Isabella Ascone
Centre national de la recherche scientifique
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Featured researches published by Isabella Ascone.
Journal of Synchrotron Radiation | 2001
Roger Fourme; Richard A. Kahn; Mohamed Mezouar; Eric Girard; C. Hoerentrup; Thierry Prangé; Isabella Ascone
A new set-up and associated methodology for the collection of angle-dispersive diffraction data from protein crystals submitted to high hydrostastic pressure have been developed on beamline ID30 at the ESRF. The instrument makes use of intense X-rays of ultra-short wavelength emitted by two collinear undulators, and combines a membrane-driven diamond-anvil cell mounted on a two-axis goniometer and an imaging-plate scanner. Sharp and clean diffraction pictures from tetragonal crystals of hen egg-white lysozyme (tHEWL) and orthorhombic crystals of bovine erythrocyte Cu, Zn superoxide dismutase (SOD) were recorded at room temperature and pressures up to 0.915 and 1.00 GPa, respectively. The compressibility of tHEWL was determined from unit-cell parameters determined at 24 different pressures up to 0.915 GPa. High-pressure diffraction data sets from several crystals of tHEWL were collected and analyzed. Merging of data recorded on different crystals at 0.30 and 0.58 GPa produced two sets of structure amplitudes with good resolution, completeness, redundancy and R(sym) values. A third set at 0.69 GPa was of a similar quality except a lower completeness. The three structures have been refined. The pressure-induced loss of crystalline order in a tHEWL crystal beyond 0.82 GPa was captured through a series of diffraction pictures.
Journal of Synchrotron Radiation | 2003
Isabella Ascone; Wolfram Meyer-Klaucke; Loretta M. Murphy
Spectroscopic techniques, like X-ray absorption spectroscopy, will provide important input for integrated biological projects in genomics and proteomics. This contribution summarizes technical requirements and typical set-ups for both simple and complex biological XAS experiments. An overview on different strategies for sample preparation is discussed in detail. Present and future BioXAS spectrometers are presented to help potential users in locating the spectrometer required for their biological application.
Inorganic Chemistry | 2008
Isabella Ascone; Luigi Messori; Angela Casini; Chiara Gabbiani; Antonella Balerna; Francesca Dell'Unto; Agostina Congiu Castellano
The reaction of bovine serum albumin (BSA) with [ trans-RuCl 4(Im)(dimethylsulfoxide)][ImH] (Im = imidazole) (NAMI-A), an experimental ruthenium(III) anticancer drug, and the formation of the respective NAMI-A/BSA adduct were investigated by X-ray absorption spectroscopy (XAS) at the sulfur and chlorine K-edges and at the ruthenium K- and L 3-edges. Ruthenium K and L 3-edge spectra proved unambiguously that the ruthenium center remains in the oxidation state +3 after protein binding. Comparative analysis of the chlorine K-edge XAS spectra of NAMI-A and NAMI-A/BSA, revealed that the chlorine environment is greatly perturbed upon protein binding. Only modest changes were observed in the sulfur K-edge spectra that are dominated by several protein sulfur groups. Overall, valuable information on the nature of this metallodrug/protein adduct and on the mechanism of its formation was gained; XAS spectroscopy turns out to be a very suitable method for the characterization of this kind of systems.
Structure | 2002
Roger Fourme; Isabella Ascone; Richard A. Kahn; Mohamed Mezouar; Pierre Bouvier; Eric Girard; Tianwei Lin; John E. Johnson
The combined use of a diamond anvil cell and ultrashort-wavelength undulator radiation has allowed the collection of high-resolution diffraction data from protein and virus crystals submitted to hydrostatic pressures beyond 2 kbar. Crystals of cubic cowpea mosaic virus (CPMV) can be compressed to at least 3.5 kbar. Diffraction from CPMV crystals displaying an unusual disorder at atmospheric pressure was considerably enhanced by application of pressure. These experiments suggest that pressure may be used in some cases to improve order in crystals.
Annual review of biophysics | 2009
Roger Fourme; Eric Girard; Richard A. Kahn; Anne-Claire Dhaussy; Isabella Ascone
A survey of the main interests of high pressure for molecular biophysics highlights the possibility of exploring the whole conformational space using pressure perturbation. A better understanding of fundamental mechanisms responsible for the effects of high pressure on biomolecules requires high-resolution molecular information. Thanks to recent instrumental and methodological progress taking advantage of the remarkable adaptation of the crystalline state to hydrostatic compression, pressure-perturbed macromolecular crystallography is now a full-fledged technique applicable to a variety of systems, including large assemblies. This versatility is illustrated by selected applications, including DNA fragments, a tetrameric protein, and a viral capsid. Binding of compressed noble gases to proteins is commonly used to solve the phase problem, but standard macromolecular crystallography would also benefit from the transfer of experimental procedures developed for high-pressure studies. Dedicated short-wavelength synchrotron radiation beamlines are unarguably required to fully exploit the various facets of high-pressure macromolecular crystallography.
Journal of Synchrotron Radiation | 2009
Isabella Ascone; Richard W. Strange
In the past seven years the size of the known protein sequence universe has been rapidly expanding. At present, more then five million entries are included in the UniProtKB/TrEMBL protein database. In this context, a retrospective evaluation of recent X-ray absorption studies is undertaken to assess its potential role in metalloproteomics. Metalloproteomics is the structural and functional characterization of metal-binding proteins. This is a new area of active research which has particular relevance to biology and for which X-ray absorption spectroscopy is ideally suited. In the last three years, biological X-ray absorption spectroscopy (BioXAS) has been included among the techniques used in post-genomics initiatives for metalloprotein characterization. The emphasis of this review is on the progress in BioXAS that has emerged from recent meetings in 2007-2008. Developments required to enable BioXAS studies to better contribute to metalloproteomics throughput are also discussed. Overall, this paper suggests that X-ray absorption spectroscopy could have a higher impact on metalloproteomics, contributing significantly to the understanding of metal site structures and of reaction mechanisms for metalloproteins.
Journal of Biological Inorganic Chemistry | 2011
Luigi Messori; A. Balerna; Isabella Ascone; C. Castellano; Chiara Gabbiani; Angela Casini; C. Marchioni; G. Jaouen; A. Congiu Castellano
Gold metallodrugs form a class of promising antiproliferative agents showing a high propensity to react with proteins. We exploit here X-ray absorption spectroscopy (XAS) methods [both X-ray absorption near-edge spectroscopy (XANES) and extended X-ray absorption fine structure (EXAFS)] to gain insight into the nature of the adducts formed between three representative gold(I, III) metallodrugs (i.e., auranofin, [Au(2,2′-bipyridine)(OH)2](PF6), Aubipy, and dinuclear [Au2(6,6′-dimethyl-2,2′-bipyridine)2(μ-O)2](PF6)2, Auoxo6) and two major plasma proteins, namely, bovine serum albumin (BSA) and human serum apotransferrin (apoTf). The following metallodrug–protein systems were investigated in depth: auranofin/apoTf, Aubipy/BSA, and Auoxo6/apoTf. XANES spectra revealed that auranofin, upon protein binding, conserves its gold(I) oxidation state. Protein binding most probably takes place through release of the thiosugar ligand and its subsequent replacement by a thiol (or a thioether) from the protein. This hypothesis is independently supported by EXAFS results. In contrast, the reactions of Aubipy with serum albumin and of Auoxo6 with serum apoTf invariantly result in gold(III) to gold(I) reduction. Gold(III) reduction, clearly documented by XANES, is accompanied, in both cases, by release of the bipyridyl ligands; for Auoxo6 cleavage of the gold–gold dioxo bridge is also observed. Gold(III) reduction leads to formation of protein-bound gold(I) species, with deeply modified metal coordination environments, as evidenced by EXAFS. In these adducts, the gold(I) centers are probably anchored to the protein through nitrogen donors. In general, these two XAS methods, i.e., XANES and EXAFS, used here jointly, allowed us to gain independent structural information on metallodrug/protein systems; detailed insight into the gold oxidation state and the local environment of protein-bound metal atoms was achieved in the various cases.
FEBS Letters | 1993
Isabella Ascone; Antonella Longo; H. Dexpert; Maria Rosa Ciriolo; Giuseppe Rotilio; Alessandro Desideri
The CU(I)GSH complex has recently been shown to be a good candidate for delivering copper to the active site of Cu‐free Cu,Zn superoxide dismutase both in vivo and in vitro. In this work X‐ray absorption spectroscopy has been used to characterize the CU(I)GSH complex and to follow in vitro the reconstitution of Cu,Zn superoxide dismutase from the copper‐free protein and this complex. The results obtained indicate that the copper is directly transferred as Cu(I) from the GSH complex into the empty copper binding site. No evidence has been obtained for a ternary complex in which the metal is bound to both GSH and the protein.
Journal of Synchrotron Radiation | 2003
Sophie Quevillon-Cheruel; Bruno Collinet; Cong-Zhao Zhou; Philippe Minard; Karine Blondeau; Gilles Henkes; Robert Aufrère; Jérôme Coutant; Eric Guittet; Anita Lewit-Bentley; Nicolas Leulliot; Isabella Ascone; Isabelle Sorel; Philippe Savarin; Ines Li de La Sierra Gallay; Françoise de la Torre; Anne Poupon; Roger Fourme; Joël Janin; Herman van Tilbeurgh
A canonical structural genomics programme is being conducted at the Paris-Sud campus area on bakers yeast proteins. Experimental strategies, first results and identified bottlenecks are presented. The actual or potential contributions to the structural genomics of several experimental structure-determination methods are discussed.
European Biophysics Journal | 1994
S. Della Longa; Isabella Ascone; A. Fontaine; A. Congiu Castellano; A. Bianconi
The ligand photodissociation of sperm whale carboxymyoglobin (MbCO) at low temperature (15 K-100 K) under extended illumination has been studied by X-ray Absorption Near Edge Structure (XANES) spectroscopy using the dispersive technique. XANES simulations through the multiple scattering (MS) approach allow one to interpret the spectroscopic data in structural terms, and to investigate the Fe site structure configurations of the states that follow the CO photodissociation as a function of temperature. The Fe site in the photoproduct is unbound, with an overall structure similar to the deoxy-form (Mb) of the protein. The Fe site structure changes from T < 30 K (Mb*) to T>50 K (Mb**), revealing the existence of a slower unbound state Mb**. A model is proposed which includes the faster state (Mb*) as a planar porphyrin ring with a displacement of Fe from the heme plane of less than 0.3 Å, and the slower state (Mb**) with a domed heme.