin V based Sperm Selection in Assisted Reproduction

Use of Annexin V based Sperm Selection in Assisted Reproduction

Entropic and Enthalpic Contributions to Annexin V-Membrane Binding

Essential Role of B-helix Calcium Binding Sites in Annexin V-Membrane Binding

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