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Dive into the research topics where Alexandre Giuliani is active.

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Featured researches published by Alexandre Giuliani.


Langmuir | 2013

Aggregation of the Salivary Proline-Rich Protein IB5 in the Presence of the Tannin EgCG

Francis Canon; Franck Paté; Véronique Cheynier; Pascale Sarni-Manchado; Alexandre Giuliani; Javier Pérez; D. Durand; Joaquim Li; Bernard Cabane

In the mouth, proline-rich proteins (PRP), which are major components of stimulated saliva, interact with tannins contained in food. We report in vitro interactions of the tannin epigallocatechin gallate (EgCG), with a basic salivary PRP, IB5, studied through electrospray ionization mass spectrometry (ESI-MS), small-angle X-ray scattering (SAXS), and dynamic light scattering (DLS). In dilute protein (IB5) solutions of low ionic strength (1 mM), the proteins repel each other, and the tannins bind to nonaggregated proteins. ESI-MS experiments determine the populations of nonaggregated proteins that have bound various numbers of tannin molecules. These populations match approximately the Poisson distribution for binding to n = 8 sites on the protein. MS/MS experiments confirm that complexes containing n = 1 to 8 EgCG molecules are dissociated with the same energy. Assuming that the 8 sites are equivalent, we calculate a binding isotherm, with a binding free energy Δμ = 7.26RT(a) (K(d) = 706 μM). In protein solutions that are more concentrated (0.21 mM) and at higher ionic strength (50 mM, pH 5.5), the tannins can bridge the proteins together. DLS experiments measure the number of proteins per aggregate. This number rises rapidly when the EgCG concentration exceeds a threshold (0.2 mM EgCG for 0.21 mM of IB5). SAXS experiments indicate that the aggregates have a core-corona structure. The core contains proteins that have bound at least 3 tannins and the corona has proteins with fewer bound tannins. These aggregates coexist with nonaggregated proteins. Increasing the tannin concentration beyond the threshold causes the transfer of proteins to the aggregates and a fast rise of the number of proteins per aggregate. A poisoned growth model explains this fast rise. Very large cationic aggregates, containing up to 10,000 proteins, are formed at tannin concentrations (2 mM) slightly above the aggregation threshold (0.2 mM).


Microscopy and Microanalysis | 2010

Synchrotron UV Fluorescence Microscopy Uncovers New Probes in Cells and Tissues

Frédéric Jamme; Sandrine Villette; Alexandre Giuliani; Valérie Rouam; Frank Wien; Bruno Lagarde; Matthieu Réfrégiers

Use of deep ultraviolet (DUV, below 350 nm) fluorescence opens up new possibilities in biology because it does not need external specific probes or labeling but instead allows use of the intrinsic fluorescence that exists for many biomolecules when excited in this wavelength range. Indeed, observation of label free biomolecules or active drugs ensures that the label will not modify the biolocalization or any of its properties. In the past, it has not been easy to accomplish DUV fluorescence imaging due to limited sources and to microscope optics. Two worlds were coexisting: the spectrofluorometric measurements with full spectrum information with DUV excitation, which lacked high-resolution localization, and the microscopic world with very good spatial resolution but poor spectral resolution for which the wavelength range was limited to 350 nm. To combine the advantages of both worlds, we have developed a DUV fluorescence microscope for cell biology coupled to a synchrotron beamline, providing fine tunable excitation from 180 to 600 nm and full spectrum acquired on each point of the image, to study DUV excited fluorescence emitted from nanovolumes directly inside live cells or tissue biopsies.


Analytical and Bioanalytical Chemistry | 2010

Ability of a salivary intrinsically unstructured protein to bind different tannin targets revealed by mass spectrometry

Francis Canon; Alexandre Giuliani; Franck Paté; Pascale Sarni-Manchado

AbstractAstringency is thought to result from the interaction between salivary proline-rich proteins (PRP) that belong to the intrinsically unstructured protein group (IUP), and tannins, which are phenolic compounds. IUPs have the ability to bind several and/or different targets. At the same time, tannins have different chemical features reported to contribute to the sensation of astringency. The ability of both electrospray ionization mass spectrometry and tandem mass spectrometry to investigate the noncovalent interaction occurring between a human salivary PRP, IB5, and a model tannin, epigallocatechin 3-O-gallate (EgCG), has been reported. Herein, we extend this method to study the effect of tannin chemical features on their interaction with IB5. We used five model tannins, epigallocatechin (EgC), epicatechin 3-O-gallate (ECG), epigallocatechin 3-O-gallate (EgCG), procyanidin dimer B2 and B2 3′-O-gallate, which cover the main tannin chemical features: presence of a gallate moiety (galloylation), the degree of polymerization, and the degree of B ring hydroxylation. We show the ability of IB5 to bind these tannins. We report differences in stoichiometries and in stability of the IB5•1 tannin complexes. These results demonstrate the main role of hydroxyl groups in these interactions and show the involvement of hydrogen bonds. Finally, these results are in line with sensory analysis, by Vidal et al. (J Sci Food Agric 83:564–573, 2003) pointing out that the chain length and the level of galloylation are the main factors affecting astringency perception. FigureCID MS/MS approach to monitor the stability of noncovalent complexes between a human salivary proline-rich protein and model tannins that cover the main chemical features of tannins


Analytical and Bioanalytical Chemistry | 2009

Characterization, stoichiometry, and stability of salivary protein–tannin complexes by ESI-MS and ESI-MS/MS

Francis Canon; Franck Paté; Emmanuelle Meudec; Thérèse Marlin; Véronique Cheynier; Alexandre Giuliani; Pascale Sarni-Manchado

AbstractNumerous protein–polyphenol interactions occur in biological and food domains particularly involving proline-rich proteins, which are representative of the intrinsically unstructured protein group (IUP). Noncovalent protein–ligand complexes are readily detected by electrospray ionization mass spectrometry (ESI-MS), which also gives access to ligand binding stoichiometry. Surprisingly, the study of interactions between polyphenolic molecules and proteins is still an area where ESI-MS has poorly benefited, whereas it has been extensively applied to the detection of noncovalent complexes. Electrospray ionization mass spectrometry has been applied to the detection and the characterization of the complexes formed between tannins and a human salivary proline-rich protein (PRP), namely IB5. The study of the complex stability was achieved by low-energy collision-induced dissociation (CID) measurements, which are commonly implemented using triple quadrupole, hybrid quadrupole time-of-flight, or ion trap instruments. Complexes composed of IB5 bound to a model polyphenol EgCG have been detected by ESI-MS and further analyzed by MS/MS. Mild ESI interface conditions allowed us to observe intact noncovalent PRP–tannin complexes with stoichiometries ranging from 1:1 to 1:5. Thus, ESI-MS shows its efficiency for (1) the study of PRP–tannin interactions, (2) the determination of stoichiometry, and (3) the study of complex stability. We were able to establish unambiguously both their stoichiometries and their overall subunit architecture via tandem mass spectrometry and solution disruption experiments. Our results prove that IB5·EgCG complexes are maintained intact in the gas phase. FigureSchematic illustrating the interaction between IB5 and EgCG, leading to IB5·EgCG complexes that remain intact in the gas phase during ESI-MS analysis. This system provides a model of biological interest with regards to astringency


Journal of Synchrotron Radiation | 2012

VUV synchrotron radiation: a new activation technique for tandem mass spectrometry

Aleksandar R. Milosavljević; Christophe Nicolas; Jean‐François Gil; Francis Canon; Matthieu Réfrégiers; Laurent Nahon; Alexandre Giuliani

A novel experimental technique for tandem mass spectrometry and ion spectroscopy of electrosprayed ions using vacuum-ultraviolet (VUV) synchrotron radiation is presented. Photon activation of trapped precursor ions has been performed by coupling a commercial linear quadrupole ion trap (Thermo scientific LTQ XL), equipped with the electrosprayed ions source, to the DESIRS beamline at the SOLEIL synchrotron radiation facility. The obtained results include, for the first time on biopolymers, photodetachment spectroscopy using monochromated synchrotron radiation of multi-charged anions and the single photon ionization of large charge-selected polycations. The high efficiency and signal-to-noise ratio achieved by the present set-up open up possibilities of using synchrotron light as a new controllable activation method in tandem mass spectrometry of biopolymers and VUV-photon spectroscopy of large biological ions.


Journal of Physical Chemistry Letters | 2012

Gas-phase protein inner-shell spectroscopy by coupling an ion trap with a soft x-ray beamline

Aleksandar R. Milosavljević; Francis Canon; Christophe Nicolas; Catalin Miron; Laurent Nahon; Alexandre Giuliani

C, N, and O near-edge ion yield spectroscopy of 8+ selected electrosprayed cations of cytochrome c protein (12 kDa) has been performed by coupling a linear quadrupole ion trap with a soft X-ray beamline. The photoactivation tandem mass spectra were recorded as a function of the photon energy. Photoionization of the precursor, accompanied by CO2 loss, is the dominant relaxation process, showing high photoion stability following direct or resonant photoionization. The partial ion yields extracted from recorded mass spectra show significantly different behaviors for single and double ionization channels, which can be qualitatively explained by different Auger decay mechanisms. However, the single ionization spectra reveal characteristic structures when compared to existing near-edge X-ray absorption fine structure (NEXAFS) spectra from thin films of peptides and proteins. Therefore, the present experiment opens up new avenues for near-edge X-ray spectroscopy of macromolecules in the gas phase, overcoming the radiation damage issue or the environmental effects as due to the surface, intermolecular interactions, and solvent.


Angewandte Chemie | 2013

Photodissociation and Dissociative Photoionization Mass Spectrometry of Proteins and Noncovalent Protein–Ligand Complexes

Francis Canon; Aleksandar R. Milosavljević; Guillaume van der Rest; Matthieu Réfrégiers; Laurent Nahon; Pascale Sarni-Manchado; Véronique Cheynier; Alexandre Giuliani

Weight of evidence: Using synchrotron radiation, photo-fragmentation of an intrinsically disordered protein is probed and compared with classical tandem mass-spectrometry activation techniques. It provides excellent sequence coverage allowing the identification of the protein noncovalent binding sites.


Nano Research | 2015

X-ray-induced radiophotodynamic therapy (RPDT) using lanthanide micelles: Beyond depth limitations

Slávka Kaščáková; Alexandre Giuliani; Sara Lacerda; Agnès Pallier; Pascal Mercère; Éva Tóth; Matthieu Réfrégiers

We report lanthanide-based micelles integrating hypericin (Hyp) for X-ray-triggered photodynamic therapy (PDT). The lanthanide luminescence induced by X-ray irradiation excites the photosensitizer, which leads to the generation of singlet oxygen. This versatile approach can be extended to other photosensitizers or other types of liponanoparticles and can allow for magnetic resonance imaging (MRI) guidance.


Journal of Chemical Physics | 2002

Electronic excitation and optical cross sections of methylamine and ethylamine in the UV-VUV spectral region

M.-J. Hubin-Franskin; J. Delwiche; Alexandre Giuliani; M.-P. Ska; F Motte-Tollet; Isobel C. Walker; Nigel J. Mason; J. M. Gingell; N. C. Jones

High resolution UV–VUV photon absorption spectra of methylamine and ethylamine have been recorded between 5.0–9.0 eV (250–140 nm) using synchrotron radiation. In methylamine, the energies of the absorption bands are confirmed as centered at 5.7, 7.2, and 8.7 eV, respectively. In ethylamine the band centers are 5.8, 7.0, and 7.9 eV, respectively; the last band is seen here for the first time. Most of the transitions exhibit rich fine structure dominated by vibrational progressions involving excitation of an amino wagging vibration. The absolute photoabsorption oscillator strengths have been measured by photon absorption over the 5–9 eV range and by dipolar electron energy loss spectroscopy from 5–14 eV (250–90 nm).


Journal of Physical Chemistry A | 2011

Gas phase photo-formation and vacuum UV photofragmentation spectroscopy of tryptophan and tyrosine radical-containing peptides.

Claire Brunet; Rodolphe Antoine; A. R. Allouche; Philippe Dugourd; Francis Canon; Alexandre Giuliani; Laurent Nahon

Tryptophan (Trp(•)) and tyrosyl (Tyr(•)) radical containing peptides were produced by UV laser-induced electron detachment from a suitable precursor. Vacuum ultraviolet (VUV) action spectra of these radical peptides were recorded with synchrotron radiation in the 4.5-16 eV range, from which fragmentation pathways and yields are measured as a function of the VUV photon energy. An enhancement in photofragmentation yields of radical species by 1 order of magnitude with respect to nonradical peptides is demonstrated here for the first time. Photofragmentation spectra are compared with absorption spectra for model chromophores calculated in the frame of the time-dependent density functional theory (TDDFT). A qualitative agreement in the position of bands in the 6-8 eV region is observed between experimental photofragmentation and calculated absorption spectra. Photofragmentation spectra of peptide radicals can be useful to better assess the complex deactivation pathways that occur following the absorption of a VUV photon in biomolecular radical anions.

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Francis Canon

Institut national de la recherche agronomique

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Matthieu Réfrégiers

Institut national de la recherche agronomique

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Matthieu Réfrégiers

Institut national de la recherche agronomique

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Guy Cernogora

Centre national de la recherche scientifique

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Frédéric Jamme

Institut national de la recherche agronomique

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