Binding of Ca2+ and Zn2+ to Human Nuclear S100A2 and Mutant Proteins

Franz, Cornelia; Durussel, Isabelle; Cox, Jos A.; Schäfer, Beat W.; Heizmann, Claus W.

doi:10.1074/jbc.273.30.18826

Cited by 47 publications

(41 citation statements)

References 56 publications

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“…Secondary Structure Probed by Circular Dichroism-The spectra shown in Fig. 6 indicate that metal-free S100A5 is rich in ␣-helical structure with a mean residue ellipticity at 222 nm of 19,000 deg cm 2 dmol Ϫ1 , compared with 15,500 for S100A2 (11). Quantitative analysis according to Johnson (33) yielded the following estimates of secondary structure: 59% ␣-helix, 8% of ␤-pleated sheet, and 5% ␤-turn.…”

Section: Ca 2ϩ and Zn 2ϩ Binding To S100a5mentioning

confidence: 99%

“…Olfactory bulb and medulla oblongata were quickly isolated on ice, frozen in powdered dry ice, and stored at Ϫ80°C. Frozen tissues were powdered, lysed, and purified over a phenyl-Sepharose column as described (11). In a final step, S100A5 was purified by reversed-phase high pressure liquid chromatography using a Brownlee C8 column.…”

Section: Methodsmentioning

confidence: 99%

“…S100A3 binds eight Zn 2ϩ , but without effect on the affinity for Ca 2ϩ (9,10). S100A2 binds four Zn 2ϩ with high affinity, in a manner antagonistic to Ca 2ϩ (11). Recently it was reported that the S100B dimer can bind four Cu 2ϩ ions with a K D of 0.46 M, most of which can be displaced with 1 mM Zn 2ϩ (12).…”

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Brain S100A5 Is a Novel Calcium-, Zinc-, and Copper Ion-binding Protein of the EF-hand Superfamily

Schäfer

Fritschy

Murmann

et al. 2000

Journal of Biological Chemistry

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S100A5 is a novel member of the EF-hand superfamily of calcium-binding proteins that is poorly characterized at the protein level. Immunohistochemical analysis demonstrates that it is expressed in very restricted regions of the adult brain. Here we characterized the human recombinant S100A5, especially its interaction with Ca 2؉ , Zn 2؉ , and Cu 2؉. Flow dialysis revealed that the homodimeric S100A5 binds four Ca 2؉ ions with strong positive cooperativity and an affinity 20 -100-fold higher than the other S100 proteins studied under identical conditions. S100A5 also binds two Zn 2؉ ions and four Cu 2؉ ions per dimer. Cu 2؉ binding strongly impairs the binding of Ca 2؉ ; however, none of these ions change the ␣-helical-rich secondary structure. After covalent labeling of an exposed thiol with 2-(4-(iodoacetamide)anilino)-naphthalene-6-sulfonic acid, binding of Cu 2؉ , but not of Ca 2؉ or Zn 2؉ , strongly decreased its fluorescence. In light of the three-dimensional structure of S100 proteins, our data suggest that in each subunit the single Zn 2؉ site is located at the opposite side of the EF-hands. The two Cu 2؉ -binding sites likely share ligands of the EF-hands. The potential role of S100A5 in copper homeostasis is discussed.Calcium, a versatile second messenger of extracellular signals inside the cell, binds to a multitude of cytosolic Ca 2ϩ -binding proteins each of which can, in turn, regulate several effector proteins. The S100 protein family (18 different members) constitutes the largest group of Ca 2ϩ -binding proteins of the EF-hand type (1, 2). At least 13 S100 genes are clustered on human chromosome 1q21, leading to the designation S100A1 to S100A13 for the protein products of these genes (3). Their expression is cell-and tissue-specific, and different human diseases have been associated with deregulated expression (4, 5). S100 proteins are non-covalent homodimers with the notable exception of the heterodimeric S100A8-S100A9. Each monomer possesses 2 EF-hands as follows: a classical C-terminal EFhand with a canonical Ca 2ϩ -binding loop of 12 amino acids and a N-terminal EF-hand with a loop of 14 amino acids which is specific for S100 proteins. This structural difference likely is the reason for the large difference in the Ca 2ϩ affinities of the N-and C-terminal EF-hands. The affinities of S100 proteins for Ca 2ϩ are in general rather low with [Ca 2ϩ ] 0.5 1 values of 100 -300 mM (for review see Refs. 2 and 6), and in 6 out of 8 studied cases binding of Ca 2ϩ displays positive cooperativity. Zn 2ϩ binds to several S100 proteins; the Zn 2ϩ and Ca 2ϩ sites are distinct and can modify the affinity for Ca 2ϩ . S100B binds four Zn 2ϩ ions with concomitant 10-fold increases of the Ca 2ϩ affinity (7). High affinity binding of two Zn 2ϩ to the S100A12 dimer leads to induction of two high affinity Ca 2ϩ -binding sites (8). S100A3 binds eight Zn 2ϩ , but without effect on the affinity for Ca 2ϩ (9, 10). S100A2 binds four Zn 2ϩ with high affinity, in a manner antagonistic to Ca 2ϩ (11). Recently it was reported...

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Section: Ca 2ϩ and Zn 2ϩ Binding To S100a5mentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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Brain S100A5 Is a Novel Calcium-, Zinc-, and Copper Ion-binding Protein of the EF-hand Superfamily

Schäfer

Fritschy

Murmann

et al. 2000

Journal of Biological Chemistry

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“…Human p53 proteins in nuclear extracts were obtained from H1299 cells after transient transfection using the calcium phosphate method. Human recombinant S100A2 protein was purified as previously described (8). Nuclear extracts were incubated in the presence of 5ϫ gel shift binding buffer (20% glycerol, 5 mM MgCl 2 , 2.5 mM EDTA, 2.5 mM dithiothreitol, 250 mM NaCl, 50 mM Tris-HCl, pH 7.5, 0.25 mg/ml poly(dI-dC)) and 0.5 and 1 g of recombinant S100A2 protein.…”

Section: Methodsmentioning

confidence: 99%

The Calcium-binding Protein S100A2 Interacts with p53 and Modulates Its Transcriptional Activity

Mueller¹,

Schäfer²,

Ferrari

et al. 2005

Journal of Biological Chemistry

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129

126

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Head and neck squamous cell carcinoma express high levels of the EF-hand calcium-binding protein S100A2 in contrast to other tumorigenic tissues and cell lines where the expression of this protein is reduced. Subtractive hybridization of tumorigenic versus normal tumor-derived mammary epithelial cells has previously identified the S100A2 protein as potential tumor suppressor. The biological function of S100A2 in carcinogenesis, however, has not been elucidated to date. Here, we report for the first time that during recovery from hydroxyurea treatment, the S100A2 protein translocated from the cytoplasm to the nucleus and co-localized with the tumor suppressor p53 in two different oral carcinoma cells (FADU and SCC-25). Co-immunoprecipitation experiments and electrophoretic mobility shift assay showed that the interaction between S100A2 and p53 is Ca 2؉ -dependent. Preliminary characterization of this interaction indicated that the region in p53 involved with binding to S100A2 is located at the C terminus of p53. Finally, luciferase-coupled transactivation assays, where a p53-reporter construct was used, indicated that interaction with S100A2 increased p53 transcriptional activity. Our data suggest that in oral cancer cells the Ca 2؉ -and cell cycle-dependent p53-S100A2 interaction might modulate proliferation.

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“…S100A2 is expressed in many cells of the normal epidermis (Table 1) Eckert et al 2004;Maelandsmo et al 1997). S100A2 is dimeric and binds both zinc (K D = 25 nM) and calcium (K D = 470 μM) and binding to zinc reduces significantly the affinity for calcium (Franz et al 1998;Koch et al 2007). S100A2 is mainly localized in the cell nucleus where it is believed to play the role of tumor suppressor (Glenney, Kindy, and Zokas 1989).…”

Section: S100a2mentioning

confidence: 99%

The Roles of S100 Proteins and RAGE in Melanoma

Leclerc¹

2011

Breakthroughs in Melanoma Research

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Binding of Ca2+ and Zn2+ to Human Nuclear S100A2 and Mutant Proteins

Abstract: The Ca 2؉ -binding protein S100A2 is an unusual member of the S100 family, characterized by its nuclear localization and down-regulated expression in tumorigenic cells. In this study, we investigated the properties of human recombinant S100A2 (wtS100A2

Cited by 47 publications

References 56 publications

Brain S100A5 Is a Novel Calcium-, Zinc-, and Copper Ion-binding Protein of the EF-hand Superfamily

Brain S100A5 Is a Novel Calcium-, Zinc-, and Copper Ion-binding Protein of the EF-hand Superfamily

The Calcium-binding Protein S100A2 Interacts with p53 and Modulates Its Transcriptional Activity

The Roles of S100 Proteins and RAGE in Melanoma

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