Le monoxyde d’azote (NO), dont le rôle biologique a été découvert à la fin du 20ème siècle, est impliqué dans la régulation de nombreux processus à l’échelle de la cellule et de l’organisme. Sa biosynthèse est réalisée par les enzymes NO synthases (NOS), et met en jeu la liaison de NADPH à leur domaine réductase suivie d’une série de transfert d’électrons vers leur domaine oxygénase, où la formation de NO se produit par oxydation de la L-arginine. En s’inspirant de mimes photo-activables de NADPH précédemment décrits dans la littérature, appelés nano-déclencheurs (NT, de l’anglais nanotriggers), induisant la production de NO par illumination, nous avons conçu et synthétisé de nouvelles générations de composés potentiellement capables d’initier l’activité catalytique de NOS sous irradiation. Ils comportent une unité de reconnaissance de NOS dérivée de l’adénosine et une unité chromophorique de type diaminophényl butadiène, liées entre elles par un groupement triazole. Ces structures modulables, facilement assemblées par chimie « click » ont permis la préparation d’une librairie de nano-déclencheurs, dont les propriétés photophysiques et la stabilité dans des conditions physiologiques ont été évaluées. Ces nouvelles générations de composés offrent des perspectives intéressantes pour le contrôle de processus biologiques par la lumière. / Nitric oxide (NO), whose biological role has been discovered in the late 20th century, is involved in the regulation of many processes in cell and organism. Its biosynthesis is carried out by enzymes named nitric oxide synthases (NOS) and involves NADPH binding to their reductase domain followed by a series of electron transfers to their oxygenase domain, where the formation of NO takes place by oxidation of L-arginine. Inspired by photoactivatable NADPH mimics called nano-triggers (NT), previously described in the literature, able to produce NO upon illumination, we designed and synthesized new generations of compounds potentially capable of initiating the catalytic activity of NOS under irradiation. They contain a recognition unit for NOS derived from adenosine and a diaminophenyl butadiene chromophoric moiety, linked together by a triazole group. These modular structures, easily assembled by "click" chemistry allowed the preparation of a library of nano-triggers, whose photophysical properties and stability under physiological conditions were evaluated. These new generations of compounds offer interesting perspectives for the control of biological processes by light.

Le visage humain joue un rôle important dans la communication interpersonnelle. La dysfonction du visage ou le défigurement due aux traumatismes ou pathologies peuvent entraver les activités sociales normales. Le traitement chirurgical est souvent nécessaire. De nos jours, le résultat du traitement chirurgical et l’état d’établissement ne sont estimé qu’avec les méthodes qualitatives telles que l’observation visuelle et la palpation. Dans l’attente de fournir des critères quantitatifs, cette thèse a pour l’objectif de modéliser la mimique faciale utilisant MEF (Méthode d’Éléments Finis) sur la base des données d’IRM (Imagerie par Résonance Magnétique). Un modèle sujet-spécifique du visage a été construit sur la base de la segmentation des données IRM ; il contient des parties osseuses, muscles de la mimique (p.ex. le muscle grand zygomatique), les tissues mous sous-cutanées et la peau. L’identification des tissus mous biologiques a été réalisée via des essais de traction bi-axiale et la modélisation numérique. Ensuite, le modèle géométrique a été maillé pour effectuer des calculs EF simulant trois mouvements mimiques du visage (sourire, prononciation du son « Pou » et « O »). Les muscles ont été modélisés comme un matériau quasi-incompressible, transversalement isotrope et hyperélastique, avec la capacité d’activation. Des informations pertinentes (p.ex. l’amplitude de contraction du muscle) utilisées dans la simulation ont été extraites de la mesure des données d’IRM. Il est à noter que les mêmes données expérimentales d’IRM telles qu’ils ont utilisées dans la modélisation ont été prises comme une référence de validation pour les résultats de simulation. Cette étude peut être appliquée cliniquement dans l’évaluation du traitement faciale et le rétablissement postopérative. / Human face plays an important role interpersonal communication. Facial dysfunction or disfigurement due to trauma or pathologies may impede normal social activities. Surgical treatment is often necessary. Nowadays, treatment outcome and rehabilitation condition are estimated only by qualitative methods, such as visual observation and palpation. In expectation of providing quantitative criteria, this thesis proposes to model facial mimics using FEM (Finite Element Method) on the basis of MRI (Magnetic Resonance Imaging) data. A subject-specific face model was reconstructed based on segmentation of MRI data; it contains bony parts, mimic muscles (e.g. zygomaticus major muscle), subcutaneous soft tissues and skin. Identification of biological soft tissues was conducted through bi-axial tension tests and numerical modeling. Then the geometric model was meshed to conduct FE calculations simulating three facial mimic movements (smile, pronunciation of sound “Pou” and “O”). Muscle was modeled as quasi-incompressible, transversely-isotropic, hyperelastic material, with activation ability. Relevant information (e.g. contraction amplitude of muscle) used in simulation was extracted from measurement of MRI data. It is to be noted that the same experimental MRI data as used in modeling was taken as validation reference for simulation results. This study can be applied clinically in evaluation of facial treatment andpostoperative recovery.

Thyroid hormones (THs; T4 and T3), secreted from thyroid gland, play an important role in human growth and development. T3 (3,5,3′-triiodothyronine) is the active hormone and the conversion of T4 (3,3′,5,5′-tetraiodothyronine) to T3 in cells is mediated by iodothyronine deiodinases enzymes (DIOs). DIOs are selenocysteine containing enzymes and are classified into three types (DIO1, DIO2 and DIO3). DIO1 catalyzes the outer-ring deiodination (ORD; T3 formation) and inner-ring deiodination (IRD; rT3 formation) reactions, involving in the activation (T4 to T3 conversion) and inactivation (T4 to rT3 conversion), respectively. DIO2 and DIO3 catalyse the ORD and IRD reactions, respectively. This homeostasis is regulated tightly and any deviation would lead to diseases like hyperthyroidism or hypothyroidism. Recently it is of interest to many research groups to develop iodothyronine deiodinase mimics and we have developed naphthalene-based peri-substituted thioselenol pair at 1,8-positions (1.25), which remove iodine selectively from inner-ring of T4. When selenium atom is substituted in place of sulfur (selenol-selenol pair; 1.26), the deiodination activity was ca. 90 times faster than with 1.25. This thesis deals with various aspects of the effect of substituents on the naphthalene-1,8-diselenol and solvent effect on the thyroid hormone deiodination by naphthalene-based iodothyronine deiodinase mimics. Figure 1. (A) Deiodination reactions by DIOs. (B) Chemical structure of 1.25 and 1.26. The thesis consists of five chapters. The first chapter provides a general overview about sialoproteins, thyroid hormone biosynthesis, thyroid hormone metabolism, halogen bonding, iodothyronine deiodinase mimics and proposed mechanisms for the deidoination of thyroid hormones. This chapter also introduces peri-naphthalene-1,8-diselenol (1.26), which is the key compound in this thesis and discusses about proposed mechanism for the deiodination of thyroxine involving co-operative halogen bonding and chalcogen bonding mechanism. Figure 2. (A) TH action. (B) Proposed mechanism for the deiodination of T4 by 1.26 involving cooperative halogen bonding and chalcogen bonding. Chapter 2 discusses about the synthesis, characterization and deiodination activity of a series of naphthalene-based peri-substituted-1,8-diselenols (Figure 3). These diselenols regioselectivity remove iodine from inner ring of thyroxine and other thyroid hormones, (T3 and 3, 5-T2). Substitution with different groups on the naphthalene ring did not change the regioselectivity of deiodination, indicating that the deiodination activity does not depend on the nature of substituents. Secondary or tertiary amine side chain group attached at the 2nd position of the naphthalene ring showed better activity. It is due to the secondary interaction, which facilitates the iodine removal. It was further confirmed with the substitutions at the 4th position of the ring to discriminate the possibility of electronic effect. The higher deiodination rate owing to the t-butyl group at second position of the ring also suggests that the steric effect may also play a role in the deiodination reaction (Figure 4). It is proposed that peri substituted naphthalene-1,8-diselenols remove iodine from thyroid hormones through halogen bonding-chalcogen bonding mechanism (Figure 2). The investigation of Se···Se bond distance from the crystal structures and through DFT calculation and NMR experiment showed that the stronger chalcogen bond could be the reason for the increase in the reactivity observed with substituted peri-naphthalene-1,8-diselenols. Figure 3. peri-substituted naphthalene-1,8-diselenols used for the study. Figure 4. Relative deiodinase activity of substituted-peri-naphthalene-1,8-diselenols with T4. In Chapter 3, we have discussed about the effect of chalcogen atom substitution in a series of deiodinase mimics on the deiodination of thyroid hormones. Moving from thiol-selenol pair (1.25) to selenol-selenol pair (1.26) in naphthalene based peri-substituted mimics, an increase in the activity was observed. In this chapter, we have shown that substituting with tellurium, as tellurium-thiol pair (3.3) and ditellurol (3.4) increases the reactivity of deiodination to several times and also regioselectivity of deiodination is changed from IRD in the case of 1.26 to both IRD and ORD for 3.3 and 3.4. The presence of two tellurol moieties (3.4) or a thiol-tellurol pair (3.3) can mediate sequential deiodination of T4, to produce all the possible thyroid hormone derivatives under physiologically relevant conditions (Figure 5). This study provided the first experimental evidence that the regioselectivity of the thyroid hormone deiodination is controlled by the nucleophilicity and the strength of halogen bond between the iodine and chalcogen atoms. Figure 5. (A) HPLC chromatograms of deiodination reaction of T4 with 3.3 and 3.4. (B) Chemical structure of 3.3 and 3.4. (C) Sequential deiodination reaction of T4 by 3.3 and 3.4. Chapter 4 describes the effect of alkyl conjugation at 4′-OH position of THs on the deiodination by iodothyronine mimics. In addition to the deiodination, iodothyronines undergo conjugation with sulfate and glucuronic acid group at 4′-hydroxyl position. Conjugation alters the physico-chemical properties of iodothyronines. For example, it is known that sulfate conjugation increases the rate of deiodination to a large extend. We have conjugated alkyl group at 4′-hydroxyl position of iodothyronines and investigated the deiodination reactions with reported peri-substituted naphthalene-1,8-diselenols. We observed that similar to sulfated thyroid hormones O-methylthyroxine also undergoes both phenolic and tyrosyl ring deiodination reactions and overall the rate of deiodination is increased at least by 5 times as compared with T4 under identical conditions. The phenolic iodine removal is favored by conjugation as compared to the tyrosyl ring iodine, which is similar to the observation made for T4S. Interestingly, when the acetamide group is conjugated at 4′-OH position, the regioselectivity of deiodination is changed exclusively to 5′-iodine. DFT calculations show that the positive potential on the iodine increase upon conjugation, which leads to stronger halogen bonding interaction with selenol, might be the reason for the change in the regioselectivity of deiodination. Figure 6. (A) HPLC chromatogram of deiodination reaction of T4(Me) with 1.26. (B) Initial rate comparison of T4 and T4(Me).(C) HPLC chromatogram of deiodination reaction of T4(AA) with 1.26 showing the formation of T3(AA) (ORD product). (D) Electron potential map of T4, T4(Me) and T4(AA) showing the increase in electro positive potential on 5′-iodine upon conjugation. Chapter 5 deals with the solvent effect on the deiodination reactions of THs by iodothyronine deiodinase mimics. As discussed in the earlier chapters, the deiodination reaction of thyroxine by naphthalene based-1,8-diselenols under physiological conditions produce, rT3 (IRD) as the only observable products. Surprisingly, when the deiodination reaction was performed in DMF or DMSO in the presence of 1.26, the regioselectivity of reaction was changed and the formation of both T3 (ORD) and rT3 was observed. In DMF or in DMSO, the deiodination reactivity of 1.26 was found to be 1000 fold higher than the reaction performed in phosphate buffer at pH 7.4. Figure 7. (A) HPLC chromatogram for the deiodination reaction of T4 in DMF by 1.26 showing both IRD and ORD. (B) A comparison of initial rate for the deiodination reactions of T4, T3 and 3,5-T2 in DMF and in DMSO by 1.26. (C) HPLC chromatograms for the deiodination reaction of T4 in DMF by 1.26 in the presence of TEMPO, showing the inhibition of deiodination (i) 0 mM TEMPO (ii) 10 mM of TEMPO (iii) 30 mM TEMPO. (D) HPLC chromatograms for the deiodination reaction of T4 in DMSO by 1.26 in the presence of TEMPO showing the inhibition of deiodination (i) 0 mM TEMPO (ii) 10 mM of TEMPO (iii) 30 mM TEMPO. 3,5-DIT was not denominated under physiological conditions, however, in DMF and in DMSO, 3,5-DIT was deiodinated by 2.4 to produce 3-MIT. We also observed that the control reactions in DMF or DMSO also showed a little deiodination activity. The very high reactivity observed in the presence of DMF or DMSO implied that the mechanism of denomination in these solvents may be different. It has been reported that DMSO or DMF radicals can be formed with small amounts of a base. Reaction mixture consisting of NaBH4 (for generating selenol from diselenide) and NaOH (T4 solution) may facilitate the radical formation. We also performed the reaction in the presence of TEMPO (free radical scavenger) and observed the inhibition of deiodination reaction. However, it is not clear whether the radical pathway could be one of the possible mechanisms of deiodination in these solvents by compounds 1.26 and 2.4. Further studies are required to propose a radical mechanism in different solvents such as DMF and DMSO.

Cette thèse a porté sur la synthèse de cycloheptatriènes (CHT) silylés et leur désymétrisation pour l’obtention de mimes de sucres. Nous avons ainsi développé une approche synthétique d’analogues de calystégines et mis en évidence un réarrangement inattendu conduisant à un squelette nortropane original. Dans un second temps, nous avons étudié l’équilibre cycloheptatriène-norcaradiène (CHT-NCD) et mis au point une méthodologie de cycloaddition de CHT silylés avec des dérivés nitroso, efficace et hautement stéréosélective, pour la synthèse d’aminocarbasucres via une cascade : électrocyclisation (CHT-NCD)-cycloaddition-ouverture cationique-cycloaddition. En 3 étapes, à partir du CHT méthylsilylé, 7 centres stéréogènes, 5 liaisons C-O et 2 C-N sont formés de façon diastéréo- et régiocontrôlée. Une réaction Hétéro- Diels-Alder énantiosélective sur les CHT silylés, qui conduit en une étape à la formation de 3 centres asymétriques, a aussi été développée. / This work dealt with the synthesis of silylated cycloheptatrienes and their desymmetrisation toward the synthesis of sugar mimics. Calystegines analogues were synthesized and an unexpected rearrangement leading to an original nortropane skeleton was emphasized. In a second part, cycloheptatriene-norcaradiene (CHT-NCD) equilibrium was studied. Silylated cycloheptatriene were also shown to react through cycloaddition reaction with acylnitroso compounds, through cascade processes: electrocyclisation (CHT-NCD)-cycloaddition- cationique cyclopropane opening-cycloaddition, leading highly selectively to aminocarbasugars. Starting from methylsilylated CHT, 7 stereogenic centers, 5 C-O bonds and 2 C-N bonds were formed in only 3 steps. An enantioselective Hetero-Diels-Alder reaction was also developed.

Antibodies have been widely used as reagents, homing devices, diagnostics and as therapeutic agents against different targets in clinic and research. Recently a number of monoclonal antibodies and their drug conjugates have been approved as therapeutic agents. While these molecules have great potential in various applications and therapeutics, extensive use of full length antibodies has been hampered by the high cost of production, large molecular weight and limited ability to penetrate tumor tissues. These limitations have led to the research for antibody alternatives with lower molecular weight, similar binding and affinity properties but without the lengthy and complicate process of generating antibodies. Some examples of these efforts include minibodies, fragment antigen binding (FAB), ScFv, and synthetic antibody mimics. Although these antibody alternatives have low molecular weight, as compared to the antibody, they are either derived from full size antibodies or by a long and tedious in vitro screening process. Therefore, a rational design of molecules that mimic antibody binding is a logical first step for the development of antibody alternatives. In this study, a novel approach to design antibody mimics without involving massive experimental screening was developed. The design was developed by mapping and identifying EGFR epitope region where Cetuximab CDR binds and modifying sequences using knob-socket computational model. The binding of antibody mimics were first analyzed by using MOE to obtain the binding energy, total and preserved interactions as compared to the interactions between EGFR and Cetuximab. Further, the designed antibody mimics were used to form a peptide drug conjugate (PDC). Antibody mimics were found to specifically bind and internalized by EGFR overexpressing cell lines with three to four folds higher than control cells. Antibody mimics showed binding in nanomolar range with Pep11 with binding affinity (K D ) of 252nM as shown by SPR studies. EGFR phosphorylation studies also showed that antibody mimics were able to inhibit the binding of EGF to the EGFR in a similar fashion as Cetuximab. Specific binding, affinity and functional activity of the antibody mimics demonstrated that these peptides were able to mimic all the three important characteristics of antibodies. Peptide drug conjugate (PDC) was found to be around 10 fold more potent as compared to the drug itself towards EGFR overexpressed cancer cells. PDC also showed more than 100 fold low potency against control cells. These studies demonstrated that a rational design of molecules to mimic the antibody characteristics is feasible. The antibody mimics were also successfully applied and used as targeting moiety to design peptide drug conjugates for efficient targeted drug delivery system than antibody drug conjugates.

Introduktion: Ischemisk stroke är en av de vanligaste folksjukdomar och kan leda till funktionsnedsättning och mortalitet. Diagnostisering och tid är faktorer som spelar en avgörande roll inför behandling och prognos av ischemisk stroke. Trots att DT idag anses vara förstahandsval vid diagnostisering, har den lägre sensitivitet än MR att upptäcka ischemi. Även om MR med DWI anses ge ökade möjligheter vid diagnostisering av ischemisk stroke, varierar idag användningen mycket i Sverige. Syfte: Att sammanställa kunskap om diffusionsviktade sekvensers användbarhet inom magnetresonanstomografi för att diagnostisera och behandling av ischemisk stroke. Metod: En allmän litteraturstudie genomfördes i enlighet med Friberg (2017), där tolv artiklar från PubMed och Cinahl with fulltext analyserades och granskades. Resultat: Resultatet visar att DWI har hög sensitivitet att diagnostisera lesioner, men vid vissa fall detekteras inga lesioner trots att patienten har typiska symtom. DWI är även bra på att utvärdera om trombolys är säkert att administrera upp till tolv timmar efter symtomdebut, samt kan användas för att beräkna trombektomins förväntade resultat utifrån lesionens storlek. Slutsats och kliniska implikationer: MR med DWI kan komma att ha en betydande roll i framtiden för att förbättra diagnostisering och behandling av patienter med akut ischemisk stroke. Fler patientgrupper kommer att få möjligheten till behandling som tidigare exkluderades, vilket minskar både dödlighet och funktionsnedsättning. Effektivare protokoll behövs för att införa MR med DWI i en större skala för att diagnostisera patienter med ischemisk stroke, vilket kommer att vara en stor utmaning som kräver fler MR apparater, röntgensjuksköterskor med specialistkompetens, multiprofessionellt arbete och mer forskning.

Rigid body modeling has historically been used to study various features of the elbow joint including both physical and computational models. Computational modeling provides an inexpensive, easily customizable, and effective method by which to predict and investigate the response of a physiological system to in vivo stresses and applied perturbations. Utilizing computer topography scans of a cadaveric elbow, a virtual representation of the joint was created using the commercially available MIMICS(TM) and SolidWorks(TM) software packages. Accurate 3D articular surfaces, ligamentous constraints, and joint contact parameters dictated motion. The model was validated against two cadaveric studies performed by Chanlalit et al. (2011, 2012) considering monopolar and bipolar circular radial head replacements in their effects on radiocapitellar stability and respective reliance upon lateral soft tissues, as well as a comparison of these with a novel anatomic radial head replacement system in an elbow afflicted with the “terrible triad” injury. Rigid body simulations indicated that the computational model was able to accurately recreate the translation of forces in the joint and demonstrate results similar to those presented in the cadaveric data in both the intact elbow and in unstable injury states. Trends in the resulting data were reflective of the average behavior of the cadaveric specimens while percent changes between states correlated closely with the experimental data. Information on the transposition of forces within the joint and ligament tensions gleaned from the computational model provided further insight into the stability of the elbow with a compromised radial head.

Thyroid hormones, which are secreted by the thyroid gland, are one of the most important halogenated compounds in the body. Thyroid hormones control almost every processes in the body including growth, body temperature, protein synthesis, carbohydrate and fat metabolism, heart rate, and cardiovascular, renal and brain function. Thyroid gland secretes L-thyroxine or 3,3',5,5'-tetraiodothyronine (T4) as a prohormone. While the biologically active hormone 3,3',5-triiodothyronine (T3) is produced by selective phenolic ring deiodination of T4, selective tyrosyl ring deiodination of T4 produces a biologically less active metabolite 3,3',5'-triiodothyronine (rT3). Tyrosyl and phenolic ring deiodination of T3 and rT3, respectively, also produces a biologically inactive metabolite 3,3'-diiodothyronine (3,3'-T2). Regioselective deiodinations of thyroid hormones are catalysed by three isoforms of a selenoenzyme iodothyronine deiodinase (DIO1, DIO2, DIO3). DIO1 can remove iodine from both the tyrosyl and phenolic rings of thyroid hormones, whereas DIO2 and DIO3 are selective towards phenolic and tyrosyl ring, respectively. Although the Figure 1. (A) Deiodination of thyroid hormones by iodothyronine deiodinases (DIOs) (A) and naphthyl-based selenium and/or sulphur compounds (B). mystery behind the origin of regioselectivity of deiodination by DIOs remains unsolved, formation of halogen bonding between selenium in the active site of DIOs and iodine of thyroid hormones has been widely accepted as the mechanism of deiodination. Halogen bonding, a noncovalent interaction between halogen and an electron donor such as nitrogen, oxygen, sulphur, selenium etc., elongates the C-I bond and impart a carbanionic character on the carbon atom that gets protonated after the removal of iodide. Apart from the deiodination, thyroid hormones also undergo decarboxylation, oxidative deamination, sulphate-conjugation to form iodothyronamines, iodothyroaetic acids and sulphated thyroid hormones, respectively. Figure 2. (A) Proposed mechanism of deiodination of thyroid hormones by deiodinase mimics. (B) Halogenation of uracil- and cytosine-containing nucleosides by hypohalous acid (HOX). Recently, naphthyl-based selenium/sulphur-containing compounds, such as compound 1 (Figure 1B), have been reported to mediate the selective tyrosyl ring deiodination of T4 and T3 to form rT3 and 3,3'-T2, respectively. Interestingly, replacement of the selenol moiety in compound 1 with a thiol decreases the activity, whereas replacement of the thiol moiety with another selenol dramatically increases the deiodination activity. Based on the detailed experimental and theoretical investigations, a mechanism involving the Se···I halogen bonding was proposed (Figure 2A). In addition to the halogen bonding between selenium and iodine atom, chalcogen bonding between two nearby chalcogen atoms was also shown to be important for the deiodination activity. Another important class of halogenated compounds in the body are the halogenated nucleosides. Myeloperoxidase and eosinophil peroxidase are heme-containing enzymes, which can convert halide ions (X¯) into a toxic reactive halogen species hypohalous acid (HOX) in presence of hydrogen peroxide (H2O2). Uracil- and cytosine-containing nucleosides are known to undergo halogenation at the 5-position of the nucleobase to form the halogenated nucleosides (Figure 2B). Interestingly, halogenated nucleosides such as 5-halo-2'-deoxyuridine are known to be incorporated in the DNA of dividing cells essentially substituting for thymidine. Incorporation of halogenated nucleosides into the DNA leads to mutagenesis, carcinogenesis and loss of genome integrity. Thymidylate synthase (TSase), the key enzyme involved in the biosynthesis of 2'-deoxythmidine-5'-monophosphate (dTMP) from 2'-deoxyuridine-5'-monophosphate (dUMP), can catalyse the dehalogenation of halogenated nucleotides in presence of external thiols. This thesis consists of five chapters. The first chapter provides a general introduction to halogen bonding, thyroid hormones and halogenated nucleosides. This chapter also briefly describes the halogen bond-mediated biochemical and biomimetic deiodinations of thyroid hormones by iodothyronine deiodinases and naphthyl-based organoselenium compounds. Dehalogenation of halogenated nucleotides by thymidylate synthase and thiol-based small molecules has also been discussed in this chapter. The second chapter of this thesis contains the regioselective deiodination of iodothyronamines (TAMs) by deiodinases mimics. TAMs are the endogenous metabolites produced by the decarboxylation of β-alanine side chain of thyroid hormones (THs). 3,3',5-triiodothyronamine (T3AM) and 3,5-diiodothyronamine (3,5-T2AM) undergoes selective tyrosyl ring deiodination by deiodinase mimics to form 3,3'-diiodothyronamine (3,3'-T2AM) and 3-iodothyronamine (3-T1AM), respectively. Interestingly, when the initial rates of deiodinations of T3 and T3AM were compared, deiodination of T3 was found to be several fold faster than that of T3AM under identical reaction conditions. To understand the ability of the iodine atoms to form Figure 3. (A) HPLC chromatogram of deiodination of T3. (B) Proposed mode of interaction of dimeric T3 and monomeric T3AM with organoselenium compounds. halogen bonding, a model selenolate (MeSe¯) was optimized with the T3 and T3AM. Although both T3 and T3AM forms the expected Se···I halogen bonding with MeSe¯, the strength of halogen bonding was found to be less for T3AM than T3. Furthermore, detailed kinetic and spectroscopic studies indicate that T3 and T3AM exist as dimeric and monomeric species in solution. The dimerization of T3 in solution was shown to have remarkable impact on the activation energy and pre-exponential factor of the deiodination reactions. Single crystal X-Ray crystallography and theoretical calculations indicated that in addition to Se···I halogen bonding, I···I halogen bonding may play an important role in the deodination of thyroid hormones by deiodinase mimics. Furthermore, the presence of heteroatoms such as nitrogen, oxygen and sulphur in the close proximity of one of the selenium atoms of deiodinase mimics was shown to have significant effect on the rate of deiodination reactions. The third chapter of the thesis focusses on the conformational polymorphism and conformation-dependent halogen bonding of L-thyroxine. Synthetic version of L-thyroxine (T4) is a life-saver for millions of people who are suffering from hypothyroidism, a thyroidal disorder recognised by low levels of T4 and elevated levels of TSH in blood plasma. Synthetic version of L-thyroxine is available in the Figure 4. Ball and stick model of the single crystal X-Ray structure of the conformational polymorphs of L-thyroxine. Form I and Form II was exclusively crystallized from methanol and acetonitrile, respectively. Water molecules are omitted for clarity. market with various brand names. However, adverse effects have been observed in the patients when they switch their brand of thyroxine. Based on these observations, the American Thyroid Association (ATA), the Endocrine Society (TES), and the American Association of Clinical Endocrinologists (AACE) declared that the different brands of T4 are not bioequivalent, thus leading to differences in the bioavailability of the drug. We have shown that the commercially available thyroxine exists in at least two stable forms (Form I and Form II) with different three-dimensional structures (Figure 4). These two forms exhibit different intermolecular interactions in crystal packing, spectral behaviours, thermal stabilities, optical activity and very interestingly, different solubility in acidic and basic pH. At pH 4, solubility of Form I is about 42% and 45% greater than that of Form II and bulk T4, respectively, whereas at pH 9, the solubility of Form II is about 38% and 42% higher than that of Form I and bulk T4, respectively. As T4 is a narrow therapeutic index drug, these differences in solubility may have remarkable impact on the bioavailability of the drug. In addition to this, we have shown that the ability of the iodine atoms in the C-I bonds to form halogen bond with donor atoms can be altered by changing the relative orientation of tyrosyl and phenolic rings in T4. In the fourth chapter, the three-dimensional structures and conformations of thyroid hormones (THs) and iodothyronamines (TAMs) are discussed. TAMs, the endogenous decarboxylated metabolites of THs, exhibit different binding affinities to the transport proteins and iodothyronine deiodinases (DIOs) compared to the THs. Figure 5. Change in the structure and conformations of thyroid hormones and iodothyronamines with the decarboxylation of amino acid side chain and deiodination of phenolic and tyrosyl ring. Furthermore, the substrate specificities of DIOs have been found to be dependent on the position of iodine atoms on the phenolic and tyrosyl ring of TAMs and THs. Single crystal X-ray structures of TAMs indicate that decarboxylation of amino acid side chain of THs induces significant changes in the structure and conformation. Furthermore, the positional isomers of THs and TAMs exhibit remarkably different conformations, which may have significant effect on the binding of these metabolites to the active site of DIOs. In addition to the structure and conformations, different categories of the intermolecular halogen···halogen (X···X) interactions in the crystal packing of THs and TAMs have also been discussed. Natural bond orbital (NBO) analysis have been done on the halogen-bonded geometries to understand the electronic nature of these interactions. In the fifth chapter, the dehalogenation of halogenated nucleosides and nucleobases by naphthyl-based sulphur/selenium compounds is discussed. Purine and pyrimidine nucleosides are halogenated at various positions of the aromatic ring by different peroxidases such as myeloperoxidase and eosinophil peroxidase present in the white blood cells. Incorporation of the halogenated nucleosides into the DNA of replicating cells leads to DNA-strand breaks, mutagenesis, carcinogenesis and loss of Figure 6. (A) Dehalogenation of halogenated nucleosides. Effect of base-pairing wih adenine and guanine on the deiodination of IU (B) and debromination of BrU (C) by compound 2. genome integrity. We have shown that the naphthalene-based organoselenium compounds such as compound 2 can mediate the dehalogenation of 5-iodo-2'-deoxyuridine (5-IdUd) and 5-bromo-2'-deoxyuridine (5-BrdUd) to produce 2'-deoxyuridine (dUd) (Figure 6A). The deiodination of 5-IdUd was found to be faster than the debromination of 5-BrdUd by compound 2. The mechanism of dehalogenation of halogenated nucleosides by compound 2 was found to be dependent on the nature of halogen. While the deiodination of 5-IdUd by compound 2 follow halogen bond-mediated pathway like thyroid hormones, debromination of 5-BrdUd follow a Michael addition-elimination pathway. Similar results were obtained when 5-iodo-2'-deoxycytidine (5-IdCd) or 5-bromo-2'-deoxycytidine (5-BrdCd) was used as substrate for dehalogenation reaction. Base-pairing of 5-iodouracil (IU) and 5-bromouracil (5-BrU) with adenine and guanine has a significant effect on the rate of dehalogenations of IU and BrU by compound 2 (Figure 6B and 6C).

Thyroxine is the main secretory hormone of thyroid gland and it is produced in thyroglobulin by thyroid peroxidase/hydrogen peroxide/iodide system. After biosynthesis and secretion of thyroxine, it undergoes multiple metabolic reactions. The most important metabolic pathway is the stepwise deiodination from the inner ring or outer ring. Removal of one of the outer ring or phenolic ring iodines of biologically less active T4, leads to the formation of 3,5,3'-triiodothyronine or T3, a compound which is biologically more active. On the other hand, removal of one of the inner ring or tyrosyl ring iodines gives 3,3',5'-triiodothyronine (3,3',5'-T3 or rT3) which is a biologically inactive thyroid hormone. Three enzymes involved in this activation and inactivation pathway of thyroid hormones are known as iodothyronine deiodinases (IDs), which are dimeric integral-membrane selenoproteins. Depending upon the sequence and substrate specificity, three iodothyronine deiodinase enzymes have been identified, iodothyronine deiodinase-1 (ID-1), iodothyronine deiodinase-2 (ID-2) and iodothyronine deiodinase-3 (ID-3). ID-1 can catalyze both inner ring and outer ring deiodination of thyroid hormones whereas, ID-2 is selective to the outer ring deiodination. The type-1 and -2 deiodinases (ID-1 and ID-2) produces the biologically active hormone 3,5,3′-triiodothyronine (T3). These two enzymes also convert 3,3′,5′-triiodothyronine (reverse T3 or rT3) to 3,3′-diiodothyronine (3,3′-T2) by outer-ring deiodination (Scheme 1). The type-3 deiodinase (ID-3) catalyzes the convertion of T4 to rT3 by an inner-ring deiodination pathway. Apart from deiodination, there are several alternate pathways of thyroid hormone metabolism, which include sulfate conjugation and glucoronidation of the phenolic hydroxyl group of iodothyronines, the oxidative deamination and decarboxylation of the alanine side chain to form thyroacetic acid and thyronamines, respectively. Glucoronidation and sulfate conjugation changes the physico-chemical properties of iodothyronines dramatically. This thesis consists of five chapters. The first chapter provides a general introduction of biosynthesis of thyroid hormones and followed by deiodination by three iodothyronine deiodinase enzyme. This chapter also provides an overview of thyroid hormone transport and different transport proteins and their mode of binding with thyroid hormones. Apart from this, this chapter also provides a brief overview on other thyroid hormone metabolites. In the second chapter of the thesis, initial attempts in the development of different iodothyronine deiodinase mimics have been discussed. Goto et al have shown that the sterically hindered selenol 1 converts the thyroxine derivative 3 (N¬butyrylthyroxine methyl ester) to the corresponding triiodo derivative 4 by an outer-ring deiodination (Scheme 2). Although the reaction was carried out in organic solvent and a relatively higher temperature (50 °C) and longer reaction time (7 days) were required for about 65% deiodination, this study also provides an experimental evidence for the formation of selenenyl iodide (2) in the deiodination of a thyroxine derivative by an organoselenol. However, only one iodine was removed from the outer ring of 3, no inner ring deiodination was detected (Scheme 2). Interestingly, when compound 5 was treated with selenol 1 under similar conditions, no deiodination was observed (Scheme 3). This leads to assumption that presence of free phenolic hydroxyl group is important for the deiodinase activity. Based on this experimental observation, they proposed a mechanism which involves an enol¬keto tautomerism of the phenolic hydroxyl group. In the case of thyroxine, the outer-ring can undergo enol-keto tautomerism, whereas due to lack of free hydroxyl group, the inner ring cannot undergo similar kind of tautomerism. The enol-keto tautomerism probably makes the outer ring iodines more reactive than the inner ring iodines of thyroxine. We have developed tthe first chemmical modell for the inneer ring deioddination of TT4 and T3 by type 33 deiodinase . We have shown that naphthyl-baseed selenol 6 bearing a thhiol group in the cloose proximitty to the sellenium act aas an excelleent model foor ID-3 by selectively deiodinatting T4 andd T3 to prodduce rT3 annd 3,3'-T2, rrespectively,, under physiological relevant conditions. When 2 equuivalent of ccompound 66 was emplooyed in the assay, an almost quuantitative cconversion oof T4 to rT3 was observeed within 300 hours and there was no indicaation of the fformation off T3 or 3,3'-TT2. When the selenol group was repplaced with a thiol group in compouund 7, the ddeiodinase activity wwas decreassed. On thee other handd, when thee thiol groupp was replaaced with selenol mmoiety in commpound 8, thhe deiodinasse activity drramatically iincreased wiithout any change iin the selecttivity. Comppounds 10 and 11 havving N-methhylamino grooup were found too be more aactive than the correspponding unssubstituted ccompounds 7 and 8, respectively. However, introduction of a secondary amine adjacent to the selenol moiety into the compound 9 significantly reduces the deiodinase activity. In the third chapter synthesis, deiodinase activity and mechanism of deiodination of a series of peri-substituted naphthalene derivatives is discussed. Iodobenzene was used as halogen bond donor for the DFT calculations. From the orbital analysis it is observed that there is perfect orbital symmetry match between the HOMO of compound 8 (selenolate form) and LUMO of iodobenzene. When the selenolate form of 1-selenonaphthol interacts with iodobenzene, a halogen bonded adduct is formed. The negative charge on the selenium center decreases as it donates electron pair to the σ* orbital of C–I bond in iodobenzene and as a consequence the positive charge on the iodine center decreases (Figure 1). Addition of iodobenzene to 1-selenonaphthol led to a significant downfield shift in 77Se NMR spectrum of 1-selenonaphthol and with an increase in the concentration of iodobenzene, more downfield shift in the signal was observed. Figure 1. The charges obtained from Natural Bond Orbital (NBO) analysis for the selenolate form of (a) 1-selenonaphthol (b) iodobenzene, (c) halogen-bonded adduct On the basis of experimental end theoretical data, a mechanism for the deiodination of T4 by compound 8 is proposed. According to the mechanism, the initial interaction of one of the selenol moieties with an iodine leads to the formation of halogen bond. The transfer of electron density from selenium to the σ* orbital of the C−I bond generates a σ-hole or partial positive charge on the selenium atom, which facilitates an interaction between the halogen bonded selenium atom and the free selenol (selenolate) moiety (intermediate 12). The selenium−selenium interaction (chalcogen bond) strengthens the halogen bond, leading to a heterolytic cleavage of the C−I bond. The protonation of the resulting carbanion leads to the formation of rT3. On the other hand, the formation of an Se−Se bond produces the diselenide 13 with elimination of iodide as HI. The reductive cleavage of the Se−Se bond in compound 13 regenerates the diselenol 8 (Figure 2). In the fourth chapter deiodination of sulfated thyroid hormones is discussed. Sulfate conjugation is an important step in in the irreversible inactivation of thyroid hormones. Sulfate conjugation of the phenolic hydroxyl group stimulates the inner ring deiodination of T4 and T3 but it blocks the outer ring deiodination of T4 by ID-1. The thyroxine sulfate (T4S) undergoes faster deiodination as compared to the parent thyroid hormone T4. Only ID-1 catalyzes the deiodination of sulfated thyroid hormones. In contrast, ID-2 and ID-3 do not accept T4S and/or T3S as substrate. We have shown that iodothyronine sulfates can be readily deiodinated by synthetic deiodinase model compound 8 and its derivatives. In contrast to the inner ring-selective deiodination of T4, the synthetic compounds loses the selectivity and mediate both inner and outer-ring deiodination of T4S and outer ring deiodination of rT3S. From this study, we have also proposed that the enol-keto tautomerism is probably not required for the outer ring deiodination and the strength of halogen bonding controls the regioselective deiodination by model compounds. In the fifth chapter, the mechanism of inhibition of iodothyronine deiodinases by PTU and IAA is discussed with the help of model compounds. In the model study, it has been observed that compound 8 does not form a stable Se-I intermediate (14), which is essential for the formation of Se-S covalent bond with PTU. As a consequence, the deiodination of T4 by compound 8 is not inhibited by PTU. This study supports the proposal that ID-3 does not follow a ping-pong bi-substrate pathway for deiodination and may not form a stable E-Se-I intermediate, which is responsible for the insensitivity of ID-3 towards PTU. The biphenyl based diselenol 15 reacts with IAA and iodoacetamide to form the corresponding carboxymethylated product 17. On the other hand, compound 8 does not undergo the expected carboxymethylation by IAA and iodoacetamide, but they readily deiodinate both IAA and iodoacetamide. Based on this model study, a possible model is proposed for the insensitivity of ID-3 towards IAA. Iopanoic acid (18) is a well known radiocontrast agent and is used as adjunctive therapy with PTU and CBZ for the treatment of thyrotoxicosis.[9] We show in this chapter that iopanoic acid undergoes monodeiodination by compound 8 under physiological relevant conditions. The deiodinated products (19 and 20) from iopanoic acid are characterized by NMR spectroscopy and single crystal X-ray crystallography. It is observed that after monodeiodination, the strength of halogen bonding decreases and therefore, the monodeiodinated products do not undergo further deiodination.

This thesis entitled “Investigating the Intercarbonyl X···C′ (X = O/S/N) Interactions in Short Peptides and Peptidomimetics. Evidence of Charge→π* Interactions. Synthesis and Characterization of Thioimidate Isostere Containing Peptidomimetics” is divided into two chapters. First chapter is further subdivided into four sections where investigation of the nature of intercarbonyl X···C′ (X = O/S/N) interactions in short peptides and peptidomimetics has been described. The second chapter also has been subdivided into three parts where the syntheses and characterization of thioimidate (1,3-thiazine) and imidate (1,3-oxazine) isostere containing peptidomimetics have been discussed. Chapter 1: Section A: Revisiting the earlier models for the intercarbonyl O···C′ interactions The proximity between carbonyls is ubiquitous in crystals. Here we review the key reports that have assigned an n→π* nature to interactions between carbonyl oxygen (O) atoms and adjacent carbonyl carbon (C′) atoms (O···C′). Based on earlier hypotheses (by Burgi-Dunitz) that suggest that “the minimum energy trajectory of a nitrogen nucleophile adding to the C′ of carbonyl is at N···C′ distances of ≤ 3.2 Å and along N···C′=O angles of 109±10o”, the optimum trajectory for addition of an O to an adjacent C′ has also been assigned to be the same (O···C′ distance ≤ 3.2 Å and O···C′=O angle is 109±10o). Additionally, all O and C′ atoms within these boundary conditions in crystal structures were assigned a status of interacting and those outside of the same as non-interacting. Based on quantum mechanical models for electronic orbitals that contain the valence electrons of such proximal O and C′ atoms – derived through NBO (Natural Bond Order) calculations (on crystal structures) – it has been proposed that the filled non-bonding lone pair orbital of the O (donor) overlaps with the empty π* orbital of the carbonyl C′ (acceptor), in these O···C′ interactions. Hence, these have been termed as n→π* interactions. Using DFT (Density Functional Theory) calculations energies for these interactions have been predicted to range from 0.5 to 5.0 kcal mol-1, which are similar to those for other strong non-covalent interactions such as H-bonding, weak cation-π, etc. This n→π* interaction model is assumed to prevail between adjacent carbonyls (Oi-1···C′i) at Xaa-Pro dipeptide motifs and to be exclusively responsible for the changes in equilibrium constant values (Kc/t) for the trans to cis isomerisation reaction at Xaa-Pro peptide bond in chosen analogue molecules. Based on this assumption, these Kc/t values have been used as direct experimental equivalents for the energies of these n→π* interactions. Simultaneous to such review of literature, this chapter highlights several anomalies in this n→π* model for the intercarbonyl O···C′ interactions. We discuss the alternate models that also exist for the O···C′ proximities and show that several features – such as improved pyramidalization at the acceptor carbonyl; decrease in Kc/t values at Xaa-Pro peptide bonds; and small changes in 13C NMR chemical shift values for the acceptor carbonyls; etc. – that accompany the shortening of O···C′ distance, can be explained without invoking the n→π* interaction model. Moreover, we discuss key observations such as the presence of near-symmetric antiparallel short contacts between carbonyl groups (C=O) in crystal structures, which cannot be explained by the quantum mechanical n→π* model for the O···C′ interactions. Chapter 1: Section B: Spectroscopic and kinetic investigations into the nature of X···C′ (X = O/S) interactions in N-acyl homoserine lactones (AHLs) In this section the key interactions involving the adjacent carbonyls in model N-acyl homoserine lactones (AHLs) (which are signalling molecules in quorum sensing) in solution, their electronic nature and their influence on solvolysis of the lactone ring have been investigated. Earlier, in the crystal structures of two sterically encumbered synthetic AHL analogues N-trimethyl acetyl homoserine lactone and N-tribromoacetyl homosrine lactone the presence of an n→π* orbital overlap type interaction between Oacyl and C′lact had been suggested. Based primarily on this, the operation of similar OacylC′lact interaction was proposed in all AHLs in their solution conformations as well. More intriguingly, the interaction was hypothesized to decrease the rates of lactone hydrolysis, rendering AHLs with longer biological half-life. This is contrary to physical organic understanding of nucleophilic catalysis of addition to carbonyls. Here we synthesize a variety of AHLs and analyze their NMR and FT-IR data in solution. The spectral data reveal that the role of the N-acyl group in AHLs is to withdraw eˉs from lactone C=O inductively and to improve electronic shielding at C′lact. Lack of appreciable changes in C=O stretching frequencies of lactone and 13C NMR chemical shift values of C′lact indicate the absence of electronic perturbation of the π* of the lactone. Similar non-variance of spectral bands with improvement in nucleophilicity of the N-acyl group indicates the absence of any evidence for n→π* nature for the O···C′ interactions (between the lone pair of eˉs from Oacyl to π* at C′lact). Further the spectroscopic data indicate that any change in charge at the acyl O is felt by C′lact and this weak interaction releases energy in the order of ≤ 0.05 kcal mol-1. The combined influence of the electron withdrawing N-acyl group and the weak Oacyl···C′lact interaction in AHLs is that, increasing the charge at Oacyl increases the rate of solvolysis of lactone. Analysis of the conformation of the lactone ring in the LuxR receptor-bound and unbound crystal structure forms reveals the flattening of the puckered ring in the LuxR bound state – facilitated by several interactions with the receptor. Conserved interactions between LuxR and AHLs lock the N-acyl carbonyl motif such that they are orthogonal to the lactone carbonyl and intramolecular interaction between Oacyl and C′lact is precluded. We propose the design of flat cyclic analogues of γ-butyrolactone bearing electron withdrawing side chains as potential molecules for taking advantage of bacterial quorum sensing in environmental applications and biotechnology. Chapter 1: Section C: Spectroscopic investigation into the nature of intercarbonyl X•••C′ (X = O/S/N) interactions: Carbamyl-cisPro model systems In this section we investigate the nature of intercarbonyl X···C′ interactions in carbamyl-Pro model systems using spectroscopic methods like FT-IR and 1D NMR. Further we derive the enthalpic and entropic contributions towards the free energy for trans to cis isomerization (Kc/t) at these model carbamyl-Pro systems. Our results reveal that changes in Kc/t values cannot always be used as proof for the presence or absence of electronic interactions, and hence to unambiguously suggest the nature of these interactions. Cis/trans isomerism exists at Xaa-Pro amide and carbamate motifs, and it was proposed that in acyl-Pro systems the O···C′ interactions are responsible for the stability of either cis or trans depending upon their direction of operation (Forward direction: O of Xaa is the donor of electrons to π* at C′ of Pro; Reverse direction: O of Pro is the donor of electrons to π* at C′ of Xaa). Investigation of the carbamyl-Pro systems can shed further light on this hypothesis. Hence we undertook the first spectroscopic and Van’t Hoff analysis of homologous carbamyl-Pro model systems. The Kc/t of the homologous series surprisingly increased with increase in the bulk at R (R varies from Me to tBu). The spectroscopic data revealed the presence of charge→σ* interactions at carbamyl groups. This interaction locks the carbamyl motif in the s-transoid conformation, along the C′-O σ-bond. Such conformational lock is observed to be greater in carbamyl groups where R has at least one Cα-H bond. Interestingly, we observe the absence of X···C′ electronic interactions that may selectively stabilize the cisPro conformer in these molecules. Van’t Hoff analyses on the other hand showed that as the number of Me substituents in R increases (R = Me to iPr), there is a favorable increase in entropy ( So) associated with the transPro to cisPro conformational isomerism. As a result, the population of the cisPro conformer improves significantly as the steric bulk at R increases. We note that the enthalpy of cisPro is however relatively small and remains unfavourable as R-bulk increases (Me to iPr). These data reveal the influence of electrostatic interactions between charged groups, on the change in entropy associated with cis/trans isomerism at carbamayl-Pro motifs. This not only opposes the n→π* model, but also provides an example for the important point that changes in Kc/t can/should not be taken as direct evidences of any single electronic interaction. Importantly, this study provides another example where electronic interactions between charged, polarized carbonyl motif rather than nonbonding lone pair eˉs of carbonyl motifs influence cis/trans isomerism at Xaa-Pro systems. Chapter 1: Section D: Investigation of the stereoelectronic nature of the X···C′ (X = O/S) contacts In this section we provide experimental evidence for the existence of inverse correlation between the charge on the O nucleophile and the O···C′ distances. We show that O and C′ atoms (of adjacent carbonyls), which are separated at distances > 3.20 Å in carefully chosen analogues, come together to σ-bonding distances when the charge on O is increased to -1. Additionally, the influence of backbone steric factors on these charge→π* interactions is investigated. A partial covalent nature was proposed for the O···C′ interactions. Our study showed that the shortest intercarbonyl O···C′ distances between the O of 1°, 2° and 3° amide carbonyls and proximal C′ in molecules found in the Cambridge Structural Database (CSD) (v5.36, November 2014) show an inverse linear correlation with the partial negative charge (δ‾) on the amide carbonyl O rendered by natural amide carbonyl polarization. These data suggest the interaction of charge on the nucleophilic O with π* of the acceptor carbonyl. Further on increasing the charge on nucleophilic carbonyl O to -1 in the model compound, we achieved the formation of σ-bond through non-native (natively disallowed) Oi‾¹→C′i-1 interaction. Here we provide the first experimental evidences that suggest the interaction between charge of O and π* at adjacent C′ (the charge→π* interaction) and the latent covalent nature of the O···C′ interactions. This charge→π* model explains the origins of variations in O···C′ distances (3.20 Å–1.43 Å) in proteins and complexes that occur to suit biological functions; and the mutual interactions between antiparallel carbonyls. Further the effect of 3 key steric factors – namely the allowed τ (N-Cα-C′) angle, entropy and allowed (ϕ,ψ) angles – on the non-native Oi→C′i-1 interactions were investigated in the model compounds. Our kinetic data revealed that, the allowed τ angles have the greatest influence on charge→π* interaction, followed by entropy. Importantly the allowed (ϕ,ψ) torsional angles for residues, that govern protein folding pathways, have little influence on the O···C′ electronic interactions. Chapter 2: Section A: Design and synthesis of novel 1,3-Thiazine containing peptidomimetics This section describes the first synthesis of peptidomimetics containing the 1,3-thiazine isostere (thioimidate isostere for the peptide bond), at the C-terminus and also at the middle of the peptide. The synthesis of the 6-membered heterocycles – 1,3-oxazine (Oxa) – have earlier been reported. Oxa motifs constrain preceding amino acid backbones into natively disallowed conformations. Here we present the first synthesis of peptidomimetics containing the 1,3-thiazine (Thi) (the thioimidate analogue of Oxa) motif, by the treatment of N-(3-hydroxypropyl)thioamides with MsCl/Et3N, which leads to intramolecular S-alkylation / cyclization. When placed at the C-terminus of acyl-Pro motifs the Thi group selectively improves the stability of the rare s-cis conformation of the acyl-Pro peptide bond. Further this method has been used to synthesize peptidomimetics in which an endogenous peptide bond is replaced with the Thi isostere. These Thi analogues are shown to be stable to standard conditions of peptide coupling and N- and C- terminus protection, deprotection and can be extended selectively at their N- or C- termini. Chapter 2: Section B: Epimerization in 1,3-Thiazine containing peptidomimetics The epimerization in 1,3-thiazine containing peptidomimetics and its mechanism has been described in this section. Further the aggregation behaviour of these thiazines, in solution and crystal structures, has been studied. It has been well-documented that epimerization (Racemization) occurs at the chiral centers at the C(2) exo methine of 1,3-thiazolines and 1,3-thiazoles. Similar epimerizations in 1,3-thiazines have however not been explored. Here we report our observation of epimerization in chiral aminoacid (non Pro) containing 1,3-thiazine peptidomimetics. Our studies revealed that, the epimerization happens at C2 positions of chiral (non-Pro) amino acids-derived 1,3-thiazine containing peptiomimetics. And NH of chiral (non-Pro) amino acid fused to Thi ring at C2 position is necessary for the epimerization. Further we investigated the Boc-Xaa*-Thi analogues in solution, which showed two resonances for the carbamate N-H (HN) and the H of Xaa*, irrespective of the side chain in Xaa, in CDCl3 a weakly polar solvent. The integral ratios of the major : minor peak increased with increase in concentration for Boc-Val*-Thi, indicating the formation of H-bonded aggregates. Even in the polar aprotic (DMSO-d6) and polar protic (D2O) solvents the two sets of resonances were observed for Boc-Val*-Thi in 1H NMR. But when the thioimidate N is protonated (N of Thi is no longer a H-bond acceptor), showed only a single set of resonances. Formation of intermolecular H-bonds involving N of Thi in solution is thus evident in the aggregates. This is further suggested by the crystal structures obtained for the peptide mimetics Boc-Val*-Thi, Boc-Leu*-Thi and Boc-Phe*-Thi in which the racemic pair, instead of one enantiomer of it, are present in the unit cell and are locked in a pair of intermolecular 10 membered H-bonding interactions between NThi and HLeu* similar to an antiparallel β-sheet. A mechanism for racemization is proposed, where this strong H-bond assists enamination/racemization process. Chapter 2: Section C: Influence of a disallowed conformation of Aib on the structure of a 310-helical fold In this section, the effect of the presence of a disallowed conformation of Aib at the C-terminus of a 310-helical peptide, on the structure and fold of the rest of the peptide body has been studied in solution. We constrain the C-terminal Aib in the Aib-rich octapeptide (N-tert-butoxycarbonyl-Leu1-Aib2-Ala3-Leu4-Aib5-Ala6-Phe7-Aib8-CO2Me (1), which adopts a complete 310-helical conformation throughout the peptide body in the crystal structure and in solution) in one of its disallowed conformations using a method earlier developed in our group. This involves the synthetic modification of the C-terminal ester (Aib8-CO2Me) in 1 to an Oxa (Aib*8-Oxa) in 2 and the study of its effect on the peptide body. Analyses of the solution FT-IR, CD, ¹H, 2D (TOCSY, HSQC, HMBC and ROESY) and solvent polarity dependent NMR data reveal that 2 adopts a 310-helical conformation similar to that of 1. The C-terminal CO2Me → Oxa (E → O) modified Aib*8-Oxa motif is constrained in a unique conformation where the two Cβ atoms of Aib*8 are staggered with respect to the Aib*8 C=O and are both interacting with the two Hβ of Phe7. Here the Aib* backbone is constrained by a 5-membered ring NOxa∙∙∙HAib* H-bond, in a C5i structure. Solvent polarity dependent ¹H NMR data indicate the formation and persistence of C5i H-bond at the Aib*8-Oxa motif in 2. Analyses of the ROESY, solvent polarity dependent ¹H NMR and CD spectra reveal that four crucial changes in ROESY cross peaks occur at the Phe7-Aib*8 motif of 2, compared to that in 1. From these spectroscopic data it has been confirmed that there is no change in the structure of 2 from Leu1 to Ala6. Whatever the crucial changes happened are at Phe7-Aib*8 motif of 2. Hence our study showed that the significant structural consequences of this disallowed conformation of Aib* are primarily observed to occur in the residue in its immediate vicinity, rather than in the whole peptide body. Presence of a disallowed fold at a residue need not result in disruption of the structure, or the overall fold, in the rest of the peptide body.