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1. Combined collision-induced dissociation and photo-selected reaction monitoring mass spectrometry modes for simultaneous analysis of coagulation factors and estrogens

Author

Quentin Enjalbert, Marion Girod, Jérémy Jeudy, Jordane Biarc, Romain Simon, Rodolphe Antoine, Philippe Dugourd, Jérôme Lemoine, and Arnaud Salvador

Subjects
Therapeutics. Pharmacology, RM1-950
Abstract

Oral estrogens are directly associated with changes in plasma levels of coagulation proteins. Thus, the detection of any variation in protein concentrations due to estrogen contraceptives, by a simultaneous analysis of both coagulation proteins and estrogens, would be a very informative tool. In the present study, the merit of photo-selected reaction monitoring (SRM), a new analytical tool, was evaluated towards estrogens detection in plasma. Then, SRM and photo-SRM detection modes were combined for the simultaneous analysis of estrogen molecules together with heparin co-factor and factor XIIa, two proteins involved in the coagulation cascade. This study shows that photo-SRM could open new multiplexed analytical routes. Keywords: Estrogen, Coagulation factor protein, Metabolite, Photo-dissociation fragmentation, SRM

Published
2014
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2. Structural basis of protein oxidation resistance: a lysozyme study.

Author

Marion Girod, Quentin Enjalbert, Claire Brunet, Rodolphe Antoine, Jérôme Lemoine, Iva Lukac, Miroslav Radman, Anita Krisko, and Philippe Dugourd

Subjects
Medicine, Science
Abstract

Accumulation of oxidative damage in proteins correlates with aging since it can cause irreversible and progressive degeneration of almost all cellular functions. Apparently, native protein structures have evolved intrinsic resistance to oxidation since perfectly folded proteins are, by large most robust. Here we explore the structural basis of protein resistance to radiation-induced oxidation using chicken egg white lysozyme in the native and misfolded form. We study the differential resistance to oxidative damage of six different parts of native and misfolded lysozyme by a targeted tandem/mass spectrometry approach of its tryptic fragments. The decay of the amount of each lysozyme fragment with increasing radiation dose is found to be a two steps process, characterized by a double exponential evolution of their amounts: the first one can be largely attributed to oxidation of specific amino acids, while the second one corresponds to further degradation of the protein. By correlating these results to the structural parameters computed from molecular dynamics (MD) simulations, we find the protein parts with increased root-mean-square deviation (RMSD) to be more susceptible to modifications. In addition, involvement of amino acid side-chains in hydrogen bonds has a protective effect against oxidation Increased exposure to solvent of individual amino acid side chains correlates with high susceptibility to oxidative and other modifications like side chain fragmentation. Generally, while none of the structural parameters alone can account for the fate of peptides during radiation, together they provide an insight into the relationship between protein structure and susceptibility to oxidation.

Published
2014
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3. Effects of calcium complexation on heparin-like disaccharides. A combined theoretical, tandem mass spectrometry and ultraviolet experiment

Author

Quentin Enjalbert, Daniel Ortiz, Philippe Dugourd, Luke MacAleese, and Jean-Yves Salpin

Subjects
Chemistry, Organic Chemistry, Photodissociation, Analytical chemistry, Tandem mass spectrometry, medicine.disease_cause, Dissociation (chemistry), Analytical Chemistry, Ion, Metal, Deprotonation, Fragmentation (mass spectrometry), visual_art, visual_art.visual_art_medium, medicine, Physical chemistry, Spectroscopy, Ultraviolet
Abstract

Rationale In order to shed light on the influence of the Ca2+ metal cation on the structure of heparin-like (Hp) disaccharides, we have explored the gas-phase structures of both [Hp, −2H]2− and [Ca(Hp), −3H]− ions by coupling experimental and theoretical methods. Methods The goal of this work was to (i) provide new evidence of the metal influence on the Hp structure, which can have important biological consequences, and (ii) to study the usefulness of metal complexation for the analytical distinction of Hp isomers. Collision-induced dissociation (CID) and ultraviolet photodissociation (UVPD) fragments, as well as optical spectra recorded in the gas phase for both [Hp, −2H]2− and [Ca(Hp), −3H]− complexes were compared for I-H, II-S and III-S isomers of Hp. Results In the case of CID fragmentation, a change in the fragmentation pattern was observed upon calcium complexation, with respect to deprotonated Hp. Conclusions Remarkably, when optical spectra are compared in the UV range, the metal effect on the carboxylic group absorption can be detected by an unambiguous blue-shift (~20 nm).

Published
2015
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4. Electron detachment/photodetachment dissociation of lasso peptides

Author

Jean-Claude Tabet, Carlos Afonso, Marie Pérot-Taillandier, Quentin Enjalbert, Rodolphe Antoine, Séverine Zirah, Philippe Dugourd, Sylvie Rebuffat, Richard B. Cole, Molécules de Communication et Adaptation des Micro-organismes (MCAM), Muséum national d'Histoire naturelle (MNHN)-Centre National de la Recherche Scientifique (CNRS), Institut Parisien de Chimie Moléculaire (IPCM), Institut de Chimie du CNRS (INC)-Université Pierre et Marie Curie - Paris 6 (UPMC)-Centre National de la Recherche Scientifique (CNRS), Chimie Organique et Bioorganique : Réactivité et Analyse (COBRA), Institut Normand de Chimie Moléculaire Médicinale et Macromoléculaire (INC3M), Institut de Chimie du CNRS (INC)-École Nationale Supérieure d'Ingénieurs de Caen (ENSICAEN), Normandie Université (NU)-Normandie Université (NU)-Institut national des sciences appliquées Rouen Normandie (INSA Rouen Normandie), Institut National des Sciences Appliquées (INSA)-Normandie Université (NU)-Institut National des Sciences Appliquées (INSA)-Université Le Havre Normandie (ULH), Normandie Université (NU)-Université de Rouen Normandie (UNIROUEN), Normandie Université (NU)-Centre National de la Recherche Scientifique (CNRS)-Université de Caen Normandie (UNICAEN), Normandie Université (NU)-Institut de Chimie du CNRS (INC)-École Nationale Supérieure d'Ingénieurs de Caen (ENSICAEN), Normandie Université (NU)-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie Organique Fine (IRCOF), Université de Rouen Normandie (UNIROUEN), Institut National des Sciences Appliquées (INSA)-Normandie Université (NU)-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS)-Centre National de la Recherche Scientifique (CNRS), Institut Lumière Matière [Villeurbanne] (ILM), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Chimie Structurale Organique et Biologique (CSOB), Institut de Chimie du CNRS (INC)-Université Pierre et Marie Curie - Paris 6 (UPMC)-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)-Université Pierre et Marie Curie - Paris 6 (UPMC)-Centre National de la Recherche Scientifique (CNRS), Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Institut de Chimie Organique Fine (IRCOF), Institut National des Sciences Appliquées (INSA)-Normandie Université (NU)-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS)-Université de Rouen Normandie (UNIROUEN), Institut National des Sciences Appliquées (INSA)-Normandie Université (NU)-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS)-Institut Normand de Chimie Moléculaire Médicinale et Macromoléculaire (INC3M), Université de Caen Normandie (UNICAEN), Normandie Université (NU)-Normandie Université (NU)-École Nationale Supérieure d'Ingénieurs de Caen (ENSICAEN), Normandie Université (NU)-Université Le Havre Normandie (ULH), Normandie Université (NU)-Institut national des sciences appliquées Rouen Normandie (INSA Rouen Normandie), Institut National des Sciences Appliquées (INSA)-Normandie Université (NU)-Institut National des Sciences Appliquées (INSA)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université de Caen Normandie (UNICAEN), Normandie Université (NU)-École Nationale Supérieure d'Ingénieurs de Caen (ENSICAEN), Normandie Université (NU)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Centre National de la Recherche Scientifique (CNRS), and Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)

Subjects
chemistry.chemical_classification, Topoisomer, Collision-induced dissociation, Electron-capture dissociation, Chemistry, Stereochemistry, 010401 analytical chemistry, Peptide, 010402 general chemistry, Condensed Matter Physics, Mass spectrometry, 01 natural sciences, Dissociation (chemistry), 0104 chemical sciences, Fragmentation (mass spectrometry), Covalent bond, Computational chemistry, [CHIM]Chemical Sciences, Physical and Theoretical Chemistry, Instrumentation, Spectroscopy, ComputingMilieux_MISCELLANEOUS
Abstract

Lasso peptides are bioactive peptides produced by bacteria, characterized by a mechanically interlocked topology where the C-terminal tail of the peptide is threaded through and trapped within an N-terminal macrolactam ring. The structural characterization of lasso peptides and differentiation from their unthreaded topoisomers by mass spectrometry are challenging tasks. We previously explored the fragmentation mechanisms of lasso peptides in positive ion mode and showed several signatures of the lasso topology under collision induced dissociation (CID) and electron capture dissociation. Here we analyzed the dissociation of the multiply-deprotonated microcin J25 (MccJ25), a 21-residue lasso peptide produced by Escherichia coli, together with its non-lasso topoisomer and several variants generated by site-directed mutagenesis, under different modes of activation. The fragmentation patterns obtained by CID for the threaded and unthreaded structures were very similar. By contrast, electron detachment dissociation (EDD) as well as activated-electron photodetachment dissociation (a-EPD) revealed very different dissociation pathways for the two topoisomers. The doubly deprotonated topoisomers showed a different deprotonation pattern, Tyr20 residue being deprotonated for MccJ25 only, yielding the singly charged [c19−C2H4O] product ion. MccJ25 also triggered several two-peptide product ions diagnostic of the lasso topology, including the [(c8)*(z2)] species, which could be maintained by either a covalent or a non-covalent linkage.

Published
2015
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5. Effects of calcium complexation on heparin-like disaccharides. A combined theoretical, tandem mass spectrometry and ultraviolet experiment

Author

Daniel, Ortiz, Quentin, Enjalbert, Luke, MacAleese, Philippe, Dugourd, and Jean-Yves, Salpin

Subjects
Isomerism, Models, Chemical, Heparin, Tandem Mass Spectrometry, Calcium, Spectrophotometry, Ultraviolet, Disaccharides
Abstract

In order to shed light on the influence of the Ca(2+) metal cation on the structure of heparin-like (Hp) disaccharides, we have explored the gas-phase structures of both [Hp, -2H](2-) and [Ca(Hp), -3H](-) ions by coupling experimental and theoretical methods.The goal of this work was to (i) provide new evidence of the metal influence on the Hp structure, which can have important biological consequences, and (ii) to study the usefulness of metal complexation for the analytical distinction of Hp isomers. Collision-induced dissociation (CID) and ultraviolet photodissociation (UVPD) fragments, as well as optical spectra recorded in the gas phase for both [Hp, -2H](2-) and [Ca(Hp), -3H](-) complexes were compared for I-H, II-S and III-S isomers of Hp.In the case of CID fragmentation, a change in the fragmentation pattern was observed upon calcium complexation, with respect to deprotonated Hp.Remarkably, when optical spectra are compared in the UV range, the metal effect on the carboxylic group absorption can be detected by an unambiguous blue-shift (~20 nm).

Published
2015

6. Combined collision-induced dissociation and photo-selected reaction monitoring mass spectrometry modes for simultaneous analysis of coagulation factors and estrogens

Author

Jordane Biarc, Quentin Enjalbert, Romain Simon, Rodolphe Antoine, Philippe Dugourd, Marion Girod, Jérôme Lemoine, Jérémy Jeudy, Arnaud Salvador, Institut Lumière Matière [Villeurbanne] (ILM), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), ANABIO-MS - Analyse biomoléculaire par spectrométrie de masse - Biological Analysis by Mass Spectrometry, Institut des Sciences Analytiques (ISA), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), and Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)

Subjects
musculoskeletal diseases, SRM, Collision-induced dissociation, Factor XIIa, medicine.drug_class, Metabolite, Photo-dissociation fragmentation, Analytical chemistry, Coagulation factor protein, Pharmaceutical Science, Pharmacy, Therapeutics, RM1-950, Mass spectrometry, Analytical Chemistry, chemistry.chemical_compound, [CHIM.ANAL]Chemical Sciences/Analytical chemistry, Drug Discovery, Electrochemistry, medicine, Coagulation (water treatment), Spectroscopy, Medicine(all), Pharmacology, Chromatography, Chemistry, Biochemistry, Genetics and Molecular Biology(all), Selected reaction monitoring, lcsh:RM1-950, Plasma levels, Estrogen, lcsh:Therapeutics. Pharmacology, Original Article, sense organs, human activities, hormones, hormone substitutes, and hormone antagonists
Abstract

Oral estrogens are directly associated with changes in plasma levels of coagulation proteins. Thus, the detection of any variation in protein concentrations due to estrogen contraceptives, by a simultaneous analysis of both coagulation proteins and estrogens, would be a very informative tool. In the present study, the merit of photo-selected reaction monitoring (SRM), a new analytical tool, was evaluated towards estrogens detection in plasma. Then, SRM and photo-SRM detection modes were combined for the simultaneous analysis of estrogen molecules together with heparin co-factor and factor XIIa, two proteins involved in the coagulation cascade. This study shows that photo-SRM could open new multiplexed analytical routes. Keywords: Estrogen, Coagulation factor protein, Metabolite, Photo-dissociation fragmentation, SRM

Published
2014
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7. Deciphering the structure of isomeric oligosaccharides in a complex mixture by tandem mass spectrometry: photon activation with vacuum ultra-violet brings unique information and enables definitive structure assignment

Author

Hélène Rogniaux, Quentin Enjalbert, Yann Bittebière, David Ropartz, Philippe Dugourd, Rodolphe Antoine, Marie-Christine Ralet, Alexandre Giuliani, Jérôme Lemoine, INRA, Ctr Rech Angers Nantes, Biopolymers Interact Assemblies UR1268, Rue Geraudiere,BP 71627, F-44316 Nantes 3, France, Biopolymers Interact Assemblies (UR1268), Institut National de la Recherche Agronomique (INRA)-Institut National de la Recherche Agronomique (INRA), ANABIO-MS - Analyse biomoléculaire par spectrométrie de masse - Biological Analysis by Mass Spectrometry, Institut des Sciences Analytiques (ISA), Centre National de la Recherche Scientifique (CNRS)-Université de Lyon-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-École normale supérieure - Lyon (ENS Lyon)-Centre National de la Recherche Scientifique (CNRS)-Université de Lyon-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-École normale supérieure - Lyon (ENS Lyon), Département Caractérisation et Elaboration des Produits Issus de l'Agriculture (CEPIA), Institut National de la Recherche Agronomique (INRA), Synchrotron SOLEIL (SSOLEIL), Centre National de la Recherche Scientifique (CNRS), ANABIO - ANAlyse BIOmoléculaire RMN et spectrométrie de masse, Université de Lyon-École normale supérieure - Lyon (ENS Lyon)-Institut Lumière Matière [Villeurbanne] (ILM), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Institut Lumière Matière [Villeurbanne] (ILM), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), ANR-08-BLAN-0065, ANR-08-BLAN-0065,SR M52,Synchrotron Radiaton for Tandem Mass Spectrometry(2008), Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), and Université de Lyon-Université de Lyon

Subjects
Spectrometry, Mass, Electrospray Ionization, Vacuum, Ultraviolet Rays, Tandem mass spectrometry, Electrospray ionization, Analytical chemistry, Oligosaccharides, GLYCOSAMINOGLYCAN TETRASACCHARIDES, 010402 general chemistry, Mass spectrometry, 01 natural sciences, Biochemistry, Dissociation (chemistry), Analytical Chemistry, Isomers, Fragmentation (mass spectrometry), Isomerism, Computational chemistry, [CHIM.ANAL]Chemical Sciences/Analytical chemistry, Vacuum ultra-violet synchrotron photon activation, Environmental Chemistry, SIALYLATED OLIGOSACCHARIDES, Spectroscopy, Chromatography, High Pressure Liquid, Ions, Chromatography, Reverse-Phase, Photons, Electron-capture dissociation, Chemistry, 010401 analytical chemistry, HEPARIN-DERIVED OLIGOSACCHARIDES, Structure, Oligogalacturonans, PERFORMANCE LIQUID-CHROMATOGRAPHY, 0104 chemical sciences, Electron-transfer dissociation, IONIZATION-TIME, Matrix-assisted laser desorption/ionization, ELECTRON DETACHMENT DISSOCIATION, LINKED GLYCOPEPTIDE IONS, CAPTURE DISSOCIATION, Synchrotrons, KAPPA-CARRAGEENAN, ULTRAVIOLET PHOTODISSOCIATION
Abstract

International audience; Carbohydrates have a wide variety of structures whose complexity and heterogeneity challenge the field of analytical chemistry. Tandem mass spectrometry, with its remarkable sensitivity and high information content, provides key advantages to addressing the structural elucidation of polysaccharides. Yet, classical fragmentation by collision-activated dissociation (CAD) in many cases fails to reach a comprehensive structural determination, especially when isomers have to be differentiated. In this work, for the first time, vacuum ultra-violet (VUV) synchrotron radiation is used as the activation process in tandem mass spectrometry of large oligosaccharides. Compared to low energy CAD (LE-CAD), photon activated dissociation brought more straightforward and valuable structural information. The outstanding feature was that complete series of informative ions were produced, with only minor neutral losses. Moreover, systematic fragmentation rules could be drawn thus facilitating the definitive assignments of fragment identities. As a result, most of the structures present in a complex mixture of oligogalacturonans could be comprehensively resolved, including many isomers differing in the position of methyl groups along the galacturonic acid backbone.

Published
2014
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8. Implementing visible 473 nm photodissociation in a Q-Exactive mass spectrometer: towards specific detection of cysteine-containing peptides

Author

Quentin Enjalbert, Philippe Dugourd, Arnaud Salvador, Rodolphe Antoine, Jordane Biarc, Jérôme Lemoine, Marion Girod, ANABIO-MS - Analyse biomoléculaire par spectrométrie de masse - Biological Analysis by Mass Spectrometry, Institut des Sciences Analytiques (ISA), Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Institut Lumière Matière [Villeurbanne] (ILM), Université Claude Bernard Lyon 1 (UCBL), and European Research Council

Subjects
Proteomics, Quantitative proteomics, Analytical chemistry, Mass spectrometry, 01 natural sciences, Biochemistry, Spectral line, Hydrolysate, Analytical Chemistry, Ion, law.invention, 03 medical and health sciences, Tandem Mass Spectrometry, law, Electrochemistry, Environmental Chemistry, [CHIM]Chemical Sciences, Cysteine, Irradiation, Chromatography, High Pressure Liquid, Spectroscopy, 030304 developmental biology, [PHYS]Physics [physics], 0303 health sciences, Chromatography, Chemistry, 010401 analytical chemistry, Photodissociation, Photochemical Processes, Laser, 0104 chemical sciences, Peptides
Abstract

International audience; Improvement of the fragmentation specificity may streamline data processing of bottom-up proteomic experiments by drastically reducing either the amount of MS/MS data to process in the discovery phase or the detection of interfering signals in targeted quantification. Photodissociation at appropriate wavelengths is a promising alternative technique to the non-discriminating conventional activation mode by collision. Here, we describe the implementation of visible LID at 473 nm in a Q-Exactive-Orbitrap mass spectrometer for the specific detection of cysteine-containing peptides tagged with a Dabcyl group. HCD cell DC offset and irradiation time were optimized to obtain high fragmentation yield and spectra free of contaminating CID product ions, while keeping the irradiation time scale compatible with chromatographic separation. With this optimized experimental set-up, the selective detection of cysteine-containing peptides in a whole tryptic hydrolysate of three combined proteins is demonstrated by comparing all ion fragmentation (AIF) spectra recorded online with and without laser irradiation.

Published
2014
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9. Multiple Electron Ejection from Proteins Resulting from Single-Photon Excitation in the Valence Shell

Author

Quentin Enjalbert, Alexandre Giuliani, Laurent Nahon, Rodolphe Antoine, Philippe Dugourd, Luke MacAleese, Institut Lumière Matière [Villeurbanne] (ILM), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Université de Lyon, Institut National de la Recherche Agronomique (INRA), Synchrotron SOLEIL (SSOLEIL), Centre National de la Recherche Scientifique (CNRS), and European Research Council

Subjects
protein polyanions, Ionic bonding, Photoionization, Electron, Photon energy, energy-range, Ion, symbols.namesake, Ionization, Physics::Atomic and Molecular Clusters, General Materials Science, molecules, vacuum-ultraviolet, Physics::Atomic Physics, Physical and Theoretical Chemistry, photoionization, Physics, negative-ions, Auger effect, charged ions, dynamics, [CHIM.THEO]Chemical Sciences/Theoretical and/or physical chemistry, 13. Climate action, symbols, [PHYS.PHYS.PHYS-CHEM-PH]Physics [physics]/Physics [physics]/Chemical Physics [physics.chem-ph], Atomic physics, Valence electron
Abstract

International audience; One-photon multiple ionization is a signature of dynamical electron correlations in atoms and small molecules, as observed in the Auger process when Auger electron emission follows core-shell ionization. In such a process, the high energy needed to remove several electrons is due to the strong Coulombic attraction between the last departing electron(s) and the ionic core. Multiply negatively charged molecules offer the possibility to overcome the Coulombic attraction, opening the way for multielectron photodetachment following valence shell excitation. Here photodetachment studies have been performed on electrosprayed protein polyanions using vacuum ultraviolet synchrotron radiation coupled to a radiofrequency ion trap. Double, triple, and quadruple electron emissions from protein polyanions resulting from single-photon excitation in the valence shell were observed with ionization thresholds below 20 eV photon energy. This suggests the existence of large electronic correlations in proteins between weakly bound electrons standing on distant sites. Besides, the resulting multiradical polyanions appear to be remarkably stable, an important issue in racliobiology.

Published
2014
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10. Structural basis of protein oxidation resistance: a lysozyme study

Author

Rodolphe Antoine, Philippe Dugourd, Anita Krisko, Quentin Enjalbert, Iva Lukac, Jérôme Lemoine, Claire Brunet, Miroslav Radman, Marion Girod, ANABIO-MS - Analyse biomoléculaire par spectrométrie de masse - Biological Analysis by Mass Spectrometry, Institut des Sciences Analytiques (ISA), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Institut Lumière Matière [Villeurbanne] (ILM), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Mediterranean Institute for Life Sciences [Split, Croatia] (MedILS), Robustesse et évolvabilité de la vie (U1001), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), European Research Council under the European Union's Seventh Framework Programme (FP7/2007-2013 Grant agreement No 320659), European Project, Centre National de la Recherche Scientifique (CNRS)-Université de Lyon-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-École normale supérieure - Lyon (ENS Lyon)-Centre National de la Recherche Scientifique (CNRS)-Université de Lyon-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-École normale supérieure - Lyon (ENS Lyon), Mediterranean Institue of Life Science, Mediterranean Institute of Life Sciences, Sch Pharm & Biomol Sci, Liverpool John Moores University (LJMU), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Descartes - Paris 5 (UPD5), Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon, and Mediterranean Institute for Life Sciences (MedILS)

Subjects
Protein Folding, CONTAINING PEPTIDES, lcsh:Medicine, 2-DIMENSIONAL LIQUID-CHROMATOGRAPHY, Protein oxidation, Biochemistry, Mass Spectrometry, Analytical Chemistry, Oxidative Damage, chemistry.chemical_compound, Protein structure, lcsh:Science, Peptide sequence, AFFINITY, Liquid Chromatography, chemistry.chemical_classification, SITES, Multidisciplinary, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Chromatographic Techniques, LIPID-PEROXIDATION, Amino acid, Chemistry, Liquid Chromatography-Tandem Mass Spectrometry, ESCHERICHIA-COLI, Physical Sciences, Protein folding, Lysozyme, Oxidation-Reduction, [CHIM.RADIO]Chemical Sciences/Radiochemistry, Research Article, Egg white, Protein Structure, Liquid Chromatography-Mass Spectrometry, Molecular Sequence Data, Oxidative phosphorylation, Molecular Dynamics Simulation, Research and Analysis Methods, [CHIM.ANAL]Chemical Sciences/Analytical chemistry, CARBONYLATED PROTEINS, Amino Acid Sequence, TANDEM MASS-SPECTROMETRY, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], IDENTIFICATION, lcsh:R, Biology and Life Sciences, Proteins, Protein Structure, Tertiary, chemistry, Gamma Rays, MOLECULAR-DYNAMICS, lcsh:Q, Muramidase
Abstract

International audience; Accumulation of oxidative damage in proteins correlates with aging since it can cause irreversible and progressive degeneration of almost all cellular functions. Apparently, native protein structures have evolved intrinsic resistance to oxidation since perfectly folded proteins are, by large most robust. Here we explore the structural basis of protein resistance to radiation-induced oxidation using chicken egg white lysozyme in the native and misfolded form. We study the differential resistance to oxidative damage of six different parts of native and misfolded lysozyme by a targeted tandem/mass spectrometry approach of its tryptic fragments. The decay of the amount of each lysozyme fragment with increasing radiation dose is found to be a two steps process, characterized by a double exponential evolution of their amounts: the first one can be largely attributed to oxidation of specific amino acids, while the second one corresponds to further degradation of the protein. By correlating these results to the structural parameters computed from molecular dynamics (MD) simulations, we find the protein parts with increased root-mean-square deviation (RMSD) to be more susceptible to modifications. In addition, involvement of amino acid side-chains in hydrogen bonds has a protective effect against oxidation Increased exposure to solvent of individual amino acid side chains correlates with high susceptibility to oxidative and other modifications like side chain fragmentation. Generally, while none of the structural parameters alone can account for the fate of peptides during radiation, together they provide an insight into the relationship between protein structure and susceptibility to oxidation.

Published
2014
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11. Vacuum ultraviolet action spectroscopy of polysaccharides

Author

A. Vernier, Philippe Dugourd, Quentin Enjalbert, Abdul-Rahman Allouche, Jérôme Lemoine, Rodolphe Antoine, Claire Brunet, Alexandre Giuliani, Laurent Nahon, Institut des Sciences Analytiques (ISA), Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Institut Lumière Matière [Villeurbanne] (ILM), Université Claude Bernard Lyon 1 (UCBL), BioMerieux SA, R&D Prote, bioMérieux SA, ANABIO - ANAlyse BIOmoléculaire RMN et spectrométrie de masse (2011-2013), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Synchrotron SOLEIL (SSOLEIL), Centre National de la Recherche Scientifique (CNRS), Département Caractérisation et Elaboration des Produits Issus de l'Agriculture (CEPIA), Institut National de la Recherche Agronomique (INRA), SOLEIL support project #20110002, Agence Nationale de la Recherche Scientifique ANR-08-BLAN-0065, École normale supérieure - Lyon (ENS Lyon)-Université Claude Bernard Lyon 1 (UCBL), ANABIO - ANAlyse BIOmoléculaire RMN et spectrométrie de masse, Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-École normale supérieure - Lyon (ENS Lyon)-Université Claude Bernard Lyon 1 (UCBL), Centre National de la Recherche Scientifique (CNRS)-Université de Lyon-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-École normale supérieure - Lyon (ENS Lyon), and Université de Lyon-École normale supérieure - Lyon (ENS Lyon)-Centre National de la Recherche Scientifique (CNRS)-Université de Lyon-Université Claude Bernard Lyon 1 (UCBL)

Subjects
CIRCULAR-DICHROISM SPECTROSCOPY, IN-VACUO, Vacuum, Photochemistry, Molecular Sequence Data, Analytical chemistry, Synchrotron radiation, Gas phase, 010402 general chemistry, medicine.disease_cause, Mass spectrometry, 01 natural sciences, GAS-PHASE, Ion, SYNCHROTRON-RADIATION, Isomerism, Structural Biology, Fragmentation, Polysaccharides, [CHIM.ANAL]Chemical Sciences/Analytical chemistry, ELECTRON PHOTODETACHMENT DISSOCIATION, medicine, Physics::Atomic and Molecular Clusters, Glycosides, Quadrupole ion trap, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], Spectroscopy, BASIS-SETS, LASER-INDUCED PHOTOFRAGMENTATION, Chemistry, 010401 analytical chemistry, Vacuum ultraviolet, OPTICAL-PROPERTIES, MASS-SPECTROMETRY, 0104 chemical sciences, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], PROTEIN POLYANIONS, Carbohydrate Sequence, Beamline, Atomic electron transition, Physics::Accelerator Physics, Spectrophotometry, Ultraviolet, Synchrotrons, Ultraviolet
Abstract

International audience; We studied the optical properties of gas-phase polysaccharides (maltose, maltotetraose, and maltohexaose) ions by action spectroscopy using the coupling between a quadrupole ion trap and a vacuum ultraviolet (VUV) beamline at the SOLEIL synchrotron radiation facility (France) in the 7 to 18 eV range. The spectra provide unique benchmarks for evaluation of theoretical data on electronic transitions of model carbohydrates in the VUV range. The effects of the nature of the charge held by polysaccharide ions on the relaxation processes were also explored. Finally the effect of isomerization of polysaccharides (with melezitose and raffinose) on their photofragmentation with VUV photons is presented.

Published
2013
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12. Improved detection specificity for plasma proteins by targeting cysteine-containing peptides with photo-SRM

Author

Romain Simon, Quentin Enjalbert, Jérôme Lemoine, Marion Girod, Fabien Chirot, Rodolphe Antoine, Philippe Dugourd, Jérémy Jeudy, Arnaud Salvador, ANABIO - ANAlyse BIOmoléculaire RMN et spectrométrie de masse (2011-2013), Institut des Sciences Analytiques (ISA), Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Institut Lumière Matière [Villeurbanne] (ILM), Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon, the French Agence National de la Recherche - photo-SRM project (ANR-11-BSV5-003-01)., ANABIO-MS - Analyse biomoléculaire par spectrométrie de masse - Biological Analysis by Mass Spectrometry, Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), and Bussy, Agnès

Subjects
musculoskeletal diseases, [CHIM.ANAL] Chemical Sciences/Analytical chemistry, Quantitative proteomics, QUANTITATIVE PROTEOMICS STRATEGY, ABUNDANCE PROTEINS, Peptide, 010402 general chemistry, Mass spectrometry, Tandem mass spectrometry, 01 natural sciences, Biochemistry, Sensitivity and Specificity, GAS-PHASE, Mass Spectrometry, Analytical Chemistry, [CHIM.ANAL]Chemical Sciences/Analytical chemistry, ELECTRON PHOTODETACHMENT DISSOCIATION, Animals, Humans, Cysteine, TANDEM MASS-SPECTROMETRY, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], ComputingMilieux_MISCELLANEOUS, chemistry.chemical_classification, Chromophore derivatization, Chromatography, Photo-SRM, QUADRUPOLE ION-TRAP, Chemistry, 010401 analytical chemistry, Selected reaction monitoring, Plasma proteins, Blood Proteins, QUANTIFICATION, Blood proteins, 0104 chemical sciences, Rats, PROSTATE-SPECIFIC ANTIGEN, Targeted mass spectrometry, Proteome, Cysteine-containing peptides quantification, ISOTOPE-DILUTION, Peptides, human activities, ULTRAVIOLET PHOTODISSOCIATION
Abstract

International audience; Targeted mass spectrometry using selected reaction monitoring (SRM) has emerged as an alternative to immunoassays for protein quantification owing to faster development time and higher multiplexing capability. However, the SRM strategy is faced with the high complexity of peptide mixtures after trypsin digestion of whole plasma or the cellular proteome that most of the time causes contamination, irremediably, by interfering compounds in the transition channels monitored. This problem becomes increasingly acute when the targeted protein is present at a low concentration. In this work, the merit of laser-induced photo-dissociation in the visible region at 473 nm implemented in an hybrid quadrupole linear ion-trap mass spectrometer (photo-SRM) was evaluated for detection specificity of cysteine-containing peptides in a group of plasma proteins after tagging with a dabcyl chromophore. Compared with conventional SRM, photo-SRM chromatograms have improved detection specificity for most of peptides monitored. Comparison of the signals obtained for the best proteotypic peptides in SRM mode and those recorded by photo-SRM of cysteine-containing peptides for the same proteins reveals either increased (up to 10-fold) or similar signal to photo-SRM detection. Finally, photo-SRM has extended response linearity across a calibration plot obtained by diluting human plasma in rat plasma, down to the lowest concentrations. Hence, photo-SRM may advantageously complement conventional SRM in assay of proteins in complex biological matrices.

Published
2013
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13. Prompt and slow electron-detachment-dissociation/electron-photodetachment-dissociation of a 21-mer Peptide

Author

Séverine Zirah, Jérôme Lemoine, Carlos Afonso, Rodolphe Antoine, Quentin Enjalbert, Marie Pérot-Taillandier, Jean-Claude Tabet, Philippe Dugourd, Sylvie Rebuffat, Institut Parisien de Chimie Moléculaire (IPCM), Université Pierre et Marie Curie - Paris 6 (UPMC)-Centre National de la Recherche Scientifique (CNRS), Molécules de Communication et Adaptation des Micro-organismes (MCAM), Muséum national d'Histoire naturelle (MNHN)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Spectrométrie Ionique et Moléculaire (LASIM), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Institut Lumière Matière [Villeurbanne] (ILM), ANABIO - ANAlyse BIOmoléculaire RMN et spectrométrie de masse (2011-2013), Institut des Sciences Analytiques (ISA), Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Chimie Organique et Bioorganique : Réactivité et Analyse (COBRA), Institut Normand de Chimie Moléculaire Médicinale et Macromoléculaire (INC3M), Institut de Chimie du CNRS (INC)-École Nationale Supérieure d'Ingénieurs de Caen (ENSICAEN), Normandie Université (NU)-Normandie Université (NU)-Institut national des sciences appliquées Rouen Normandie (INSA Rouen Normandie), Institut National des Sciences Appliquées (INSA)-Normandie Université (NU)-Institut National des Sciences Appliquées (INSA)-Université Le Havre Normandie (ULH), Normandie Université (NU)-Université de Rouen Normandie (UNIROUEN), Normandie Université (NU)-Centre National de la Recherche Scientifique (CNRS)-Université de Caen Normandie (UNICAEN), Normandie Université (NU)-Institut de Chimie du CNRS (INC)-École Nationale Supérieure d'Ingénieurs de Caen (ENSICAEN), Normandie Université (NU)-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie Organique Fine (IRCOF), Université de Rouen Normandie (UNIROUEN), Institut National des Sciences Appliquées (INSA)-Normandie Université (NU)-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS)-Centre National de la Recherche Scientifique (CNRS), Histoire naturelle de l'Homme préhistorique (HNHP), Centre National de la Recherche Scientifique (CNRS)-Muséum national d'Histoire naturelle (MNHN)-Université de Perpignan Via Domitia (UPVD), École normale supérieure - Lyon (ENS Lyon)-Université Claude Bernard Lyon 1 (UCBL), ANABIO - ANAlyse BIOmoléculaire RMN et spectrométrie de masse, Centre National de la Recherche Scientifique (CNRS)-Université de Lyon-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-École normale supérieure - Lyon (ENS Lyon)-Centre National de la Recherche Scientifique (CNRS)-Université de Lyon-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-École normale supérieure - Lyon (ENS Lyon), Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie Organique Fine (IRCOF), Normandie Université (NU)-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS)-Université de Rouen Normandie (UNIROUEN), Normandie Université (NU)-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Institut de Chimie Organique Fine (IRCOF), Institut National des Sciences Appliquées (INSA)-Normandie Université (NU)-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS)-Université de Rouen Normandie (UNIROUEN), Institut National des Sciences Appliquées (INSA)-Normandie Université (NU)-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS)-Institut Normand de Chimie Moléculaire Médicinale et Macromoléculaire (INC3M), Université de Caen Normandie (UNICAEN), Normandie Université (NU)-Normandie Université (NU)-École Nationale Supérieure d'Ingénieurs de Caen (ENSICAEN), Normandie Université (NU)-Université Le Havre Normandie (ULH), Normandie Université (NU)-Institut national des sciences appliquées Rouen Normandie (INSA Rouen Normandie), Institut National des Sciences Appliquées (INSA)-Normandie Université (NU)-Institut National des Sciences Appliquées (INSA)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université de Caen Normandie (UNICAEN), Normandie Université (NU)-École Nationale Supérieure d'Ingénieurs de Caen (ENSICAEN), Normandie Université (NU)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Centre National de la Recherche Scientifique (CNRS), Muséum national d'Histoire naturelle (MNHN)-Université de Perpignan Via Domitia (UPVD)-Centre National de la Recherche Scientifique (CNRS), and Institut National des Sciences Appliquées (INSA)-Normandie Université (NU)-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS)

Subjects
Kinetics, Molecular Sequence Data, Analytical chemistry, Peptide, Electrons, Electron, 010402 general chemistry, Mass spectrometry, 01 natural sciences, Catalysis, Dissociation (chemistry), Mass Spectrometry, Ion, Deprotonation, [CHIM.ANAL]Chemical Sciences/Analytical chemistry, Side chain, Amino Acid Sequence, chemistry.chemical_classification, Chemistry, 010401 analytical chemistry, Organic Chemistry, General Chemistry, Photochemical Processes, 0104 chemical sciences, body regions, Physical chemistry, Peptides
Abstract

International audience; Electron detachment dissociation (EDD) and electron photodetachment dissociation (EPD) are relatively new dissociation methods that involve electron detachment followed by radical-driven dissociation from multiply deprotonated species. EDD yields prompt dissociation whereas only electron detachment is obtained by EPD; subsequent vibrational activation of the charge-reduced radical anion is required to obtain the product ions. Herein, the fragmentation patterns that were obtained by EDD and by vibrational activation of the charge-reduced radical anions that were produced through EDD or EPD (activated-EDD and activated-EPD) were compared. The observed differences were related to the dissociation kinetics and/or the contribution of electron-induced dissociation (EID). Time-resolved double-resonance experiments were performed to measure the dissociation rate constants of the EDD product ions. Differences in the formation kinetics were revealed between the classical EDD/EPD 'a(.) (i) /''x(j) complementary ions and some 'a(.) (i) /c(i) /'''z(.) (j) product ions, which were produced with slower dissociation rate constants, owing to the presence of specific neighbouring side chains. A new fragmentation pathway is proposed for the formation of the slow-kinetics 'a(.) (i) ions.

Published
2013
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14. Visible Photodissociation in a Q-Exactive mass spectrometer: specific detection of cysteine-containing peptides

Author

Marion GIROD, Jordane Biarc, Quentin Enjalbert, Jéremy Jeudy, Rodolphe Antoine, Jérôme Lemoine, Philippe Dugourd, Bussy, Agnès, ANABIO-MS - Analyse biomoléculaire par spectrométrie de masse - Biological Analysis by Mass Spectrometry, Institut des Sciences Analytiques (ISA), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Institut Lumière Matière [Villeurbanne] (ILM), and Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)

Subjects
[CHIM.ANAL] Chemical Sciences/Analytical chemistry, [CHIM.ANAL]Chemical Sciences/Analytical chemistry, ComputingMilieux_MISCELLANEOUS
Abstract

communication par affiche; International audience

Published
2013

15. Evaluation of hydrophilic interaction chromatography (HILIC) versus C-18 reversed-phase chromatography for targeted quantification of peptides by mass spectrometry

Author

Jordane Biarc, Romain Simon, Jérôme Lemoine, Arnaud Salvador, Quentin Enjalbert, ANABIO - ANAlyse BIOmoléculaire RMN et spectrométrie de masse (2011-2013), Institut des Sciences Analytiques (ISA), Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), and Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)

Subjects
SRM, ELECTROSPRAY-IONIZATION, Electrospray ionization, BIOMARKERS, Analytical chemistry, ABUNDANCE PROTEINS, Context (language use), Peptide, 2-DIMENSIONAL LIQUID-CHROMATOGRAPHY, 010402 general chemistry, Mass spectrometry, 01 natural sciences, Biochemistry, Analytical Chemistry, Quantitation, SERUM, VERIFICATION, [CHIM.ANAL]Chemical Sciences/Analytical chemistry, ASSAYS, HILIC, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], chemistry.chemical_classification, Chromatography, PLASMA, CHALLENGES, Chemistry, Hydrophilic interaction chromatography, Protein, 010401 analytical chemistry, Organic Chemistry, Selected reaction monitoring, General Medicine, Reversed-phase chromatography, Small molecule, 0104 chemical sciences, SEPARATION, Peptides, Supercharging reagents
Abstract

International audience; Hydrophilic-interaction liquid chromatography (HILIC) is a widely used technique for small polar molecule analysis and offers the advantage of improved sensitivity in mass spectrometry. Although HILIC is today frequently employed as an orthogonal fractionation method for peptides during the proteomic discovery phase, it is still seldom considered for quantification. In this study, the performances in terms of peak capacity and sensitivity of 3 HILIC columns were compared to traditional reversed phase liquid C-18 column in the context of targeted quantification of proteotypic peptides using selected reaction monitoring mode (SRM). The results showed that the maximum sensitivity in HILIC chromatography was achieved by using an amide column without salt buffer and that the signal increased compared to classic reversed phase chromatography. However, the intensity improvement is quite low compared to the one obtained for small molecules. This is due on one hand to a higher matrix effect in HILIC and on the other hand to a change of charge states of peptides in organic solvent (doubly charged to monocharged). The doubly charged ions can be more readily dissociated than singly charged ions, making them ideal for SRM peptide quantification. As a result "supercharging" reagents are added to the mobile phase to shift from predominant singly charged ions to the more favorable doubly charged species. Using such optimized conditions, peptide signal is improved by a factor of between two and ten for 88% of the peptides of the 81 peptides investigated

Published
2012
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16. Evaluation of hydrophilic interaction chromatography (HILIC) versus C₁₈ reversed-phase chromatography for targeted quantification of peptides by mass spectrometry

Author

Romain, Simon, Quentin, Enjalbert, Jordane, Biarc, Jérôme, Lemoine, and Arnaud, Salvador

Subjects
Humans, Female, Peptides, Mass Spectrometry, Chromatography, Liquid
Abstract

Hydrophilic-interaction liquid chromatography (HILIC) is a widely used technique for small polar molecule analysis and offers the advantage of improved sensitivity in mass spectrometry. Although HILIC is today frequently employed as an orthogonal fractionation method for peptides during the proteomic discovery phase, it is still seldom considered for quantification. In this study, the performances in terms of peak capacity and sensitivity of 3 HILIC columns were compared to traditional reversed phase liquid C(18) column in the context of targeted quantification of proteotypic peptides using selected reaction monitoring mode (SRM). The results showed that the maximum sensitivity in HILIC chromatography was achieved by using an amide column without salt buffer and that the signal increased compared to classic reversed phase chromatography. However, the intensity improvement is quite low compared to the one obtained for small molecules. This is due on one hand to a higher matrix effect in HILIC and on the other hand to a change of charge states of peptides in organic solvent (doubly charged to monocharged). The doubly charged ions can be more readily dissociated than singly charged ions, making them ideal for SRM peptide quantification. As a result "supercharging" reagents are added to the mobile phase to shift from predominant singly charged ions to the more favorable doubly charged species. Using such optimized conditions, peptide signal is improved by a factor of between two and ten for 88% of the peptides of the 81 peptides investigated.

Published
2012

17. Optical properties of a visible push-pull chromophore covalently bound to carbohydrates: solution and gas-phase spectroscopy combined to theoretical Investigations

Author

Claire Loison, Sébastien Redon, Rodolphe Antoine, Amandine Racaud, Quentin Enjalbert, Yann Bretonnière, Chantal Andraud, Stéphane Chambert, Philippe Dugourd, Jérôme Lemoine, Mehmet Menaf Ayhan, ANABIO - ANAlyse BIOmoléculaire RMN et spectrométrie de masse (2011-2013), Institut des Sciences Analytiques (ISA), Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Spectrométrie Ionique et Moléculaire (LASIM), Université Claude Bernard Lyon 1 (UCBL), Institut de Chimie et Biochimie Moléculaires et Supramoléculaires (ICBMS), Université de Lyon-Université de Lyon-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut de Chimie du CNRS (INC)-École Supérieure Chimie Physique Électronique de Lyon-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Chimie - UMR5182 (LC), Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-École normale supérieure - Lyon (ENS Lyon)-Institut de Chimie du CNRS (INC), Spectrobio, Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), the National Research Agency (ANR-09-JCJC-0077-01), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Université de Lyon-Université de Lyon-École Supérieure de Chimie Physique Électronique de Lyon (CPE)-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), and École normale supérieure de Lyon (ENS de Lyon)-Université Claude Bernard Lyon 1 (UCBL)

Subjects
Absorption spectroscopy, BIOMOLECULAR SYSTEMS, Carbohydrates, Molecular Dynamics Simulation, LABELED OLIGOSACCHARIDES, 010402 general chemistry, Photochemistry, 01 natural sciences, [CHIM.ANAL]Chemical Sciences/Analytical chemistry, Materials Chemistry, Molecule, Physical and Theoretical Chemistry, Quadrupole ion trap, Maltose, Spectroscopy, ATOM FORCE-FIELD, LASER-INDUCED PHOTOFRAGMENTATION, 010405 organic chemistry, Chemistry, Photodissociation, RESP MODEL, Solvation, Models, Theoretical, Chromophore, SIMULATIONS, 0104 chemical sciences, Surfaces, Coatings and Films, Solutions, [CHIM.THEO]Chemical Sciences/Theoretical and/or physical chemistry, MOLECULAR-DYNAMICS, Spectrophotometry, Ultraviolet, Gases, BIOSENSORS, EXCITATION-ENERGIES, Absorption (chemistry), ULTRAVIOLET PHOTODISSOCIATION
Abstract

International audience; The use of visible absorbing and fluorescent tags for sensing and structural analysis of carbohydrates is a promising route in a variety of medical, diagnostic, and therapeutic contexts. Here we report an easy method for covalent attachment of nonfluorescent push-pull chromophores based on the 4-cyano-5-dicyanomethylene-2-oxo-3-pyrroline ring to carbohydrate moieties. The impact of sugar grafting on the optical properties of the push-pull chromophore in the gas phase and in solution was investigated by absorption and action spectroscopy and theoretical methods. The labeled sugars efficiently absorb photons in the visible range, as demonstrated by their intense photodissociation in a quadrupole ion trap. A strong blue shift (-70 nm) of the gas-phase photodissociation intensity maximum is observed upon sugar grafting, whereas no such effect is visible on the solution absorption spectra. Molecular dynamics simulations of labeled maltose in the gas phase describe strong interactions between the sulfonated chromophore and the carbohydrate, which lead to cyclic conformations. These are not observed in the simulations with explicit solvation. Time-dependent density functional theory (TD-DFT) calculations on model molecules permit us to attribute the observed shift to the formation of such cyclic conformations and to the displacement of the negative charge relative to the aromatic moiety of the chromophore.

Published
2012
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18. Photo-SRM: laser-induced dissociation improves detection selectivity of Selected Reaction Monitoring mode

Author

Sébastien Redon, Rodolphe Antoine, Arnaud Salvador, Mehmet Menaf Ayhan, Quentin Enjalbert, Jérôme Lemoine, Stéphane Chambert, Yann Bretonnière, Philippe Dugourd, Florence Darbour, Romain Simon, ANABIO - ANAlyse BIOmoléculaire RMN et spectrométrie de masse (2011-2013), Institut des Sciences Analytiques (ISA), Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Spectrométrie Ionique et Moléculaire (LASIM), Université Claude Bernard Lyon 1 (UCBL), Institut de Chimie et Biochimie Moléculaires et Supramoléculaires (ICBMS), Université de Lyon-Université de Lyon-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut de Chimie du CNRS (INC)-École Supérieure Chimie Physique Électronique de Lyon-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Chimie - UMR5182 (LC), Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-École normale supérieure - Lyon (ENS Lyon)-Institut de Chimie du CNRS (INC), the National Research Agency (ANR-09-JCJC-0077-01), the 'Service de Cooperation et d'Action Culturelle de l'Ambassade de France en Turquie' (MMA)., Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Université de Lyon-Université de Lyon-École Supérieure de Chimie Physique Électronique de Lyon (CPE)-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), and École normale supérieure de Lyon (ENS de Lyon)-Université Claude Bernard Lyon 1 (UCBL)

Subjects
Models, Molecular, Sucrose, Calibration curve, Analytical chemistry, MESH: Trypsin, Tandem mass spectrometry, Oxytocin, 01 natural sciences, Mass Spectrometry, Analytical Chemistry, DERIVATIZATION, MESH: Blood Proteins, Trypsin, MESH: Peptide Fragments, Spectroscopy, Chemistry, PEPTIDES, Blood Proteins, Photochemical Processes, Small molecule, MESH: Lasers, MESH: Models, Molecular, QUANTITATION, MESH: Ions, Ion-mobility spectrometry, 010402 general chemistry, Mass spectrometry, Sensitivity and Specificity, [CHIM.ANAL]Chemical Sciences/Analytical chemistry, ELECTRON PHOTODETACHMENT DISSOCIATION, MESH: Oxytocin, Humans, Quadrupole ion trap, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], Detection limit, Ions, MESH: Mass Spectrometry, Chromatography, QUADRUPOLE ION-TRAP, MESH: Humans, POLYANIONS, Lasers, 010401 analytical chemistry, Organic Chemistry, Selected reaction monitoring, MESH: Sucrose, MASS-SPECTROMETRY, Peptide Fragments, MESH: Sensitivity and Specificity, 0104 chemical sciences, PROSTATE-SPECIFIC ANTIGEN, CAPTURE DISSOCIATION, MESH: Photochemical Processes, MESH: Chromatography, Liquid, Chromatography, Liquid, ULTRAVIOLET PHOTODISSOCIATION
Abstract

International audience; Selected Reaction Monitoring (SRM) carried out on triple-quadrupole mass spectrometers coupled to liquid chromatography has been a reference method to develop quantitative analysis of small molecules in biological or environmental matrices for years and is currently emerging as a promising tool in clinical proteomic. However, sensitive assays in complex matrices are often hampered by the presence of co-eluted compounds that share redundant transitions with the target species. On-the-fly better selection of the precursor ion by high-field asymmetric waveform ion mobility spectrometry (FAIMS) or increased quadrupole resolution is one way to escape from interferences. In the present work we document the potential interest of substituting classical gas-collision activation mode by laser-induced dissociation in the visible wavelength range to improve the specificity of the fragmentation step. Optimization of the laser beam pathway across the different quadrupoles to ensure high photo-dissociation yield in Q2 without detectable fragmentation in Q1 was assessed with sucrose tagged with a push-pull chromophore. Next, the proof of concept that photo-SRM ensures more specific detection than does conventional collision-induced dissociation (CID)-based SRM was carried out with oxytocin peptide. Oxytocin was derivatized by the thiol-reactive QSY® 7 C(5)-maleimide quencher on cysteine residues to shift its absorption property into the visible range. Photo-SRM chromatograms of tagged oxytocin spiked in whole human plasma digest showed better detection specificity and sensitivity than CID, that resulted in extended calibration curve linearity. We anticipate that photo-SRM might significantly improve the limit of quantification of classical SRM-based assays targeting cysteine-containing peptides.

Published
2011
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