References of "Morsa, Denis"
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See detailCan we correlate ion mobility mass spectrometry data with native solution structures? A crosslinking approach
Hanozin, Emeline ULiege; Grifnée, Elodie ULiege; Morsa, Denis ULiege et al

Poster (2018, June)

Introduction: The structural characterization of biomolecules is of prime importance in the understanding of biological processes at the molecular level. In this context, the development of soft ... [more ▼]

Introduction: The structural characterization of biomolecules is of prime importance in the understanding of biological processes at the molecular level. In this context, the development of soft ionization techniques, namely electrospray ionization, and the advent of ion mobility (IM) separation tools open new perspectives for the analysis of large intact molecular ions by mass spectrometry (MS). Previous (IM-)MS experiments emphasized that the solution phase structures may be (partially) preserved in the gas phase under soft ionization conditions. However, several factors (e.g. coulombic interactions, accelerating voltages) have an influence on the conformation of biomolecular ions in the gas phase and should be carefully considered. Consequently, some fundamental questions remain elusive: “Can we correlate ion mobility mass spectrometry data to native solution structures?” Methods: To bring innovative answers to these longtime debated questions, we here propose an early stage study targeting constrained “native-like” conformations in the gas phase. To this end, we capitalize on a crosslinking approach to establish chemical links between lysine residues located in close spatial proximity within the protein. The so-created covalent network is here used as a scaffold to prevent the collapsing and unfolding of biomolecular ions once desolvated in the gas phase. These “frozen” native solution conformations are then interrogated using ion mobility as implemented on the commercial SYNAPT G2 HDMS spectrometer. Preliminary data: Practically, we used cytochrome c as a model system that we crosslinked with BS3 linkers. Using ion mobility, we monitored the evolution of the collision cross section (CCS) quantity, a rotationally averaged 2D projection of the ion conformation, for each charge state as function of the number of covalent intramolecular linkers. These data were readily compared with those obtained in similar conditions for the non-crosslinked cytochrome c as well as with a benchmark corresponding to the native solution conformations as resolved by NMR spectroscopy. Our results highlight that the crosslinked cytochrome c adopts more compact conformations in the gas phase, closer to values monitored for the native solution conformation, compared to its non-crosslinked homologue. In addition, we found that a critical number of intramolecular linkers were required to prevent structural unfolding from Coulomb repulsions. Novel aspect: Bring innovative answers to the longtime debated native MS questions based on an alternative application of intramolecular covalent linkers. [less ▲]

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See detailPredicting Ion Mobility-Mass Spectrometry Trends of Polymers using the Concept of Apparent Densities
Haler, Jean ULiege; Morsa, Denis ULiege; Lecomte, Philippe ULiege et al

in Methods (2018), 144

Ion Mobility (IM) coupled to Mass Spectrometry (MS) has been used for several decades, bringing a fast separation dimension to the MS detection. IM-MS is a convenient tool for structural elucidation. The ... [more ▼]

Ion Mobility (IM) coupled to Mass Spectrometry (MS) has been used for several decades, bringing a fast separation dimension to the MS detection. IM-MS is a convenient tool for structural elucidation. The fold- ing of macromolecules is often assessed with the support of computational chemistry. However, this strat- egy is strongly dependent on computational initial guesses. Here, we propose the analysis of the Collision Cross-Section (CCS) trends of synthetic homopolymers based on a fitting method which does not rely on computational chemistry a prioris of the three-dimensional structures. The CCS trends were evaluated as a function of the polymer chain length and the charge state. This method is also applicable to mobility trends. It leads to two parameters containing all information available through IM(-MS) measurements. One parameter can be interpreted as an apparent density. The second parameter is related to the shape of the ions and leads us to introduce the concept of trends with constant apparent density. Based on the two fitting parameters, a method for IM trend predictions is elaborated. Experimental deviations from the predictions facilitate detect- ing structural rearrangements and three-dimensional structure differences of the cationized polymer ions. This leads for instance to an easy identification and prediction of the presence of different polymer topologies in complex polymer mixtures. The classification of predicted trends could as well allow for software-assisted data processing. Finally, we suggest the link between the CCS trends of homopolymers and those obtained from (monodisperse) biomolecules to interpret potential folding differences during IM-MS studies. [less ▲]

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See detailWhere Ion Mobility and Molecular Dynamics Meet to Unravel the (Un)Folding Mechanisms of an Oligorotaxane Molecular Switch
Hanozin, Emeline ULiege; Mignolet, Benoît ULiege; Morsa, Denis ULiege et al

in ACS Nano (2017), 11(10), 10253-10263

At the interface between foldamers and mechanically interlocked molecules, oligorotaxanes exhibit a spring-like folded secondary structure with remarkable mechanical and physicochemical properties. Among ... [more ▼]

At the interface between foldamers and mechanically interlocked molecules, oligorotaxanes exhibit a spring-like folded secondary structure with remarkable mechanical and physicochemical properties. Among these properties, the ability of oligorotaxanes to act as molecular switches through controlled modulations of their spatial extension over (un)folding dynamics is of particular interest. The present study aims to assess and further characterize this remarkable feature in the gas phase using mass spectrometry tools. In this context, we focused on the [4]5NPR+12 oligorotaxane molecule complexed with PF6 - counterion and probed its co-conformational states as a function of the in-source-generated charge states. Data were interpreted in light of electronic secondary structure computations at the PM6 and DFT levels. Our results highlight two major co-conformational groups associated either with folded compact structures, notably stabilized by intramolecular π-π interactions and predominant for low charge states or with fully stretched structures resulting from significant Coulombic repulsions at high charge states. Between, the oligorotaxane adopts intermediate folded co-conformations, suggesting a stepwise unfolding pathway under increasing repulsive Coulombic constraints. The reversibility of this superstructural transition was next interrogated under electron-driven (nondissociative electron transfer) and heat-driven (collision-induced unfolding) activation stimuli. The outcomes support the feasibility to either unfold or (partially) refold the oligorotaxane foldamer on purpose in the gas phase. Our results show that the balance between the stabilizing π-π interactions and the versatile Coulomb interactions dictates the elongation state of the foldamer in the gas phase and emphasizes the adequacy of mass spectrometry tools for the superstructural characterization of desolvated prototypical artificial molecular machines. © 2017 American Chemical Society. [less ▲]

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See detailCoulombic driven multi-conformational aspects of oligorotaxane switches studied by ion mobility mass spectrometry and molecular dynamics
Hanozin, Emeline ULiege; Mignolet, Benoît ULiege; Morsa, Denis ULiege et al

Conference (2017, June)

Introduction Artificial Molecular Machines (AMMs), such as Mechanically Interlocked Molecules (MIMs) and foldamers, have recently raised tremendous interest due to their unique properties. Under the ... [more ▼]

Introduction Artificial Molecular Machines (AMMs), such as Mechanically Interlocked Molecules (MIMs) and foldamers, have recently raised tremendous interest due to their unique properties. Under the influence of an appropriate stimuli (pH, redox potential, light…), such molecules are able to reversibly switch between distinct conformational states. Scientists may capitalize on such exclusive properties to get a better understanding of the biomacromolecular level or to design innovative “smart” materials. At the interface between foldamers and MIMs, oligorotaxanes exhibit a spring-like folded secondary structure with remarkable mechanical and physicochemical properties. In the present study, we use ion mobility coupled with mass spectrometry (IM-MS) to probe the conformational states of differentially charged oligorotaxanes in the gas phase. Method Oligorotaxanes are donor-acceptor polymers composed of a π electron-donating dumbbell over which a discrete number of π electron-accepting tetracationic cyclophanes are threaded. The numerous intra-molecular interactions provide them a highly-stabilized rigid rod-like structure in solution. We use IM-MS as implemented in the Synapt G2 HDMS (Waters, Manchester, UK) to investigate the structure of the ionized oligorotaxanes. Our purposes are to probe (i) the different populations of stable conformers generated according to the charge state z and (ii) the reversibility of an electron-driven or thermal-driven conformational change in the gas phase implemented via an electron transfer or collisional activation process prior to the mobility separation. Our experimental observations are supported by electronic structure optimizations at the PM6 and DFT levels coupled with Born-Oppenheimer Molecular Dynamics simulations. Preliminary data Our results highlight a progressive elongation of the oligorotaxane structure with increasing charge numbers until it reaches a maximum extension state. Matching the experimental data with theoretical simulations, we find that the oligorotaxanes adopt an entropically-favored globular shape at low z. As z increases, coulombic repulsions occurring between the cyclophanes gradually outweigh the stabilizing π-stacking interactions and force the structure to elongate. This process occurs in a multistep fashion, each corresponding to a distinct group of helical-shaped conformers, before it eventually results in a fully stretched structure. On the other hand, our results also highlight that a charge reduction driven by a non-dissociative electron transfer process leads to a refolding of the structure so that it adopts a size similar to its electrospray-generated counterpart when the appropriate number of electrons is added. This observation may be imparted to the gradual decrease of the Coulomb repulsions between the cyclophanes mediated through increasing numbers of transferred electrons. These results suggests that the transition from one conformer to another is reversible so that the electrostatic balance between the cyclophanes may be used to further tune the structural state adopted by this artificial molecular switch. The second stimulus relied on collisional activation whose inelastic component provides a way to build up energy into the accessible vibrational degrees of freedom. The conformational landscapes of such-activated oligorotaxanes ions were found unchanged in term of collision cross section position but the repartition of population was altered with a promotion of the most elongated conformer, provided the absence of selective fragmentation. Altogether, these results highlight the feasibility of handling the elongation state of oligorotaxanes in the gas phase through appropriate inputs and underline its conformational reversibility properties. Novel aspect Stimuli-induced reversible conformational rearrangements of innovative AMMs studied by IM-MS and molecular dynamics in the gas phase. [less ▲]

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See detailFirst Results Using TIMS on Systems Requiring High IMS Resolution
Haler, Jean ULiege; Massonnet, Philippe ULiege; Morsa, Denis ULiege et al

Conference (2016, June 05)

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See detailContribution of Capillary Electrophoresis and Ion Mobility Spectrometry to probe conformational change during desolvation
Far, Johann ULiege; Delvaux, Cédric ULiege; Kune, Christopher ULiege et al

Conference (2016, January 28)

The transfer of ions from the solution to the gas phase is a critical step to produce « native species ». Coming from a highly solvating medium, ionic species will tend to find a new equilibrium ... [more ▼]

The transfer of ions from the solution to the gas phase is a critical step to produce « native species ». Coming from a highly solvating medium, ionic species will tend to find a new equilibrium conformation in the gas phase. The pathway to reach the thermodynamically stable conformation involves crossing potential barriers of different heights. When these barriers are too high compared to the internal energy of the ions, it will result in “partial memories” (as structural preservation) of the conformation in solution. In order to evaluate the effect of the solvent evaporation and of the various collision processes encountered by the ions in the mass spectrometer, we developed two strategies: The first strategy consists in comparing in a single experiment the shape of the ions in solution and in the gas phase. Data are obtained by coupling Capillary Electrophoresis (CE) with Ion Mobility Mass Spectrometry (IMS). Drift times in solution and in the gas phase are directly compared. Deviations from their correlation points out changes in folding upon desolvatation. Preliminary results show that some of peptides issued from tryptic digest of BSA clearly change their conformation during desolvatation. The second strategy consists to monitor the shape of the different conformers observed for the intact cytochrome C during native runs in capillary electrophoresis and “native” ion mobility mass spectrometry, both using a physiological TRIS acetic pH7 buffer compared to a “MS friendly” ammonium acetate pH7 buffer. Finally as preliminary study, we proposed to use homopolymers for better understanding about the folding behavior in gas phase and the resulting shape of these ions using ion mobility spectrometry. [less ▲]

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See detailCoulombic driven multi-conformational aspects of oligorotaxane switches studied by ion mobility mass spectrometry
Hanozin, Emeline ULiege; Mignolet, Benoît ULiege; Morsa, Denis ULiege et al

Conference (2016)

Molecular machines, such as Mechanically Interlocked Molecules (MIMs) and foldamers, have recently raised tremendous interest due to their unique properties. Under the influence of an appropriate stimuli ... [more ▼]

Molecular machines, such as Mechanically Interlocked Molecules (MIMs) and foldamers, have recently raised tremendous interest due to their unique properties. Under the influence of an appropriate stimuli (pH, redox potential, light…), such molecules are able to reversibly switch between distinct conformational states. Scientists may capitalize on such exclusive properties to get a better understanding of the biomacromolecular level or to design innovative “smart” materials. At the edge of foldamers and MIMs, oligorotaxanes exhibit a spring-like folded secondary structure with remarkable mechanical and physicochemical properties. In the present study, we use ion mobility coupled with mass spectrometry (IM-MS) to probe the conformational states of differentially charged oligorotaxanes in the gas phase. Oligorotaxanes are donor-acceptor polymers composed of a π electron-donating dumbbell over which a discrete number of π electron-accepting tetracationic cyclophanes are threaded. The numerous intra-molecular interactions provide them a highly-stabilized rigid rod-like structure in solution. We presently use IM-MS as implemented in the Synapt G2 HDMS (Waters, Manchester, UK) to investigate the structure of the ionized three-ring oligorotaxane. Our purposes are to probe (i) the different populations of stable conformers generated according to the charge state z and (ii) the reversibility of an electron-driven conformational change in the gas phase implemented via an electron transfer process (ETnoD) prior to the mobility separation. Our experimental observations are supported by electronic structure optimizations at the PM6 and DFT levels coupled with Born-Oppenheimer Molecular Dynamic (BOMD) simulations. Our results highlight a progressive elongation of the oligorotaxane structure with increasing charge numbers until it reaches a plateau. Matching the experimental data with theoretical simulations, we find that the oligorotaxanes adopt an entropically-favored globular shape at low z. As z increases, coulombic repulsions occurring between the cyclophanes gradually outweigh the stabilizing π-stacking interactions and force the structure to elongate. This process occurs in a multistep fashion, each corresponding to a distinct group of conformers, before it eventually results in a fully stretched structure. On the other hand, our results also highlight that a charge reduction driven by the ETnoD process leads to a refolding of the structure so that it adopts a size similar to its electrospray-generated counterpart, therefore attesting of the conformational reversibility of olixorotaxanes in the gas phase. Altogether, our results show that the oligorotaxanes adopt different conformers associated with increasing levels of extension as a function of the charge state. The transition from one conformer to another is reversible so that the electrostatic balance between the cyclophanes may be used to further tune the structural state adopted by this switch in the gas phase. [less ▲]

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See detailDo pi-stacking interactions preserve the solution-state structure un the gas phase? Study of oligorotaxanes by ion mobility mass spectrometry
Hanozin, Emeline ULiege; Mignolet, Benoît ULiege; Morsa, Denis ULiege et al

Conference (2016)

The development of the so-called soft ionization techniques, namely ESI and MALDI, opened new perspectives for the analysis of large intact molecular ions by mass spectrometry (MS) and set the basis of ... [more ▼]

The development of the so-called soft ionization techniques, namely ESI and MALDI, opened new perspectives for the analysis of large intact molecular ions by mass spectrometry (MS) and set the basis of the native MS field. In this context, the conservation of nonbonded interactions maintaining the physiological conformation of an ion through the ESI process is of major interest. Among these, π-stacking interactions are ubiquitous both within biological (DNA) and synthetic (organic electronic devices) systems. In the present investigation we attempt to probe the conservation of π-stacking interactions on oligorotaxanes, a synthetic π-stacked polymers, after its transfer process to the gas phase. The foundations of native mass spectrometry still constitute a controversial topic. In this context, we here investigate this eventuality using ion mobility on electrosprayed mechanically interlocked molecules (MIMS) called oligorotaxanes. These ones are π-stacked polymers composed of a π electron-donating dumbbell over which a discrete number of π electron-accepting tetracationic cyclophanes are threaded5. The numerous intra-molecular interactions confer to these molecules a highly-stabilized rigid rod-like structure and make them ideal candidates to probe the structural fate of ions following the electrospray process. Both the effects of electrostatic repulsions and of collision-driven internal activation on the ion conformation are considered. The resulting isolated ions were then structurally interrogated by means of ion mobility directly hyphenated with MS (Synapt G2 HDMS, Manchester, UK). Dealing with different polymer lengths and charge states, our results highlight a progressive elongation of the oligorotaxane structure with increasing charge density until it reaches a plateau. Using theoretical models built around a linear assemble of monomeric units with tunable sizes as a reference tool, we show that, at low charge states, the “native” π-stacked rod-like structure of oligoratoxanes is expected to be preserved. However, as the charge density increases, coulombic repulsions occurring between the cyclophanes gradually outweigh the stabilizing π-stacking interactions and force the structure to elongate, eventually resulting in the formation of fully stretched structure. Geometries optimized at a PM6 and DFT level also support these conclusions. Meanwhile, collisional activation experiments highlight potent population shifts towards the formation of slightly elongated conformers. They however remain insufficient to promote the formation of fully extended conformations before the fragmentation occurs, emphasizing the high stability of oligorotaxanes. Altogether, our results show that the structure of strongly stabilized oligorotaxanes could be preserved from the solution to the gas phase provided that the electrostatic repulsions and the internal heating are kept minimal. Moreover, our results may also constitute a starting point to quantitatively study the strength of π-stacking interactions in the gas phase. [less ▲]

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See detailIon Mobility-Mass Spectrometry as a Tool for the Structural Characterization of Peptides Bearing Intramolecular Disulfide Bond(s)
Massonnet, Philippe ULiege; Haler, Jean ULiege; Upert, Gregory et al

in Journal of the American Society for Mass Spectrometry (2016), 27(10), 16371646

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See detailEnergetics and Structural Characterization of Isomers Using Ion Mobility and Gas-phase H/D Exchange: Learning from Lasso Peptides
Hanozin, Emeline ULiege; Morsa, Denis ULiege; De Pauw, Edwin ULiege

in Proteomics (2015), early view

State-of-the-art characterization of proteins using mass spectrometry namely relies on fragmentation methods which allows exploring featured dissociative reaction pathways. These pathways are often ... [more ▼]

State-of-the-art characterization of proteins using mass spectrometry namely relies on fragmentation methods which allows exploring featured dissociative reaction pathways. These pathways are often initiated by a series of potentially informative mass-constant conformational changes that are nonetheless frequently overlooked by lack of adequate investigation techniques. In the present study, we propose a methodology to readily address both structural and energetic aspects of stereoisomerization reactions using ion mobility coupled with mass spectrometry. To this end, a commercial spectrometer was used as a reactor comprising an energy resolved collisional activation step intended at promoting controlled conformational changes and a structural assignment step dedicated to the identification of the generated isomers. This identification relies on ion mobility and other on-line coupled techniques, namely an originally designed gas-phase H/D exchange experiment. We here apply this methodology to characterize the isomerization kinetics of capistruin, a 19-residue long lasso-folded peptide. We expect this approach to bring insights into the physical origin of global dissociation thresholds monitored in tandem mass spectrometry experiments and to set a promising basis for quantitative investigations of the stability of different molecular folds. [less ▲]

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See detailIs the natural shape of ions in the gas phase spherical ? The allegory of the cave (Plato) applied in mass spectrometry
Far, Johann ULiege; Haler, Jean ULiege; Kune, Christopher ULiege et al

Conference (2015, May 13)

The transfer of ions from solution to gas phase is a critical step to produce « native species ». Coming from a highly solvating medium, ionic species will tend to find new equilibrium conformations in ... [more ▼]

The transfer of ions from solution to gas phase is a critical step to produce « native species ». Coming from a highly solvating medium, ionic species will tend to find new equilibrium conformations in the gas phase. The pathway to reach the thermodynamically stable conformation(s) involves crossing potential barriers according to the type of interactions involved. When these barriers are too high compared to the internal energy of the ions, it may result in a “partial memory” (as structural preservation) of the conformation in solution. In order to evaluate the effect of the solvent evaporation and of the various collision processes encountered by the ions in the mass spectrometer we based our strategy on the determination of deviations from a spherical shape, supposed to be the natural state of ions in the gas phase. [less ▲]

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