Mass-selected infrared multiple photon spectroscopy (IRMPD), Fourier transform ion cyclotron resonance (FT-ICR) kinetic experiments, RRKM and electronic structure calculations have been performed in order to propose a complex mechanism involving the formation of the proton-bound dimer of water (H5O2+) from 1,1,3,3-tetrafluorodimethyl ether. It has been found that the reaction is facilitated by a series of sequential exothermic bimolecular ion-molecule reactions. Evidence for the dominant mechanistic pathway involving the reaction of CF2H-O=CHF+, an ion of m/z 99, with water is presented. The primary channel occurs via nucleophilic attack of water on the ion of m/z 99 (CF2H-O=CHF+), to lose formyl fluoride and yield protonated difluoromethanol (m/z 69). Association of a second water molecule with protonated difluoromethanol generates a reactive intermediate which decomposes via a 1,4-elimination to release hydrogen fluoride and yield the proton-bound dimer of water and formyl fluoride (m/z 67). The 1,4-elimination of hydrogen fluoride is found to be strongly supported by the results of both RRKM theory and electronic structure calculations. Lastly, the elimination of formyl fluoride occurs by the association of a third water molecule to produce H5O2+ (m/z 37). The most probable isomeric forms of the ions with m/z 99 and 69 were found using IRMPD spectroscopy and electronic structure theory calculations. Thermochemical information for reactant, transition and product species was obtained using MP2/aug-cc-pVQZ//MP2(full)/6-31G(d) level of theory. Ionic hydrogen bond (IHB) interactions, resulting from the association of ammonia and two of the protonated methylxanthine derivatives, caffeine and theophylline, have been characterized using mass-selected IRMPD and electronic structure calculations at the MP2/aug-cc-pVTZ//B3LYP/6-311+G(d,p) level of theory. It was found that the formation of a proton-bound dimer (PBD) of caffeine and ammonia was elusive under the experimental conditions. The low binding energy of the caffeine and ammonia PBD is responsible for the perceived difficulty in obtaining an IRMPD spectrum. The IRMPD spectrum of the PBD of theophylline and ammonia was obtained and revealed bidentate IHB formation within the complex, which greatly increased the binding energy relative to the most stable isomer of the PBD of caffeine and ammonia. The IRMPD spectra of the protonated forms of caffeine and theophylline have also obtained. The spectrum of protonated caffeine showed the dominant existence of a single isomer, whereas the spectrum of protonated theophylline showed a mixture of isomers. The mixture of isomers of protonated theophylline resulted as a consequence of proton-transport catalysis (PTC) occurring within the PBD of theophylline and ammonia. All calculated harmonic spectra have been produced at the B3LYP/6-311+G(d,p) level of theory with fundamental frequencies scaled by 0.9679; calculated anharmonic spectra have also been provided at the same level of theory and were found to greatly improve the match with the IRMPD spectra obtained in all cases. Ionic hydrogen bond (IHB) interactions, resulting from the association of caffeine and theophylline with their protonated counterparts, forming proton-bound homodimers, have been characterized using mass-selected IRMPD and electronic structure calculations at the MP2/6-311+G(2d,2p)//B3LYP/6-311+G(d,p) level of theory. It is found that the IRMPD spectra of the proton-bound homodimers of caffeine and theophylline are complicated resulting from the existence of several pairs of enantiomers separated by a narrow range of relative Gibbs free energies (298 K) of 15.6 and 18.2 kJ mol-1, respectively. The IRMPD spectrum of the proton-bound homodimer of theophylline is dominated by a unique isomer facilitated by formation of a bidentate IHB. Formation of this interaction lowers the relative Gibbs free energy of the ion to 9.75 kJ mol-1 below that of the most favourable pair of enantiomers. The IRMPD spectrum of the PBD of caffeine is complicated by the existence of at least two pairs of enantiomers with the strong likelihood of the spectral contributions of a third pair existing. The most favourable enantiomeric pair involves the formation of a O-H+⋯O IHB. However, verification of a pair of enantiomeric PBDs containing a N-H+⋯O IHB is also observed in the IRMPD spectrum of the PBD of caffeine due to the presence of three free carbonyl stretching modes located at 1731, 1751 and 1785 cm-1. The mass-selected IRMPD spectra of the sodium cation-bound dimers (SCBD) of caffeine and theophylline also have been obtained. Both the mass-selected IRMPD spectra and electronic structure calculations predict the most likely structure of the SCBDs of caffeine and theophylline to form by an efficient O⋯Na+⋯O interaction between C=O functional groups possessed by each monomer. The frequencies of the C=O-Na+ stretch are found to be nearly identical in the IRMPD spectra for both of the SCBDs of caffeine and theophylline at 1644 and 1646 cm-1, respectively. However, the degenerate free C=O symmetric and asymmetric stretches for the SCBDs of caffeine and theophylline found at 1732 and 1758 cm^(-1), respectively, demonstrating a red-shift for caffeine possibly linked to a steric interaction absent in theophylline. Free rotation about the O⋯Na+⋯O bond is found to greatly decrease the complexity of the IRMPD spectra of the SCBDs of caffeine and theophylline and demonstrates excellent agreement between the IRMPD and calculated spectra. Electronic structure calculations have been done at the MP2(full)/aug-cc-pCVTZ/6-311+G(2d,2p)//B3LYP/6-311+G(d,p) level of theory using the aug-cc-pCVTZ basis set for Na+ and all Na+-interacting heterotatoms, and the 6-311+G(2d,2p) basis set for all non-interacting atoms within the SCBDs, in order to provide accurate electronic energies. Currently, installation and implementation of a pulsed electrospray high pressure ion source mated to an existing high pressure mass spectrometer (HPMS) is underway. The new ion source will greatly increase the range of possibilities for the study of ion-molecule reactions in the McMahon laboratory. One of the unique features of the new design is the incorporation of a gas-tight electrospray interface, allowing for more possibilities than only the study of cluster-ion equilibria involving hydration (H2On⋯S+), where S+ is an ion produced by electrospray. Other small prototypical biological molecules such as amines and thiols can be used without concern for the toxicity of these species. Another unique design feature allows electrosprayed ions to associate with neutral solvent species in an electric field free reaction chamber (RC). This ensures that values of equilibrium constants determined are truly representative of ions in states of thermochemical equilibrium. The existing HPMS in the McMahon laboratory is limited to the study of small volatile organic molecules. The new ion source will permit the exploration of systems involving non-volatile species, doubly charged ions and many biologically relevant molecules such as amino acids, peptides, nucleobases and carbohydrates.

Mass-selected infrared multiple photon spectroscopy (IRMPD), Fourier transform ion cyclotron resonance (FT-ICR) kinetic experiments, RRKM and electronic structure calculations have been performed in order to propose a complex mechanism involving the formation of the proton-bound dimer of water (H5O2+) from 1,1,3,3-tetrafluorodimethyl ether. It has been found that the reaction is facilitated by a series of sequential exothermic bimolecular ion-molecule reactions. Evidence for the dominant mechanistic pathway involving the reaction of CF2H-O=CHF+, an ion of m/z 99, with water is presented. The primary channel occurs via nucleophilic attack of water on the ion of m/z 99 (CF2H-O=CHF+), to lose formyl fluoride and yield protonated difluoromethanol (m/z 69). Association of a second water molecule with protonated difluoromethanol generates a reactive intermediate which decomposes via a 1,4-elimination to release hydrogen fluoride and yield the proton-bound dimer of water and formyl fluoride (m/z 67). The 1,4-elimination of hydrogen fluoride is found to be strongly supported by the results of both RRKM theory and electronic structure calculations. Lastly, the elimination of formyl fluoride occurs by the association of a third water molecule to produce H5O2+ (m/z 37). The most probable isomeric forms of the ions with m/z 99 and 69 were found using IRMPD spectroscopy and electronic structure theory calculations. Thermochemical information for reactant, transition and product species was obtained using MP2/aug-cc-pVQZ//MP2(full)/6-31G(d) level of theory. Ionic hydrogen bond (IHB) interactions, resulting from the association of ammonia and two of the protonated methylxanthine derivatives, caffeine and theophylline, have been characterized using mass-selected IRMPD and electronic structure calculations at the MP2/aug-cc-pVTZ//B3LYP/6-311+G(d,p) level of theory. It was found that the formation of a proton-bound dimer (PBD) of caffeine and ammonia was elusive under the experimental conditions. The low binding energy of the caffeine and ammonia PBD is responsible for the perceived difficulty in obtaining an IRMPD spectrum. The IRMPD spectrum of the PBD of theophylline and ammonia was obtained and revealed bidentate IHB formation within the complex, which greatly increased the binding energy relative to the most stable isomer of the PBD of caffeine and ammonia. The IRMPD spectra of the protonated forms of caffeine and theophylline have also obtained. The spectrum of protonated caffeine showed the dominant existence of a single isomer, whereas the spectrum of protonated theophylline showed a mixture of isomers. The mixture of isomers of protonated theophylline resulted as a consequence of proton-transport catalysis (PTC) occurring within the PBD of theophylline and ammonia. All calculated harmonic spectra have been produced at the B3LYP/6-311+G(d,p) level of theory with fundamental frequencies scaled by 0.9679; calculated anharmonic spectra have also been provided at the same level of theory and were found to greatly improve the match with the IRMPD spectra obtained in all cases. Ionic hydrogen bond (IHB) interactions, resulting from the association of caffeine and theophylline with their protonated counterparts, forming proton-bound homodimers, have been characterized using mass-selected IRMPD and electronic structure calculations at the MP2/6-311+G(2d,2p)//B3LYP/6-311+G(d,p) level of theory. It is found that the IRMPD spectra of the proton-bound homodimers of caffeine and theophylline are complicated resulting from the existence of several pairs of enantiomers separated by a narrow range of relative Gibbs free energies (298 K) of 15.6 and 18.2 kJ mol-1, respectively. The IRMPD spectrum of the proton-bound homodimer of theophylline is dominated by a unique isomer facilitated by formation of a bidentate IHB. Formation of this interaction lowers the relative Gibbs free energy of the ion to 9.75 kJ mol-1 below that of the most favourable pair of enantiomers. The IRMPD spectrum of the PBD of caffeine is complicated by the existence of at least two pairs of enantiomers with the strong likelihood of the spectral contributions of a third pair existing. The most favourable enantiomeric pair involves the formation of a O-H+⋯O IHB. However, verification of a pair of enantiomeric PBDs containing a N-H+⋯O IHB is also observed in the IRMPD spectrum of the PBD of caffeine due to the presence of three free carbonyl stretching modes located at 1731, 1751 and 1785 cm-1. The mass-selected IRMPD spectra of the sodium cation-bound dimers (SCBD) of caffeine and theophylline also have been obtained. Both the mass-selected IRMPD spectra and electronic structure calculations predict the most likely structure of the SCBDs of caffeine and theophylline to form by an efficient O⋯Na+⋯O interaction between C=O functional groups possessed by each monomer. The frequencies of the C=O-Na+ stretch are found to be nearly identical in the IRMPD spectra for both of the SCBDs of caffeine and theophylline at 1644 and 1646 cm-1, respectively. However, the degenerate free C=O symmetric and asymmetric stretches for the SCBDs of caffeine and theophylline found at 1732 and 1758 cm^(-1), respectively, demonstrating a red-shift for caffeine possibly linked to a steric interaction absent in theophylline. Free rotation about the O⋯Na+⋯O bond is found to greatly decrease the complexity of the IRMPD spectra of the SCBDs of caffeine and theophylline and demonstrates excellent agreement between the IRMPD and calculated spectra. Electronic structure calculations have been done at the MP2(full)/aug-cc-pCVTZ/6-311+G(2d,2p)//B3LYP/6-311+G(d,p) level of theory using the aug-cc-pCVTZ basis set for Na+ and all Na+-interacting heterotatoms, and the 6-311+G(2d,2p) basis set for all non-interacting atoms within the SCBDs, in order to provide accurate electronic energies. Currently, installation and implementation of a pulsed electrospray high pressure ion source mated to an existing high pressure mass spectrometer (HPMS) is underway. The new ion source will greatly increase the range of possibilities for the study of ion-molecule reactions in the McMahon laboratory. One of the unique features of the new design is the incorporation of a gas-tight electrospray interface, allowing for more possibilities than only the study of cluster-ion equilibria involving hydration (H2On⋯S+), where S+ is an ion produced by electrospray. Other small prototypical biological molecules such as amines and thiols can be used without concern for the toxicity of these species. Another unique design feature allows electrosprayed ions to associate with neutral solvent species in an electric field free reaction chamber (RC). This ensures that values of equilibrium constants determined are truly representative of ions in states of thermochemical equilibrium. The existing HPMS in the McMahon laboratory is limited to the study of small volatile organic molecules. The new ion source will permit the exploration of systems involving non-volatile species, doubly charged ions and many biologically relevant molecules such as amino acids, peptides, nucleobases and carbohydrates.

Proteins have an indispensable role in the cell. They carry out a wide variety of structural, catalytic and signaling functions in all known biological systems. To perform their biological functions, proteins establish interactions with other bioorganic molecules including other proteins. Therefore, protein-protein interactions is one of the central topics in molecular biology. My thesis is devoted to three different topics in the field of protein-protein interactions. The first one focuses on solvent contribution to protein interfaces as it is an important component of protein complexes. The second topic discloses the structural and functional potential of fluorine's unique properties, which are attractive for protein design and engineering not feasible within the scope of canonical amino acids. The last part of this thesis is a study of the impact of charged amino acid residues within the hydrophobic interface of a coiled-coil system, which is one of the well-established model systems for protein-protein interactions studies. I. The majority of proteins interact in vivo in solution, thus studies of solvent impact on protein-protein interactions could be crucial for understanding many processes in the cell. However, though solvent is known to be very important for protein-protein interactions in terms of structure, dynamics and energetics, its effects are often disregarded in computational studies because a detailed solvent description requires complex and computationally demanding approaches. As a consequence, many protein residues, which establish water-mediated interactions, are neither considered in an interface definition. In the previous work carried out in our group the protein interfaces database (SCOWLP) has been developed. This database takes into account interfacial solvent and based on this classifies all interfacial protein residues of the PDB into three classes based on their interacting properties: dry (direct interaction), dual (direct and water-mediated interactions), and wet spots (residues interacting only through one water molecule). To define an interaction SCOWLP considers a donor–acceptor distance for hydrogen bonds of 3.2 Å, for salt bridges of 4 Å, and for van der Waals contacts the sum of the van der Waals radii of the interacting atoms. In previous studies of the group, statistical analysis of a non-redundant protein structure dataset showed that 40.1% of the interfacial residues participate in water-mediated interactions, and that 14.5% of the total residues in interfaces are wet spots. Moreover, wet spots have been shown to display similar characteristics to residues contacting water molecules in cores or cavities of proteins. The goals of this part of the thesis were: 1. to characterize the impact of solvent in protein-protein interactions 2. to elucidate possible effects of solvent inclusion into the correlated mutations approach for protein contacts prediction To study solvent impact on protein interfaces a molecular dynamics (MD) approach has been used. This part of the work is elaborated in section 2.1 of this thesis. We have characterized properties of water-mediated protein interactions at residue and solvent level. For this purpose, an MD analysis of 17 representative complexes from SH3 and immunoglobulin protein families has been performed. We have shown that the interfacial residues interacting through a single water molecule (wet spots) are energetically and dynamically very similar to other interfacial residues. At the same time, water molecules mediating protein interactions have been found to be significantly less mobile than surface solvent in terms of residence time. Calculated free energies indicate that these water molecules should significantly affect formation and stability of a protein-protein complex. The results obtained in this part of the work also suggest that water molecules in protein interfaces contribute to the conservation of protein interactions by allowing more sequence variability in the interacting partners, which has important implications for the use of the correlated mutations concept in protein interactions studies. This concept is based on the assumption that interacting protein residues co-evolve, so that a mutation in one of the interacting counterparts is compensated by a mutation in the other. The study presented in section 2.2 has been carried out to prove that an explicit introduction of solvent into the correlated mutations concept indeed yields qualitative improvement of existing approaches. For this, we have used the data on interfacial solvent obtained from the SCOWLP database (the whole PDB) to construct a “wet” similarity matrix. This matrix has been used for prediction of protein contacts together with a well-established “dry” matrix. We have analyzed two datasets containing 50 domains and 10 domain pairs, and have compared the results obtained by using several combinations of both “dry” and “wet” matrices. We have found that for predictions for both intra- and interdomain contacts the introduction of a combination of a “dry” and a “wet” similarity matrix improves the predictions in comparison to the “dry” one alone. Our analysis opens up the idea that the consideration of water may have an impact on the improvement of the contact predictions obtained by correlated mutations approaches. There are two principally novel aspects in this study in the context of the used correlated mutations methodology : i) the first introduction of solvent explicitly into the correlated mutations approach; ii) the use of the definition of protein-protein interfaces, which is essentially different from many other works in the field because of taking into account physico-chemical properties of amino acids and not being exclusively based on distance cut-offs. II. The second part of the thesis is focused on properties of fluorinated amino acids in protein environments. In general, non-canonical amino acids with newly designed side-chain functionalities are powerful tools that can be used to improve structural, catalytic, kinetic and thermodynamic properties of peptides and proteins, which otherwise are not feasible within the use of canonical amino acids. In this context fluorinated amino acids have increasingly gained in importance in protein chemistry because of fluorine's unique properties: high electronegativity and a small atomic size. Despite the wide use of fluorine in drug design, properties of fluorine in protein environments have not been yet extensively studied. The aims of this part of the dissertation were: 1. to analyze the basic properties of fluorinated amino acids such as electrostatic and geometric characteristics, hydrogen bonding abilities, hydration properties and conformational preferences (section 3.1) 2. to describe the behavior of fluorinated amino acids in systems emulating protein environments (section 3.2, section 3.3) First, to characterize fluorinated amino acids side chains we have used fluorinated ethane derivatives as their simplified models and applied a quantum mechanics approach. Properties such as charge distribution, dipole moments, volumes and size of the fluoromethylated groups within the model have been characterized. Hydrogen bonding properties of these groups have been compared with the groups typically presented in natural protein environments. We have shown that hydrogen and fluorine atoms within these fluoromethylated groups are weak hydrogen bond donors and acceptors. Nevertheless they should not be disregarded for applications in protein engineering. Then, we have implemented four fluorinated L-amino acids for the AMBER force field and characterized their conformational and hydration properties at the MD level. We have found that hydrophobicity of fluorinated side chains grows with the number of fluorine atoms and could be explained in terms of high electronegativity of fluorine atoms and spacial demand of fluorinated side-chains. These data on hydration agrees with the results obtained in the experimental work performed by our collaborators. We have rationally engineered systems that allow us to study fluorine properties and extract results that could be extrapolated to proteins. For this, we have emulated protein environments by introducing fluorinated amino acids into a parallel coiled-coil and enzyme-ligand chymotrypsin systems. The results on fluorination effect on coiled-coil dimerization and substrate affinities in the chymotrypsin active site obtained by MD, molecular docking and free energy calculations are in strong agreement with experimental data obtained by our collaborators. In particular, we have shown that fluorine content and position of fluorination can considerably change the polarity and steric properties of an amino acid side chain and, thus, can influence the properties that a fluorinated amino acid reveals within a native protein environment. III. Coiled-coils typically consist of two to five right-handed α-helices that wrap around each other to form a left-handed superhelix. The interface of two α-helices is usually represented by hydrophobic residues. However, the analysis of protein databases revealed that in natural occurring proteins up to 20% of these positions are populated by polar and charged residues. The impact of these residues on stability of coiled-coil system is not clear. MD simulations together with free energy calculations have been utilized to estimate favourable interaction partners for uncommon amino acids within the hydrophobic core of coiled-coils (Chapter 4). Based on these data, the best hits among binding partners for one strand of a coiled-coil bearing a charged amino acid in a central hydrophobic core position have been selected. Computational data have been in agreement with the results obtained by our collaborators, who applied phage display technology and CD spectroscopy. This combination of theoretical and experimental approaches allowed to get a deeper insight into the stability of the coiled-coil system. To conclude, this thesis widens existing concepts of protein structural biology in three areas of its current importance. We expand on the role of solvent in protein interfaces, which contributes to the knowledge of physico-chemical properties underlying protein-protein interactions. We develop a deeper insight into the understanding of the fluorine's impact upon its introduction into protein environments, which may assist in exploiting the full potential of fluorine's unique properties for applications in the field of protein engineering and drug design. Finally we investigate the mechanisms underlying coiled-coil system folding. The results presented in the thesis are of definite importance for possible applications (e.g. introduction of solvent explicitly into the scoring function) into protein folding, docking and rational design methods. The dissertation consists of four chapters: ● Chapter 1 contains an introduction to the topic of protein-protein interactions including basic concepts and an overview of the present state of research in the field. ● Chapter 2 focuses on the studies of the role of solvent in protein interfaces. ● Chapter 3 is devoted to the work on fluorinated amino acids in protein environments. ● Chapter 4 describes the study of coiled-coils folding properties. The experimental parts presented in Chapters 3 and 4 of this thesis have been performed by our collaborators at FU Berlin. Sections 2.1, 2.2, 3.1, 3.2 and Chapter 4 have been submitted/published in peer-reviewed international journals. Their organization follows a standard research article structure: Abstract, Introduction, Methodology, Results and discussion, and Conclusions. Section 3.3, though not published yet, is also organized in the same way. The literature references are summed up together at the end of the thesis to avoid redundancy within different chapters.

Thèse numérisée par la Division de la gestion de documents et des archives de l'Université de Montréal

Les matériaux moléculaires amorphes, aussi appelés verres moléculaires, sont constitués de molécules organiques de petite taille capables de s’organiser de façon désordonnée. En plus de présenter certaines des propriétés analogues à celles des polymères, ils offrent des avantages supplémentaires, puisqu’ils sont des espèces isomoléculaires dont la synthèse, la purification et la mise en œuvre sont facilitées par leur viscosité relativement faible. Toutefois, la préparation souvent exigeante de ces matériaux et leur durée de vie utile limitée par leur tendance à relaxer vers l’état cristallin demeurent des obstacles à leur utilisation pour certaines applications, e.g. opto-électronique, nanolithographie, pharmaceutique. Le développement de stratégies visant à faciliter la préparation de la phase vitreuse et éviter sa cristallisation est donc essentiel à la conception de matériaux moléculaires amorphes fonctionnels. L’objectif principal de cette thèse est d’établir des relations entre la structure moléculaire des verres moléculaires et leurs propriétés. Pour y arriver, différentes librairies de composés modèles, des dérivés analogues de triazine ayant démontré une excellente capacité à former une phase vitreuse, sont utilisées pour i) déterminer l’influence de la nature et de la position des groupements sur la triazine; ii) explorer l’influence des liaisons hydrogène sur les propriétés des verres lorsque leur structure comporte des groupements fonctionnels reconnus pour faciliter la cristallisation et lorsque leurs conditions de préparation se rapprochent de celles employées en industrie et iii) exploiter la phase amorphe afin d’étudier la photosensibilité des azobenzènes (azo) en vue d’optimiser leur utilisation dans des applications. Tout d’abord, l’influence des différents groupes substituants sur la triazine (groupements de tête, auxiliaires et liants) sur la capacité des composés à former une phase vitreuse (GFA), sur sa stabilité cinétique (GS) et sur sa température de transition vitreuse (Tg) est étudiée. Un système de classification des composés développé à partir de mesures de calorimétrie différentielle à balayage (DSC) et des mesures de spectroscopie infrarouge (IR) à température variable combinées à des analyses chimiométriques facilitent la rationalisation des rôles joués par chaque groupe. L’impact des liaisons hydrogène (H), de la barrière énergétique de rotation et de l’encombrement stérique des groupements est ainsi déterminé, permettant de conclure que le groupe de tête est le plus influent et que la présence de liaisons H n’est pas essentielle au GFA mais qu’elle est importante pour obtenir une Tg élevée. Ensuite, l’influence des liaisons H sur les propriétés des verres se rapprochant de ceux exploités dans l’industrie est explorée. Des mesures de spectroscopie IR à température variable, de DSC et de résolution de structures cristallines ont permis de conclure que les liaisons H réussissent à nuire à la cristallisation des composés et ce, même s’ils sont simultanément fonctionnalisés avec des motifs qui favorisent la cristallisation (empilements π-π entre dérivés stilbènes fluorés et non fluorés). De plus, trois composés analogues fonctionnalisés avec un groupement de tête possédant une capacité décroissante à établir des liaisons H (donneur, accepteur, aucune) ont été déposés en phase vapeur (PVD), une technique employée entre autres dans l’industrie opto-électronique pour évaluer leur capacité à former des verres ultrastables. Les films ainsi préparés présentent tous des propriétés similaires à celles des verres ultrastables précédemment étudiés, telles qu’une plus grande densité et anisotropie, et sont tous plus stables que ceux préparés par refroidissement à partir de l’état liquide. Toutefois, le verre formé du composé avec un groupement de tête donneur de liaisons H est moins stable que les autres d’au moins un ordre de grandeur, suggérant que les liaisons H limitent le niveau de stabilité atteignable par PVD. Finalement, un verre à base de triazine fonctionnalisé avec un groupement azo est employé pour étudier d’un point de vue moléculaire les perturbations provoquées par la photoisomérisation de l’azo. Grâce à une nouvelle méthode de spectroscopie IR, il est possible d’observer un gradient d’environnement moléculaire le long de la molécule lors de la photoisomérisation, permettant de soutenir certaines hypothèses relatives au déplacement macroscopique de la matière qui en résulte. Les mélanges de verres à base de triazine servent aussi de plateforme idéale pour découpler l’influence de la Tg et du contenu en azo sur la photo-orientation de l’azo, mais aussi sur la cinétique d’écriture et l’efficacité des réseaux de diffraction (SRG). Ce travail permet ainsi de déterminer une zone optimale de Tg pour l’inscription de SRG. Ces nouvelles connaissances mèneront à la conception plus rationnelle de nouveaux verres moléculaires, pouvant s’étendre à d’autres matériaux amorphes. / Amorphous molecular materials, also known as molecular glasses, are small organic molecules capable of being organized in a disordered manner. In addition to sharing some of the useful properties of polymers, they offer additional advantages because they are isomolecular species for which synthesis, purification and processing are facilitated by a relatively low viscosity. However, the usually demanding preparation conditions of these materials and their limited functional lifetime due to their tendency to relax to the crystalline state remain obstacles to their use for certain applications, e.g. opto-electronics, nanolithography, pharmaceuticals. The development of strategies to facilitate the preparation of the vitreous phase and avoid its crystallization is therefore essential for the design of functional amorphous molecular materials. The main objective of this thesis is to establish relationships between the molecular structure of molecular glasses and their properties. To achieve it, various libraries of model compounds, analogues of triazine derivatives that have demonstrated excellent glass-forming ability, are used to i) determine the influence of the nature and the position of the groups on the triazine; ii) explore the influence of hydrogen (H) bonds on the properties of glasses when their structure includes functional groups known to facilitate crystallization and when their preparation conditions are similar to those used in industry; and iii) exploit the amorphous phase in order to study the photoresponsiveness of azobenzenes (azo) in order to optimize their use in different applications. The influence of the various substituent groups on the triazine (headgroup, ancillary and linkers) on the glass-forming ability (GFA), the kinetic glass stability (GS) and the glass transition temperature (Tg) of the compounds is first studied. A classification system based on differential scanning calorimetry (DSC) and variable temperature infrared spectroscopy (IR) measurements combined to chemometrics analyses facilitate the rationalization of the roles played by each group. The impact of the H-bonds, the energy of the rotation barrier, and the steric hindrance of the groups is determined, leading to the conclusion that the headgroup is the most influential group and that the presence of H-bonds is not essential to the GFA, but important to obtain a high Tg. The influence of the H-bonds on the properties of glasses approaching those exploited in industry is then explored. Variable temperature IR spectroscopy measurements, DSC studies, and single crystal structure resolution have led to the conclusion that H-bonds impede the crystallization of the compounds even though they are simultaneously functionalized with moieties that promote crystallization (π-π stacking between fluorinated and non-fluorinated stilbene groups). In addition, three similar compounds functionalized with a headgroup presenting a decreasing capability to establish H-bonds (donor, acceptor, none) were vapor-deposited (PVD), a technique used, among others, in the opto-electronic industry, to evaluate their capability to form ultrastable glasses. These PVD glasses all show properties that are similar to those previously reported for ultrastable glasses, including higher density and anisotropy, and are all more kinetically stable than glasses prepared by cooling from the viscous state. However, the PVD glasses prepared with a H-bond donor headgroup are less stable than the others by at least an order of magnitude, suggesting that H-bonds limit the level of kinetic stability achievable by PVD. Finally, a triazine molecular glass functionalized with an azo group is used to study, from a molecular point of view, the perturbations caused by the photoisomerization of the azo. A new IR spectroscopy method was developed to observe a molecular environment gradient along the molecule during photoisomerization, making it possible to support certain hypotheses concerning the resulting macroscopic transport of the material. Triazine-based molecular glass blends are also used as an ideal platform for decoupling the influence of Tg and azo content on the azo photo-orientation, but also on the inscription kinetics and the diffraction efficiency of surface relief gratings (SRGs). This work enables the determination of an optimal Tg range for the inscription of SRGs. Altogether, these new insights will lead to a more rational design of new molecular glasses, which can extend to other amorphous molecular materials.

Proteins have an indispensable role in the cell. They carry out a wide variety of structural, catalytic and signaling functions in all known biological systems. To perform their biological functions, proteins establish interactions with other bioorganic molecules including other proteins. Therefore, protein-protein interactions is one of the central topics in molecular biology. My thesis is devoted to three different topics in the field of protein-protein interactions. The first one focuses on solvent contribution to protein interfaces as it is an important component of protein complexes. The second topic discloses the structural and functional potential of fluorine's unique properties, which are attractive for protein design and engineering not feasible within the scope of canonical amino acids. The last part of this thesis is a study of the impact of charged amino acid residues within the hydrophobic interface of a coiled-coil system, which is one of the well-established model systems for protein-protein interactions studies. I. The majority of proteins interact in vivo in solution, thus studies of solvent impact on protein-protein interactions could be crucial for understanding many processes in the cell. However, though solvent is known to be very important for protein-protein interactions in terms of structure, dynamics and energetics, its effects are often disregarded in computational studies because a detailed solvent description requires complex and computationally demanding approaches. As a consequence, many protein residues, which establish water-mediated interactions, are neither considered in an interface definition. In the previous work carried out in our group the protein interfaces database (SCOWLP) has been developed. This database takes into account interfacial solvent and based on this classifies all interfacial protein residues of the PDB into three classes based on their interacting properties: dry (direct interaction), dual (direct and water-mediated interactions), and wet spots (residues interacting only through one water molecule). To define an interaction SCOWLP considers a donor–acceptor distance for hydrogen bonds of 3.2 Å, for salt bridges of 4 Å, and for van der Waals contacts the sum of the van der Waals radii of the interacting atoms. In previous studies of the group, statistical analysis of a non-redundant protein structure dataset showed that 40.1% of the interfacial residues participate in water-mediated interactions, and that 14.5% of the total residues in interfaces are wet spots. Moreover, wet spots have been shown to display similar characteristics to residues contacting water molecules in cores or cavities of proteins. The goals of this part of the thesis were: 1. to characterize the impact of solvent in protein-protein interactions 2. to elucidate possible effects of solvent inclusion into the correlated mutations approach for protein contacts prediction To study solvent impact on protein interfaces a molecular dynamics (MD) approach has been used. This part of the work is elaborated in section 2.1 of this thesis. We have characterized properties of water-mediated protein interactions at residue and solvent level. For this purpose, an MD analysis of 17 representative complexes from SH3 and immunoglobulin protein families has been performed. We have shown that the interfacial residues interacting through a single water molecule (wet spots) are energetically and dynamically very similar to other interfacial residues. At the same time, water molecules mediating protein interactions have been found to be significantly less mobile than surface solvent in terms of residence time. Calculated free energies indicate that these water molecules should significantly affect formation and stability of a protein-protein complex. The results obtained in this part of the work also suggest that water molecules in protein interfaces contribute to the conservation of protein interactions by allowing more sequence variability in the interacting partners, which has important implications for the use of the correlated mutations concept in protein interactions studies. This concept is based on the assumption that interacting protein residues co-evolve, so that a mutation in one of the interacting counterparts is compensated by a mutation in the other. The study presented in section 2.2 has been carried out to prove that an explicit introduction of solvent into the correlated mutations concept indeed yields qualitative improvement of existing approaches. For this, we have used the data on interfacial solvent obtained from the SCOWLP database (the whole PDB) to construct a “wet” similarity matrix. This matrix has been used for prediction of protein contacts together with a well-established “dry” matrix. We have analyzed two datasets containing 50 domains and 10 domain pairs, and have compared the results obtained by using several combinations of both “dry” and “wet” matrices. We have found that for predictions for both intra- and interdomain contacts the introduction of a combination of a “dry” and a “wet” similarity matrix improves the predictions in comparison to the “dry” one alone. Our analysis opens up the idea that the consideration of water may have an impact on the improvement of the contact predictions obtained by correlated mutations approaches. There are two principally novel aspects in this study in the context of the used correlated mutations methodology : i) the first introduction of solvent explicitly into the correlated mutations approach; ii) the use of the definition of protein-protein interfaces, which is essentially different from many other works in the field because of taking into account physico-chemical properties of amino acids and not being exclusively based on distance cut-offs. II. The second part of the thesis is focused on properties of fluorinated amino acids in protein environments. In general, non-canonical amino acids with newly designed side-chain functionalities are powerful tools that can be used to improve structural, catalytic, kinetic and thermodynamic properties of peptides and proteins, which otherwise are not feasible within the use of canonical amino acids. In this context fluorinated amino acids have increasingly gained in importance in protein chemistry because of fluorine's unique properties: high electronegativity and a small atomic size. Despite the wide use of fluorine in drug design, properties of fluorine in protein environments have not been yet extensively studied. The aims of this part of the dissertation were: 1. to analyze the basic properties of fluorinated amino acids such as electrostatic and geometric characteristics, hydrogen bonding abilities, hydration properties and conformational preferences (section 3.1) 2. to describe the behavior of fluorinated amino acids in systems emulating protein environments (section 3.2, section 3.3) First, to characterize fluorinated amino acids side chains we have used fluorinated ethane derivatives as their simplified models and applied a quantum mechanics approach. Properties such as charge distribution, dipole moments, volumes and size of the fluoromethylated groups within the model have been characterized. Hydrogen bonding properties of these groups have been compared with the groups typically presented in natural protein environments. We have shown that hydrogen and fluorine atoms within these fluoromethylated groups are weak hydrogen bond donors and acceptors. Nevertheless they should not be disregarded for applications in protein engineering. Then, we have implemented four fluorinated L-amino acids for the AMBER force field and characterized their conformational and hydration properties at the MD level. We have found that hydrophobicity of fluorinated side chains grows with the number of fluorine atoms and could be explained in terms of high electronegativity of fluorine atoms and spacial demand of fluorinated side-chains. These data on hydration agrees with the results obtained in the experimental work performed by our collaborators. We have rationally engineered systems that allow us to study fluorine properties and extract results that could be extrapolated to proteins. For this, we have emulated protein environments by introducing fluorinated amino acids into a parallel coiled-coil and enzyme-ligand chymotrypsin systems. The results on fluorination effect on coiled-coil dimerization and substrate affinities in the chymotrypsin active site obtained by MD, molecular docking and free energy calculations are in strong agreement with experimental data obtained by our collaborators. In particular, we have shown that fluorine content and position of fluorination can considerably change the polarity and steric properties of an amino acid side chain and, thus, can influence the properties that a fluorinated amino acid reveals within a native protein environment. III. Coiled-coils typically consist of two to five right-handed α-helices that wrap around each other to form a left-handed superhelix. The interface of two α-helices is usually represented by hydrophobic residues. However, the analysis of protein databases revealed that in natural occurring proteins up to 20% of these positions are populated by polar and charged residues. The impact of these residues on stability of coiled-coil system is not clear. MD simulations together with free energy calculations have been utilized to estimate favourable interaction partners for uncommon amino acids within the hydrophobic core of coiled-coils (Chapter 4). Based on these data, the best hits among binding partners for one strand of a coiled-coil bearing a charged amino acid in a central hydrophobic core position have been selected. Computational data have been in agreement with the results obtained by our collaborators, who applied phage display technology and CD spectroscopy. This combination of theoretical and experimental approaches allowed to get a deeper insight into the stability of the coiled-coil system. To conclude, this thesis widens existing concepts of protein structural biology in three areas of its current importance. We expand on the role of solvent in protein interfaces, which contributes to the knowledge of physico-chemical properties underlying protein-protein interactions. We develop a deeper insight into the understanding of the fluorine's impact upon its introduction into protein environments, which may assist in exploiting the full potential of fluorine's unique properties for applications in the field of protein engineering and drug design. Finally we investigate the mechanisms underlying coiled-coil system folding. The results presented in the thesis are of definite importance for possible applications (e.g. introduction of solvent explicitly into the scoring function) into protein folding, docking and rational design methods. The dissertation consists of four chapters: ● Chapter 1 contains an introduction to the topic of protein-protein interactions including basic concepts and an overview of the present state of research in the field. ● Chapter 2 focuses on the studies of the role of solvent in protein interfaces. ● Chapter 3 is devoted to the work on fluorinated amino acids in protein environments. ● Chapter 4 describes the study of coiled-coils folding properties. The experimental parts presented in Chapters 3 and 4 of this thesis have been performed by our collaborators at FU Berlin. Sections 2.1, 2.2, 3.1, 3.2 and Chapter 4 have been submitted/published in peer-reviewed international journals. Their organization follows a standard research article structure: Abstract, Introduction, Methodology, Results and discussion, and Conclusions. Section 3.3, though not published yet, is also organized in the same way. The literature references are summed up together at the end of the thesis to avoid redundancy within different chapters.

Thèse numérisée par la Division de la gestion de documents et des archives de l'Université de Montréal.

Die Energierelaxationsdynamik und strukturelle Dynamik der DNA spielen eine entscheidende Rolle für das Verständnis der photochemischen Eigenschaften und biologischen Funktion von DNA. Die schnellsten Prozesse geschehen dabei auf Zeitskalen im Femto- bis Pikosekundenbereich und können durch zeitaufgelöste (ultraschnelle) Messungen an molekularen Schwingungsanregungen in Echtzeit beobachtet werden. In dieser Arbeit werden NH-Streck Schwingungsmoden der Nukleobasen sowie OH-Streckmoden der Hydrathülle mittels linearer infrarot (IR) Spektroskopie sowie ultraschneller Pump-Probe und zweidimensionaler IR Spektroskopie untersucht. Messungen an monomerartigen Nukleobasen, wasserstoffverbrückten Nukleobasenpaaren und kurzen DNA Fragmenten in Doppelhelixstruktur ermöglichen es, die Effekte der verschiedenen Wasserstoffbrücken und Schwingungskopplungen separat voneinander zu untersuchen. Im Zusammenspiel mit exzitonischen und quantenchemischen Rechnungen werden so weitreichende Einsichten in die spektroskopischen Eigenschaften und die Relaxationsdynamik von Schwingungsanregungen in DNA gewonnen. Es wird u.a. gezeigt, dass Wasserstoffbrücken zwischen Nukleobasen eine Lebensdauer größer 1 ps besitzen und eine beschleunigte Energierelaxation durch Fermiresonanzen mit niederfrequenten Schwingungsmoden des Fingerprintbereichs bewirken. Die DNA-Hydrathülle zeigt eine ultraschnelle strukturelle Dynamik unabhängig von der Basenpaarzusammensetzung und fungiert als effiziente Wärmesenke für hochfrequente Schwingungsanregungen. / Energy relaxation and structural dynamics in DNA play a crucial role for the understanding of the photochemical properties and biological function of DNA. The fastest of such processes occur on the femto- to picosecond time scale and can be followed in real time through time-resolved (ultrafast) measurements on molecular vibrations. In this work, NH stretching excitations of nucleobases and OH stretching modes of the hydration shell are analyzed through linear infrared (IR) spectroscopy as well as ultrafast pump-probe and two-dimensional IR spectroscopy. Measurements on monomeric nucleobases, hydrogen-bonded nucleobase pairs, and short DNA fragments in double helix structure allow one to examine the effects of the different hydrogen bonds and vibrational couplings separately from each other. The combination with excitonic and quantum chemical calculations provides profound new insights into the spectroscopic properties and relaxation dynamics of vibrational excitations in DNA. This work shows that hydrogen bonds between nucleobases have a lifetime greater than 1 ps and lead to an accelerated dissipation of energy due to Fermi resonances with vibrational modes in the fingerprint range. The hydration shell of DNA exhibits ultrafast structural dynamics independent of the base pair composition and serves as an efficient heat sink for high-energy vibrational excitations.

Für die makroskopischen Eigenschaften und Funktionen biologisch relevanter Materie spielen schwache, intra- und intermolekulare Wechselwirkungen dispersiver und elektrostatischer Natur auf molekularem Niveau eine große Rolle. Um diese schwachen Wechselwirkungen zu untersuchen, können Modellsysteme, isoliert in der Gasphase, herangezogen werden. Benzoldimer, ein schwach gebundener Van der Waals Komplex, kann beispielsweise als Modellsystem für dispersive Wechselwirkungen dienen. In der vorliegenden Arbeit werden die strukturellen Eigenschaften und die (interne) Dynamik des Benzoldimers mit Hilfe spektroskopischer Methoden in den Energiebereichen der Rotationen, Vibrationen und elektronischen Übergänge untersucht und im Kontext der Symmetrie diskutiert. Die in dieser Arbeit vorgestellten Experimente tragen zu einem tieferen Verständnis des Benzoldimers bei, jedoch zeigt das Experiment zur internen Dynamik auch, dass eine ausreichende theoretische Beschreibung des Benzoldimers nach wie vor eine Herausforderung darstellt. Schwingungsübergänge hochsymmetrischer Moleküle sind oft optisch inaktiv, können jedoch mit der hier vorgestellten Methode der Symmetrieerniedrigung durch Komplexierung zugänglich gemacht werden, wie am Beispiel des Benzols demonstriert wird. Außerdem wird ein Mechanismus vorgstellt, der kollisionsinduzierte Konformationsänderungen in einem Molekularstrahl beschreibt. Dieses Modell kann generell für Molekularstrahlexperimente an flexiblen Molekülen hilfreich sein, einerseits um die beobachtete Konformationsverteilung zu verstehen, andererseits um die experimentellen Parameter gezielt zu verändern und somit Konformerpopulationen zu manipulieren. Die in dieser Dissertation vorgestellten spektroskopischen Experimente liefern einerseits molekülspezifische Informationen und ermöglichen andererseits, Modelle, die von allgemeiner Bedeutung sind, zu entwickeln. / The macroscopically observable properties and functionalities of biological matter are often determined by weak intra- and intermolecular interactions on the microscopic level. Such weak interactions are for example hydrogen bonding and van der Waals interactions and can be investigated best on isolated model systems in the gas phase. The benzene dimer, for example, is a prototype system to investgate dispersive interactions. The spectroscopic experiments, covering the energy ranges of rotations, vibrations and electronic transitions, presented in this thesis, contribute to a deeper understanding of the benzene dimer. However, from the experiments investigating the internal dynamics it becomes clear that an appropriate theoretical description of the benzene dimer is still a challenge. Vibrational transitions of highly symmetric molecules, as for example of the benzene, are often optically inactive. Here, a method is presented, which exploits symmetry reduction upon complexation and thus allows one to access such modes. Furthermore, a model is proposed describing collision induced conformational interconversion in a molecular beam. This model can be helpful for molecular beam experiments of flexible molecules to understand the observed relative conformational population and to adapt the experimental conditions allowing for the manipulation of the relative conformer abundances. In this thesis, results are presented that allow one on the one hand to deduce molecular specific information and that on the other hand also give a broader insight into phenomena of general importance.