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This thesis describes the preparation of halogen bonding and chalcogen bonding macrocyclic and mechanically interlocked systems for the recognition of anions and ion-pairs. Chapter 1 introduces the field of supramolecular host-guest chemistry, with particular focus on the recognition of anions and ion-pair species. The synthesis and applications of mechanically interlocked molecules are also reviewed. Chapter 2 details the fundamental charged guest recognition properties of a novel family of σ-hole [2]rotaxanes incorporated with multiple halogen bonding and chalcogen bonding triazole-based donor motifs. Chapter 3 investigates the anion sensing capability of halogen bonding receptors functionalised with photo- and redox-active reporter groups. This entails the construction of zinc(II) metalloporphyrin-BODIPY [2]rotaxanes for the dual optical- and electrochemical sensing of anions. Chapter 4 describes the exploitation of halogen bonding and chalcogen bonding interactions to mediate ion-pair recognition in heteroditopic/multitopic macrocyclic and catenane host systems. Chapter 5 summarises the conclusion of this thesis. Chapter 6 provides the experimental procedures employed in this work, including synthetic protocols and characterisation of novel compounds.

Human immunodeficiency virus (HIV) and other RNA viruses pose a major threat to human health. Current therapeutic options are limited due to the multifaceted protein-RNA interactions that drive the lifecycle of HIV. The human heterogeneous nuclear ribonucleoprotein (hnRNP) H is a dynamic RNA-binding protein (RBP) that participates in many stages of RNA metabolism in humans and viruses alike, including in HIV type 1 (HIV-1). G-tracts—stretches of RNA consisting of three or more consecutive guanines—are the binding partners of hnRNP H. G-tracts are important splicing elements, and, due to their abundance, HIV-1 can recruit hnRNP H to bind to viral G-tracts within its genome. The interactions between hnRNP H and G-tracts can be studied using common 2D NMR techniques such as heteronuclear single quantum correlation (HSQC) or nuclear Overhauser effect spectroscopy (NOESY), but they are often cumbersome and require isotopic enrichment in nutrient-poor media. Fluorine (19F) NMR spectroscopy is a viable 1D alternative to 2D methods used for characterizing proteins, owing to both the excellent sensitivity of 19F and the much lower time cost it takes in running 1D versus 2D NMR experiments. The usefulness of 19F NMR is highlighted through two separate projects that reveal mechanistic insights into hnRNP H:G-tract molecular recognition. 19F is NMR active and is not found in essentially any biological system, making it suitable for studying protein-nucleic-acid interactions. 19F NMR spectroscopy can be coupled with paramagnetic relaxation enhancement (PRE), the effect of which can be measured in order to study non-specific interactions in a dynamic environment. Here, 19F NMR PREs are used in order to gain both qualitative and quantitative distance-dependent information about hnRNP H:G-tract binding (Chapter 2). 19F NMR spectroscopy can also be used to monitor changes in protein-RNA complexes resulting from drug binding as part of a larger screening pipeline dubbed “Targeting of Specific Protein-RNA Interfaces detected by NMR” (To-SPRIN) (Chapter 3). From these projects, greater insight is obtained into the molecular mechanisms that drive the interactions be-tween hnRNP H and its G-tract binding partners.

Molecular recognition at biomembranes is one of the more poorly understood aspects of fundamental research in physical organic chemistry. Our aim was to improve our understanding of the molecular recognition of polysaccharides at biomembranes, in particular developing synthetic lipids that will recognise and report on the presence of glycosaminoglycans (GAG polysaccharides), like heparin and hyaluronic acid. Elevated levels of hyaluronic acid have been implicated in bladder carcinoma and osteoarthritis, and the use of heparin for medical applications is well documented. We synthesised a boronic acid capped lipid that also bore a fluorinated fluorescent reporter group, which could report on multivalent recognition events at bilayer membranes by fluorescent quenching and changes in the lateral distribution of the reporter groups. These preliminary studies showed these boronic acid capped fluorinated lipids gave a fluorescent signal upon interaction with simple mono- and poly- saccharides, albeit with unexpectedly weak binding to these saccharides. To understand and quantify the weaker binding of saccharides to membrane bound boronic acids a series of novel fluorescent and chromogenic lipids were synthesised that bore the reporter group close to the boronic acid. These studies revealed several underlying factors that had important roles in the recognition of oligosaccharides by boronic acid capped lipids. For the first time the effect of the bilayer on saccharide/boronic acid recognition was quantified, with the membrane weakening the interaction 33-fold. We were able to propose a model for the interaction of saccharides for membrane bound boronic acids that explained many of these unexpected observations.We also devised a parallel approach using GAGs to open or close synthetic membrane channels. Using a GAG to switch on the release of an ion or dye would generate a fluorescent signal that amplifies the original recognition event and improves sensitivity for GAGs. Proof-of-principle studies using palladium ions to open dye-transporting channels were successful and these studies were followed by the synthesis of boronic acid-capped cholates. Incorporation of boronic acid-capped cholates into membranes caused changes in the rate of release of alkali metal ions, which caused an enclosed fluorescent dye to give a signal, in the presence or absence of saccharides. These compounds successfully gave a response to the simple saccharide D-fructose but gave no response to other saccharides tested, including various hyaluronic acids. Although we were not able to develop a selective sensor for GAGs, we have developed a model for saccharide/boronic acid interactions that is a valuable addition to the physical organic chemistry of membranes.

Molecular recognition is essential to all biological interactions and processes. Knowledge of the structural basis of recognition offers a powerful mechanism for understanding, predicting and controlling the behaviour of biological systems. In this thesis, we present, firstly a computational and crystallographic analysis of molecular recognition in protein-ligand systems; and secondly, progress towards the synthesis of a fluorescent probe for calcium ion recognition. Class I phosphoinositide 3-kinases (PI3Ks), in particular PI3Kγ, have long been considered promising drug targets for the treatment of inflammatory and autoimmune disorders. As a step towards improved understanding of PI3K binding preferences, we examine the basis on which PI3Kγ distinguishes γ-selective inhibitors AS-605240 and AS-604850, with a ~30-fold preference for the former. Interestingly, despite the chemical similarity of the two ligands, the X-ray structures for their PI3Kγ complexes exhibit the molecules in different conformers, s-cis for AS-604850 and s-trans for AS-605240. Here, we re-examine the PI3Kγ/AS-605240 crystallographic data and find that not only is a s-cis conformation possible but in fact it has a much higher occupancy (87%) than the originally modelled s-trans isomer (13%). Subsequently, to account for the isomeric complexities presented by the ligands, we perform 140 ns MD simulations of the four PI3Kγ complexes in explicit solvent: this reveals similar conformational flexibility at the active site for all systems. Yet, the conformations sampled by the s-cis isomers are more consistent with the conformations reported by the X-ray crystal structures. Subsequent energetic analysis was performed incorporating ensemble-averaging and desolvation effects via the Poisson-Boltzmann continuum solvent model. For both inhibitors the s-cis isomers are predicted to be the most favourable conformations. Additionally, the results indicate a preference for AS-605240, as observed experimentally. The molecular basis for this preference is discussed, together with a comparison of molecular mechanical and quantum chemical treatments of the key ligand-Val882 interaction. This study provides structural, dynamical and energetic insights into the subtle basis of molecular recognition by PI3Kγ.Fluorescent probes have evolved into an extremely useful tool for the detection of calcium in biological systems. Benzothiazole derivatives BTC, and its iminocoumarin analogue BTIC, are two low affinity calcium indicators featuring many desirable properties for cellular calcium measurement. In an effort to produce fluorophores that can be chemically conjugated with a screening protein, such as Green Fluorescent Protein (GFP), we aimed to derivatise BTC and BTIC. Two synthetic approaches towards the synthesis of these potential fluorescent probes are outlined.

The failure of the pharmaceutical industry to increase the delivery of new drugs into the market is driving a re-assessment of practices and methods in drug discovery and development. In particular alternative strategies are being pursued to find therapeutics that are more selective, including small molecules that target protein-protein interactions. However, success depends on improving our understanding of the recognition of small molecules by interfaces in order to develop better methods for maximising their affinity and selectivity, whilst trying to confer an appropriate therapeutic profile. This thesis starts with the description of the creation of TIMBAL, a database that holds small molecules disrupting protein-protein interactions. The thesis then focuses on the analysis of these molecules and their interactions in a medicinal chemistry and structural biology context. TIMBAL molecules are profiled against other sets of molecules (drugs, drug-like and screening compounds) in terms of molecular properties. Using the structural databases in the Blundell group, the atomic detail of the interaction patterns of TIMBAL molecules with their protein targets are compared with other molecules interacting with proteins, comprising natural molecules, small peptides, synthetic small molecules (including drug-like and drugs) and other proteins. The structural features and composition of the binding sites of these complexes are also analysed. Keeping in mind that current drug candidates are somewhat too lipophilic to succeed, these interaction profiles are defined in terms of polar and apolar contacts, with the aim of migrating natural patterns into the design of new therapeutics.

Dynamic combinatorial chemistry (DCC) is a powerful method for synthesising complex molecules and identifying unexpected receptors. Chapter 1 gives an overview of the concept of DCC and its applications, and discusses its evolution to date. Chapter 2 describes the discovery of a new generation of donor-acceptor [2]catenanes in aqueous dynamic combinatorial systems. The assembly of these [2]catenanes is promoted by a high salt concentration (1 M NaNO3), which raises the ionic strength and encourages hydrophobic association. More importantly, a mechanism that explains and predicts the structures formed is proposed, giving a fundamental insight into the role played by hydrophobic effect and donor-acceptor interactions in this process. Building on these results, Chapter 3 describes the assembly in high salt aqueous libraries of a larger structure: a [3]catenane. Remarkably, the [3]catenane exhibits strong binding interactions with a biologically relevant target - spermine - in water under near-physiological conditions. Its synthesis is improved if the salt is replaced by a sub-mM concentration of spermine, acting as a template. Chapter 4 explores in further detail how subtle variations in the building block design influence the selective formation of either [2] or [3]catenanes. This last section underlines both the advantages and the limitations of the method developed in Chapter 3. After a short conclusion (Chapter 5), Chapter 6 gives experimental details.

This thesis describes the synthesis and investigation of novel anion templated interlocked structures which incorporate the positively charged imidazolium and triazolium binding motifs for applications in anion recognition and sensing. Chapter One introduces the fields of anion supramolecular chemistry and mechanically interlocked structures, focusing on topics of particular relevance to this thesis, including anion recognition, anion sensing and the templated synthesis of interlocked architectures. Chapter Two details the incorporation of the imidazolium motif into the axle components of anion templated rotaxane hosts to achieve selective anion recognition by virtue of their interlocked binding cavities. The effects of exploiting imidazolium motifs with contrasting hydrogen bond donor arrangements and reducing the macrocycle size on the anion recognition properties of such systems were investigated using 1H NMR spectroscopy. Chapter Three reports the work undertaken to utilise fluorescent reporter groups as stoppers in the synthesis of anion sensing rotaxanes. Imidazolium- and triazolium based systems containing either luminescent ruthenium(II) bipyridyl complexes or the organic fluorophore anthracene were prepared and their anion sensing behaviours explored using fluorescence spectroscopy. Synthetic efforts to construct suitable photo-active rotaxanes are detailed. Chapter Four describes investigations of the novel naphthalimide triazolium motif both for use in interlocked molecular motion systems, and for fluorescence sensing applications. The preparation of a naphthalimide triazolium rotaxane, capable of selective, anion-induced, uni-directional shuttling which was investigated extensively using 1H NMR spectroscopy and optically signalled by perturbations in the UV/Vis spectrum, is detailed. Preliminary research studies into the potential to exploit this motif for surface based fluorescence sensing devices are also included. Chapter Five presents research into the utilisation of acyclic receptors displaying considerable binding induced conformational changes for fluorescence anion sensing. The recognition properties of a series of imidazolium-based receptors were studied, whilst the ability of a pyrene appended analogue to signal anion complexation via changes in excimer emission is reported. The control of interpenetrated assembly formation using anion-induced conformational changes within the threading component of a pseudorotaxane is also discussed. Chapter Six provides the experimental procedures and characterisation details for the compounds synthesised in this thesis. Chapter Seven is a summary of conclusions from Chapters Two, Three, Four and Five. Supplementary information relating to titration protocols, crystallographic data and surface studies is provided in the Appendices.

Molecularly imprinted polymers (MIPs) are a novel, promising family of porous materials with potential applications ranging from separations, chemical sensing and catalysis to drug delivery and artificial immunoassays. The unique feature of these materials is their biomimetic molecular recognition functionality. Molecular recognition is the biological phenomenon of specific, selective and strong association between a substrate and a ligand. In man made MIPs this functionality is implemented via templated synthesis protocol. MIPs are synthesized in the presence of additional template molecules which form complexes with functional monomers in the pre‐polymerization mixture. After polymerization, the template is removed, leaving cavities in the structure which are complementary in shape and interaction patterns to the template molecules. These cavities act as mimics of biological receptors and are able to recognize and rebind template molecules. Although the imprinting concept is simple in principle, synthesis of MIPs with precisely controlled characteristics and performance remains a challenging task. Composition, polymerization conditions, template removal process and application conditions all affect the properties of MIPs. The material is affected at different scales, but crucially at the microscopic level, the number, fidelity and accessibility of binding sites are dependent on all the factors mentioned. The full potential of these materials can only be achieved if researchers can control and optimize the properties of MIPs through detailed understanding of adsorption and molecular recognition processes in these materials. The objective of this work is to, using computer simulations and statistical mechanics; develop a fundamental description of MIP formation and function, and to link morphological features of the model materials to their molecular recognition capabilities. In general, molecular simulations employed in this study should allow easier and more efficient exploration of a vast number of factors influencing the behaviour of MIPs. At the heart of the approach developed in this thesis is a computational strategy that imitates all the stages of MIP formation and function. First, the model simulates the pre‐polymerization mixture, allowing the formation of template‐functional monomer complexes. (This stage is implemented via canonical Monte Carlo simulation). Complexes can have different structures, depending on the chemical nature of template and functional monomer; therefore complexes can have a range of association constants. The distribution of template‐functional monomer complexes also translates into a distribution of binding sites of different specificity after template removal. In the second stage of the process, adsorption simulations (grand canonical Monte Carlo) are performed for a variety of model MIPs prepared to assess the role of various processing conditions such as composition, density and binding sites degeneration. This strategy was first applied to a simplified description of MIP species in order to identify the minimal model capable of molecular recognition and thus shed the light on the very nature of this phenomenon. In the developed model, the molecular species are constructed from hard spheres, featuring small interaction sites on their surfaces. The bond between two interaction sites has the strength and topological features of a typical hydrogen bond. The model exhibits molecular recognition, being able to preferentially adsorb template molecules. The associations between template and functional monomers were analyzed and classified to describe the distribution of binding sites and their heterogeneity. Using this model, several experimental trends typically observed in MIP studies could be explained, such as maximum in the selectivity as a function of monomer concentration. Using this model, we were also able to explore hypothetical, alternative protocols for MIP synthesis in order to improve their performance. These include the use of alternative templates and the post‐synthetic surface modifications of MIPs. The general strategy to modelling MIP, employed in this thesis, was then applied to a more detailed description of MIPs with realistic force field potentials for all the species involved. This more elaborate model is simulated with a combination of molecular dynamics (MD) and Monte Carlo techniques. This detailed model provided a wealth of information on various types of complexes observed in the pre‐polymerization mixture. Specifically, it revealed the relative weight of different interactions in the complex and their role in the binding energy of adsorbates. These simulations also provided the comparison of the relative contribution of different types of interactions (van der Waals, Coulombic) involved in a molecular recognition process. We believe the insights gained in this work will contribute to the development of rational MIP design strategies. In the discussion of the results of the thesis we speculate on how these models can be further developed in order to generate quantitative predictions and what type of systems it would be interesting and important to investigate in the future.

This thesis is concerned with the synthesis and study of novel anion templated rotaxanes and catenanes for electrochemical anion sensing, as well as interlocked structures that possess different anion binding properties, higher-order topologies and the ability to undergo molecular motion. Chapter One provides an introduction to anion recognition and the preparation of interlocked structures. A short summary of fundamental aspects of supramolecular chemistry is followed by detailed surveys of current approaches to anion binding and sensing, as well as the templated synthesis of rotaxanes and catenanes. Chapter Two describes the preparation of rotaxanes and catenanes appended with ferrocene to allow for electrochemical anion sensing. The anion recognition properties of a [2]rotaxane and a [2]catenane, as investigated by ¹H NMR spectroscopy and electrochemical methods, are presented. The utilization of a ferrocene-appended macrocycle in the construction of surface confined anion templated rotaxanes and catenanes is also discussed. Chapter Three reports the work carried out to achieve electrochemical anion sensing by the incorporation of redox-active groups into the integral structures of interlocked structures. The syntheses of a bis-stoppered 1, 2, 3, 4, 5-pentaphenylferrocene [2]rotaxane and a ferrocene containing [3]rotaxane are presented, along with their subsequent anion recognition studies. In addition, attempts to incorporate ferrocene into the macrocyclic components of rotaxanes and catenanes are outlined. Chapter Four details further investigations into the use of interlocked structures to achieve anion recognition. Doubly-charged [2]catenanes able to bind anions in aqueous solvent media, as well as the incorporation of alternative anion binding motifs into interlocked architectures are reported. The exploitation of anion templated synthesis to allow for the construction of higher order structures (including [3]catenanes, a “handcuff” catenane and a Janus [2]rotaxane), as well as a [2]catenane system with anion controlled molecular motion is also described. Chapter Five presents the experimental procedures and characterization data relating to the compounds prepared in Chapters Two, Three and Four. Chapter Six summarizes the main conclusions of the work contained within this thesis. Supplementary experimental information relating to titration protocols, investigations into self-assembled monolayers (SAMs) and crystallographic data are provided in Appendices I, II and III.

Development of selective artificial metalloenzymes by combining the biological concepts for selective recognition with those of transition metal catalysis has received much attention during the last decade. Targeting covalent incorporation of organometallic catalysts into proteins, we explored site-selective covalent coupling of phosphane and N–containing ligands. The successful approach for incorporation of phosphane ligands we report herein consists of site-specific covalent coupling of a maleimide functionalized hydrazide into proteins, followed by coupling of aldehyde functionalized phosphanes via a hydrazone linkage. Site selective incorporation of N–containing ligands was obtained by coupling maleimide functionalized N–ligands to proteins via Michael addition to the maleimide double bond. These two methods can be easily applied to virtually any protein displaying a single reactive cysteine and allows a wide range of possibilities in terms of cofactor design. Site-specific covalent incorporation of transition metal complexes of phosphane ligands into proteins was successfully obtained. The success of the approach is influenced by several factors like the metal precursor, the phosphane type and the protein scaffold. Metal complexes of 5–maleimido–1,10–phenanthroline modified proteins were formed in situ, via addition of a metal precursor to the phenanthroline modified proteins or by coupling preformed metal complexes to proteins via Michael addition of the thiol group from a cysteine residue to the maleimide double bond of the N-ligand. These successful coupling methods enable the use of a wide range of protein structures as templates for the preparation of artificial transition metalloenzymes, which opens the way to full exploration of the power of selective molecular recognition of proteins in transition metal catalysis.

Until very recently, synthetic chemistry has focussed on the creation of chemical entities with desirable properties through the programmed application of isolated chemical reactions, either individually or in a cascade that afford a target compound selectively. By contrast, biological systems operate using a plethora of complex interconnected signaling and metabolic networks with multiple checkpoint controls and feedback loops allowing biological systems to adapt and respond rapidly to external stimuli. Systems chemistry attempts to capture the complexity and emergent phenomena prevalent in the life sciences within a wholly synthetic chemical framework. In this approach, complex phenomena are expressed by a group of synthetic chemical entities designed to interact and react with many partners within the ensemble in programmed ways. In this manner, it should be possible to create synthetic chemical systems whose properties are not simply the linear sum of the attributes of the individual components. Chapter 1 discusses the role of complex networks in various aspects of chemistry- related research from the origin of life to nanotechnology. Further, it introduces the concept of Systems chemistry, giving various examples of dynamic covalent networks, self-replicating systems and molecular logic gates, showing the applications of complex system research. Chapter 2 discusses the components of replicator design. Further, it introduces a network based on recognition mediated reactions that is implemented by length- segregation of the substrates and displays properties of self-sorting. Chapter 3 presents a fully addressable chemical system based on auto- and cross- catalytic properties of product templates. The system is described by Boolean logic operations with different template inputs giving different template outputs. Chapter 4 introduces a dynamic network which fate is determined by a single recognition event. The replicator is capable of exploiting and dominating the exchanging pool of reagents in order to amplify its own formation at the expense of other species through the non-linear kinetics inherent in minimal replication. Chapter 5 focuses on the development of complex dynamic systems from structurally simple molecules. The new approach allows creating multicomponent networks with many reaction pathways operating simultaneously from readily available substrates.

In the last twenty years, chemists have devised numerous synthetic chemical systems in which self-replication operates, demonstrating that molecules can replicate themselves without the aid of enzymes and that self-replication is not a prerogative of nucleic acids only. However, the coupling of replication to other recognition-mediated events and its exploitation in the amplification of large supramolecular assemblies, such as mechanically interlocked molecules, have remained unexplored areas. Among mechanically interlocked molecules, rotaxanes represent particularly attractive targets because of their application as molecular switches. This thesis describes how the recognition-mediated synthesis of a rotaxane can be combined to the amplification of its structure by replication. Kinetic models for the integration of self-replication with the formation of a rotaxane are presented. The logical steps required to convert these models into molecular structures through consideration of the design criteria highlighted by the models are discussed and executed. The macrocyclic component is an essential part of a rotaxane. The synthesis of several novel macrocycles is presented. Their ability to bind guests in their cavities through hydrogen bonds was probed. The best macrocycle/guest pairs were integrated in the formation of rotaxanes. Further investigations on the stoppering reaction and on the various recognition processes involved in the system lead ultimately to the construction of self-replicating rotaxanes.

In the last 20 years there has been a number of synthetic and natural product based molecular replicators published in the literature. The majority of these systems have focused on the minimal model with only a few examples of cross-catalytic or reciprocal replication. Of the cross-catalytic systems investigated the majority focus around the use of natural products, oligonucleotides, peptides etc. This thesis will investigate the design, synthesis and kinetic analysis of both synthetic minimal and reciprocal replicating systems, and how these two forms of replication interact in a complex hypercyclic network. Chapter 1 introduces key concepts such as molecular recognition, intramolecularity/ enzyme kinetic, bisubstrate systems and the work conducted into replication systems to date. Chapter 2 describes the design, synthesis and kinetic analysis of a reciprocal replicating system, based on Diels-Alder and 1,3-dipolar cycloadditions, before going on to discuss what we have learned and how this system can be improved. Chapter 3 focuses on the design, synthesis and kinetic analysis of a replicating network (minimal and reciprocal replication), based on 1,3-dipolar cycloadditions. Initial individual systems are examined in isolation to determine their behavior and nature. After which the systems are combined to observe how each species interacts in a potential complex hypercyclic network. Chapter 4 investigates the redesign of the replicating network in Chapter 3 in order to overcome the problems identified from its kinetic analysis. Chapter 5 introduces the shift in direction away from kinetically controlled replicating networks towards systems in thermodynamic equilibrium.

The structures of nucleic acids play crucial roles in biological functions but are challenging to rapidly identify and classify. Our study demonstrates the effectiveness of an arrayed suite of synthetic hosts and dyes in detecting and distinguishing various DNA secondary structures. By utilizing multivariate analysis of fluorescence profiles which were generated from cationic dyes with affinities for both DNA and synthetic hosts, discrimination between G-quadruplex structures of identical length and similar topological types, i.e., parallel, antiparallel, and hybrid, is achieved. The host:guest sensor array enables the classification of diverse noncanonical DNA structures, including G-quadruplexes, Hoogsteen triplexes, hairpins, and i-motifs with high selectivity, as well as differentiation between native G-quadruplexes and those with a vacancy or bulge. Machine learning algorithms applied to the sensing data allow for accurate prediction of unknown DNA folding states. Furthermore, the sensor array can detect single oxidation or methylation modifications in one guanine base of G-quadruplexes, even in diluted serum, which provides a powerful tool for identifying structural changes caused by DNA damage. In addition, the host:guest complexes can function for “turn-on” or “turn-off” indicator displacement sensing of neurotransmitter analogs in cellular environments. The dyes selectively accumulate in nucleotide-rich regions of the cell nucleus and cytoplasm. The hosts bind the dyes and promote their relocation to the outer cell membrane. Incubating the cells with structurally similar biomarkers illustrates the selective recognition—choline and butyrylcholine can be bound by the hosts, but minimal binding is seen with betaine or cholamine. Overall, our study highlights the capability of supramolecular host:guest systems for the recognition of nucleic acid secondary structures and offers promising applications in cellular sensing.

Given that the world's population is growing quickly and is predicted to reach 9 billion by 2050, food production and sustainability are among humanity's top priorities. Since phosphorus is one of the three fundamental nutrients commonly integrated in commercial fertilizers, phosphorus (phosphate) plays a crucial role in the food production processes. Additionally, global demand for phosphorus increases roughly 3% every year, and yet, most phosphorus ends up in the waste! Most of the wasted phosphorus enters our water systems (rivers, lakes, seas, oceans) resulting in the eutrophication phenomena. The excessive amount of phosphorus and phosphates from various anthropogenic and natural resources create eutrophication, a dangerous form of water body pollution. Inorganic phosphates have enormous application in industry as both raw materials and fertilizers, often over-used in the agricultural settings. Organophosphates are a type of phosphorusbased compounds that are found to be harmful to the environment. For instance, abuse of chemical warfare agents since their invention in the 1930s has had a harmful effect on the society. Overuse of the commercially available herbicide Glyphosate (Roundup®) has been linked to the cancerogenic effects to humans.To overcome these social, ecological, and economical burdens, new scientific and technological progresses have been developed. Chemistry has been one of the most important scientific branches that has played a substantial role in tackling environmental issues, thus dramatically improving the quality of life. A qualitative and quantitative determination of inorganic and organic phosphates demands development of chemosensors capable of operating in the various media, preferably in water. The present work describes the design and synthesis of macrocyclic fluorescent based chemosensors and their utilization in the state-of-the-art sensing methods employing pattern recognition protocols for identification and quantification of various phosphates and organophosphates. Aim and contribution of this work are to demonstrate practical utility of a single selective, yet cross-reactive sensing probe for discrimination of phosphates and their mixtures, thereby simplifying pattern recognition protocols while maintaining the high overall reliability of the sensing process.

Molecular recognition between two or more molecular binding partners plays an essential role in many cellular processes and pathogen entry steps. It is widely accepted now that molecular recognition is mediated through non-covalent interactions (aka non-bonded interactions) like hydrogen bond (HB), salt bridge, π-system interactions like cation–π interaction, π–π stacking interaction, CH-π interaction, and XH-π interaction (X=N, S, O); and van der Waals interaction (VDW). In this dissertation, we have studied molecular recognition in three important biological systems, i.e., agonists/antagonists in G protein-coupled receptors, guanine in GTP-binding proteins, and spike proteins of novel coronavirus with the human ACE2 receptor. It is expected that the knowledge of molecular recognition in these selected proteins would help design of drugs targeting GPCRs and GTP-binding pockets, or design of neutralizing antibodies targeting the spike protein (RBD) of the novel coronavirus.In the first system, molecular recognition of ligands that binds to GPCRs, mainly A2A receptors, was studied. It has been reported that nearly 34% of FDA-approved drugs target GPCRs. Most drugs function as either an antagonist or an agonist of GPCRs. The molecular recognition of the ligands in A2A receptors represents a topic of great importance because of the significance of adenosine and cognate ligands in cellular physiology and the necessity to develop safe and effective medications for many pathophysiological issues. A total of 5 unique agonist-bound and 18 unique antagonist-bound complexes of adenosine A2A receptor were systematically studied. Interestingly, all agonists and antagonists feature a central heterocycle that is capable of hydrogen bonding and π-system interactions. For each complex, the interaction modes between an agonist/antagonist and its interacting residues were examined to identify all the non-covalent interactions necessary for molecular recognition. The interaction mode analysis revealed that three major interaction modes, i.e., hydrogen bond from N253, π-π stacking interactions from F168, and CH-π interaction from M177, L249 and I274, are responsible for the binding of the central heterocycle to the A2A receptor. Subsequently, strengths of intermolecular interactions are quantified by means of advanced quantum chemical calculations to determine the relative importance of different modes of non-covalent interactions in A2A-ligand binding. The interaction energies were calculated at the B2PLYP level of theory, in combination with Grimme’s D3BJ dispersion correction. It was found that these 5 residues (“key” residues) contribute significantly to ligand binding. The strength of π-π stacking interactions from F168 is the strongest. Residues M177, L249 and I274 offer some strong CH-π interactions. The strength of the hydrogen bond from N253 is comparable to that of CH-π interactions. These interactions from the “key” residues can be considered the major molecular determinant responsible for the molecular recognition of ligands in the A2A receptor. Our findings are in good agreement with site-directed mutagenesis data which showed that the mutation of these “key” residues to alanine leads to the total abolition of agonist or antagonist binding. In the second system, the molecular recognition in GTP-binding proteins, was studied. G proteins, including small monomeric GTPases (small G proteins) and heterotrimeric G proteins, are the family of proteins that act as molecular switches inside cells and play an essential role in cellular signal transduction. Thus, molecular recognition of GTP/GDP in G proteins is a subject of great importance for understanding the enzymatic mechanism and designing drugs targeting the GTP binding pocket.A large-scale data mining of Protein Data Bank was performed, which resulted in 302 GTP-binding protein complexes. To decipher specific interactions responsible for molecular recognition, the intermolecular interaction modes of hydrogen bond interaction, cation-π interaction, and π-π stacking interaction between guanine and its surrounding residues were systematically analyzed. A database of such interactions has been created.We discovered 6 different hydrogen bonding patterns in which the surrounding interacting residues prefer to interact with N2 and O6 of the guanine base via hydrogen bond interaction. This results in selectivity to the guanine base because these two atoms are absent in the adenine base. In 86.1 % (260 out of 302) of GTP-binding proteins, O6 is accepting an additional hydrogen bond from the main chain amino group of alanine and/or its succeeding residue from (T/G)(C/S)A sequence motif or any non-conserved residues. The N2, on the other hand, donates two hydrogens, N2H1, and N2H2, directly to the surrounding residue. In 90.1 % (272 out of 302) of complexes, N2H1 is being donated to surrounding residues, and in 21.9 % (66 out of 302) of the complexes, N2H2 is donated to the surrounding resides. High-level quantum chemical methods like B2PLYP and MP2 methods were applied to quantify the energetic contribution of each type of interaction. Overall, our analysis showed the significant contribution of cation-π interaction (between guanine and a positively charged residue) to the binding of guanine. In 86.4% of complexes (261 out of 302), at least one cation-π interaction does exist between the guanine base and positively charged side chains. The presence of π-π stacking interactions is less than cation-π interactions. In 54.3 % of complexes (164 out of 302), π-π stacking interactions exist between the guanine base and aromatic side chains. It was observed that the strength of the intermolecular cation-π interaction depends upon the combination of three factors, i.e., distance, the extent of overlap between the side chain of positively charged residue and the guanine ring, and multiple modes of interactions (cation-π and hydrogen bond). The strength of the intermolecular π-π stacking interaction is dependent upon a combination of three factors, i.e., distance, the angle between the aromatic rings, and the extent of π-π stacking.In order to get a detailed picture of molecular recognition in a whole GTP-binding protein, a complete analysis of the entire binding pocket was carried out on a representative GTP-binding protein, the p21-ras protein. The resulting interaction energies showed the most significant contributor to guanine binding energy is cation-π interaction which contributes almost half of the guanine binding energy, -18.2 kcal/mol (49.3 %). The remaining contribution is from hydrogen bond, -15.7 kcal/mol (42.5%), and π-π stacking interactions (8.2 %). The frequent presence of cation-π interaction (86.4% of complexes) and its significant energetic contribution to the binding of guanine suggest that cation-π interaction is one of the main molecular determinants for guanine binding in GTP-binding proteins besides hydrogen bonding.The final system studied was a protein-protein complex consisting of the spike protein of novel coronavirus (SARS-CoV-2) and the human ACE2 receptor. The development of therapies that target spike protein and the prevention of viral entry depend heavily on our understanding of molecular recognition between the spike protein and ACE2 or between spike and neutralizing antibodies. Binding of the spike protein with ACE2 initiates the viral entry, and the receptor binding domain (RBD) of the spike protein is directly involved. We identified all the residues at the binding interface of RBD, and their modes of interactions to unravel the molecular determinants responsible for recognition of spike protein. 13 hydrogen bonds, 2 salt bridges, 14 CH-π, 3 π-π, 1 cation-π, 1 NH-π and 1 S-π interaction are found at the binding interface. Our high-level quantum chemical analysis revealed three hot spot areas consisting of nine strong-binding residues, and one isolated strong-binding residue. Strong-binding residues K417, Y449, L455, F456, F486, N487, Y489, T500, N501, and Y505 possess interaction energies of greater than -4.0 kcal/mol. Remarkably, some of these strong-binding residues are among the epitope residues of several potent neutralizing antibodies, indicating that these residues also play a crucial role in strong antibody binding. The MM/PBSA method of binding free energy calculation is close to the experimental value, and it can be an efficient method to calculate the binding free energy of new variants or antibodies for which the complex structure is available from experiment or homology modeling.The impact of the N501Y mutation on the binding affinity between the spike protein and the ACE2 was investigated in another protein-protein system. The N501Y mutation occurs in several highly contagious alpha-, beta-, gamma-, and omicron-strains of SARS-CoV-2, and has been linked to enhanced transmission and immune evasion. There is an urgent need to understand this mutation at the molecular level so that effective vaccines and antibody therapies against quickly evolving variants can be designed. We performed a comparative examination of the molecular recognition of the wild-type and mutant spike proteins by the human cell surface receptor ACE2. It was found that the N501Y mutation results in a gain in new interaction modes and a gain in interaction energy by almost -2.0 kcal/mol. In addition, molecular dynamics simulation showed that Y501 is surrounded by a network of water molecules, facilitating the formation of a water mediated hydrogen bond with the hydroxyl group of Y501. It is concluded that the increased binding affinity in the N501Y mutants may be partially responsible for the greater transmissibility of the VOCs with the N501Y mutation.

Drugs with desired kinetic properties have better efficacy. Non-covalent small molecule drugs can bind/unbind from their targets, which normally are macro biomolecules. Experimental methods can measure kinetics parameters at ensemble level, but they are unable to track the exact drug binding/unbinding pathways or explain whether drugs show different kinetic behaviors from different pathways. Investigation of ligand binding/unbinding pathways can deepen our understanding of ligand-protein molecular recognition. In this work, possible pathways of ligands-protein were sampled and post-analysis were conducted to investigate ligand binding kinetics (residence time), free energy landscape of ligand unbinding, and key factors that affect binding kinetics. We are the first utilizing unbiased molecular dynamics to sample a pair of ligands, ritonavir and xk263, unbinding from HIV protease comprehensively, and classify the unbinding pathways based on the contacts between ligand-protein regions during dissociation. I identify key residues that form hydrogen bond with ligands that results in a meta-stable state for ligand-protein complex. Four distal mutation sites were observed forming interactions with ligands during unbinding, which explains why distal mutations in HIV protease affect drug binding affinity. Molecular mechanics Poisson–Boltzmann surface area (MM/PBSA) was performed to calculate free energy landscape during ligands dissociation. HIV protease backbone conformations during ligands binding/unbinding were compared based on root-mean-square deviation (RMSD). High similarity of protein conformation during ritonavir binding/unbinding suggests that ritonavir follows conformation-selection mechanism during dissociation. In the contrast, low similarity of protein conformation during xk263 binding/unbinding suggests that xk263 follows induced-fit mechanism during dissociation.Reconstruction of real free energy landscape of ritonavir-HIV protease dissociation was performed using Binding Kinetic Toolkit (BKiT) package. First two principal components (PC) of alpha carbon of protein along with heavy atoms in ritonavir were used as reaction coordinates to guide dissociation path on PC space. Energy barriers on free energy landscape under different unbinding pathways were explained by the molecular recognition of ritonavir-HIV protease. Furthermore, free energy landscape using ligand RMSD were computed, giving a much more accurate binding free energy and residence time approximation but in the lack of detailed ligand-protein interactions.

Molecular recognition is a fundamental part of chemical processes, especially those relevant to biology. It refers to the process by which two molecules diffuse and eventually bind with one another to form a complex. This can be broken down into to broad aspects: kinetics and thermodynamics. Kinetics refers to the motion of molecules and the rates of their reactions with each other. Thermodynamics refers to the transfers of energy that drive the reaction when molecules bind together. The work in this dissertation uses computational methods to study both aspects of molecular recognition in a range of systems, and it includes the application of existing methods and development of new tools for simulation and analysis.A strong focus is given to protein-ligand systems, in which the ligand is an inhibitory drug designed to shut down function of its target protein. The concept of developing drugs (inhibitors) that bind with and disrupt the activity of their targets is the basis for much of modern medicine, and has had incredible success. The development of more effective inhibitors is a constant challenge. The physical and chemical principles that predict an inhibitor’s effectiveness have a complex interplay, and an understanding of these principles is challenging and highly sought after. Two projects described here use molecular dynamics (MD) simulations to elucidate these principles through better understanding inhibitor binding thermodynamics. These techniques are applied to a host of inhibitors for the carcinogenic CDK8 protein and to inhibitors of a protease (PLpro) of the recent SARS-CoV2 virus. Novel inhibitors of PLpro are also developed and validated. The other work described herein studies molecular binding kinetics in both natural and engineered systems. Brownian dynamics simulation software is described which has been developed by the author and other group members. Its goal is to provide a robust tool with which researchers can study molecular recognition and association in a range of systems under varied conditions. This program, called GeomBD3, has been applied to study association kinetics and mechanisms in enzyme bioconjugates, protein-ligand systems, and nucleic acid biosensors. These studies are included in this dissertation, and we thus demonstrate that Brownian dynamics simulations can aid in rational bio/chemical engineering design efforts and supplement experimental analysis.

The precision medicine initiative was launched by the Obama administration in 2015. Major technological challenges exist, however, preventing the clinical realization of precision medicine. This dissertation specifically addresses two of these challenges, namely the need to engineer materials with specific affinity for protein analytes, and the need for customizable nanocarriers that target individual patients’ ailments. To address the first challenge, I developed multifunctional, molecularly imprinted hydrogels for protein recognition. Molecular imprinting within anionic microparticles increased their capacity to load high isoelectric point proteins, but failed to impart single-protein specificity. To further understand the impact of material composition on protein adsorption, I constructed a library hydrogel microparticles comprised of hydrophilic, anionic, and hydrophobic components. Analysis of model protein adsorption to each of these formulations elucidated the interaction between Coulombic attraction, hydrophobic contact, and size exclusion in dynamic protein-hydrogel interfaces. This analysis was the first to characterize an affinity-permeability tradeoff, where moieties that increase solute affinity simultaneously decrease the maximum solute partitioning. I also uncovered the critical role of solvent-accessible arginine content in the affinity of high isoelectric point proteins for amphiphilic, anionic copolymers. In the second portion of the dissertation, I describe the fabrication of tunable nanoscale hydrogels by inverse emulsion polymerization. A base poly(methacrylic acid-co-acrylamide) formulation was modified by altering the composition of the monomer feed or completing bioconjugation reactions off of acid moieties. Modified polymer conjugates included poly(methacrylic acid-co-acrylamide) with pendant aromatic moieties, cationic groups, fluorophores, peptide oligomers, and proteins. Modified nanogels’ physicochemical properties determined their cytocompatibility, environmental responsiveness, and solute affinity. They facilitated combination therapies (i.e. chemotherapy delivery and photothermal therapy) for precision medicine, and exhibited predictable biodegradation kinetics. As proof-of-concept, I concluded with a tailored peptide-modified nanogel that promoted carrier uptake by colorectal tumor cells. In the future, we will design formulations that bind, sequester, or deliver biomolecules reliably by applying known relationships governing protein-biomaterial interactions in physiological fluids. Furthermore, diverse and modular nanomaterial platforms will enable the rapid and rational construction of patient or pathology-specific drug delivery devices.

Self-assembled metal organic cages are supramolecular compounds with a unique internal cavity capable of creating an alternate nanophase inside of a reaction vessel (e.g. creating a hydrophilic environment inside of an organic solvent like acetonitrile). By exploiting this internal cavity, the complexes may be used for a wide array of applications in molecular recognition, small molecule transport, catalysis and even as molecular machines. Often the function of these cages becomes limited due to the flat aromatic ligands used in the assembly process. To improve the function of these cages, new methods for installing reactive groups on the interior of the cage must be explored.This work focuses on new methods of introducing function to self-assembled cages through modifications of the ligand and cage with the goal of creating new biomimetic catalysts. These modifications can be used introduce new reactivity. By introducing native fluorescence to the chelating motif, a new class of metal selective fluorophores was discovered. While adding reactivity can be more challenging, it can be simplified to three classes: reactions that modify a preformed cage, reactions activated by the cage, and reactions hosted within the cage. Cages can be modified after their assembly through the introduction of reactive components. Internalized doubly benzylic methylene subunits can be altered post-assembly via radical promoted oxidation, with the resulting products exhibiting an impressive level of stereocontrol. The use of a strained complex enables transamination reactions to be performed at ambient temperatures giving rise to rare intermediates along the assembly pathway. A large Fe2+-iminopyridine cage bearing twelve internal carboxylic acid groups was synthesized which can affect 1000-fold rate enhancements for several reactive processes at high turnover. Further, internalization of the acid functional groups allows a tandem cage-to-cage reaction and can vary the mechanism reactions, which is unattainable by similar “free” acid catalysts. This combined process of molecular recognition and substrate activation is reminiscent of that found in the active site of enzymes. Finally, a unfunctionalized tetrahedral cage exhibiting high binding affinity, up to 200,000 M-1, was synthesized. This high binding can be used to encapsulate a catalyst to accelerate substitution reactions. This reactivity is akin to the symbiotic relationship of an apoenzyme and cofactor and is unprecedented in synthetic hosts. This is because the selective simultaneous encapsulation of multiple different guests is extremely difficult and exhibits high entropic penalties. The results of this process not only affect the reaction rate but also biased selectivity of substrates in forming quaternary complexes enabling a controlled change in the reaction mechanism.

The study of synthetic model systems and biological counterparts has developed an extraordinary symbiosis, helping to decipher important chemical phenomena observed in nature. This Thesis is dedicated to the understanding of molecular recognition processes of neutral achiral and chiral small molecules by enantiopure receptors. Despite the progress in the design and construction of enantiopure receptors, examples of optically pure systems that effectively differentiate chiral neutral small molecules are still rare. The general notion prevails that strong directional interactions between the host and the guest are required. In order to question this idea, we constructed enantiopure alleno-acetylenic cage (AAC) receptors that bind molecules purely based on dispersion interactions largely in the absence of directional interactions. Subsequently, we extended our molecular recognition studies to halogen-bonding and hydrogen-bonding interactions. In the second chapter, we describe the synthesis and properties of enantiopure alleno-acetylenic cage (AAC) receptors. AACs are constructed from a methylene-bridged resorcin[4]arene core to which four homochiral alleno-acetylenes with OH termini are attached to, giving access to both (P)4- and (M)4 configured AACs. Detailed analysis of the structure-property relationship of the receptor allowed to identify important conformational features of the receptor, enabling quantification of guest uptake and release: the receptors undergoes solvent-dependent binary conformational switching accompanied by strong changes in the associated electronic circular dichroism (ECD) spectra with ΔΔɛ = 882 M–1 cm–1 at �������� = 304 nm, allowing for a sensitive spectroscopic readout of the conformational changes. In the closed cage conformation, the OH-termini of the alleno-acetylenic arms form a cyclic four-fold hydrogen-bonding array, creating a highly confined cavity. The directional nature of the H-bonding array – clockwise for (P)4-configured AACs and counter-clockwise for (M)4-configured AACs – was identified to contribute to the unprecedentedly large change in chiroptical properties of the assembly. A general method to obtain single crystals of the solid-state inclusion complexes was developed and relies on the guest-induced switching of the receptor from its open state (in CH3CN/H2O 9:1) to the closed state upon encapsulation of guest molecules. The combination of a highly shape-persistent, confined chiral cavity, capable of guest encapsulation, together with the spectroscopic and crystallographic readout for cage inclusion, made the AAC receptors ideal model system to study chiral recognition. We first investigated the molecular recognition of achiral and chiral cyclic alkanes, where complexation is purely based on non-directional dispersion interactions. X-ray co-crystal structures revealed a size adaptability of the receptor towards the guest, thereby optimizing the packing coefficient of the ensemble. At the optimal packing coefficient of ∼55%, the enantiopure receptor showed complete selectivity towards (±)-trans-1,2-dimethylcyclohexane, where the (P)4-configured host solely bound the (R,R)-configured guest, and the (M)4-configured receptor exclusively bound the (S,S)-configured guest. X-ray co-crystal structures of the host-bound guests revealed the exclusive complexation of their higher-energy diaxial conformation, with the diaxial dihedral angle deviating strongly from the commonly accepted value of 180° down to 146°. Subsequent theoretical investigations demonstrated negligible influence of the host on the guest structures. We validated the utility of the host for the structural elucidation of the (di)axial conformations of cyclohexane derivatives by expanding the series of guest molecules to monohalo- and (±)-trans-1,2 dihalocyclohexanes. The molecular structures of the host–guest complexes, obtained through single-crystal X-ray diffraction, showed the guests exclusively bound in their axial and diaxial chair conformation, with dihedral angles ϑa,a (X-C(1)-C(2)-H/X) deviating substantially from 180°. Increasing deviation from this angle was observed for the monohalocyclohexanes (up to 25°) to (±)-trans-1,2-dihalocyclohexanes (up to 33°). Substantial bond-length and bond-angle alteration in the carbon scaffold was assumed to reduce the strain caused by the 1,3-diaxial interactions of the guests in their diaxial conformation. Solution complexation studies supported the exclusive complexation of the guests in their (di)axial chair conformation, where slow host–guest exchange allowed for full characterization of the guest in the interior of the host. Theoretical analysis of the isolated guest molecules showed close agreement of the complexed and the isolated guest structures, validating the utility of the AACs to capture single conformers of derivatives of cyclohexane for their structural elucidation. X-ray co-crystal structures of the host-guest complexes further revealed a yet hardly studied halogen-bonding contact: the C–X⋯⫴ contact. Theoretical studies on the C–X⋯⫴ interaction substantiated its halogen-bonding character. Solution binding constants, along with the theoretical calculations on the conformational energies (A-values) of the guests, indicated a contribution of the C–Br⋯⫴ halogen-bonding contact of ΔΔG F→Br = – 0.9 kcal mol–1. The C–X⋯⫴ contact appeared to majorly influence the enantioselectivity of the enantiopure receptor towards the chiral guests, with increased enantioselectivity with increased halogen-bonding strength (Cl < Br). The overall enantioselectivity towards (±)-trans-1,2-dihalocyclohexanes was lower compared to (±) trans-1,2-dimethylcyclohexanes (complete enantioselectivity). This finding was counter-intuitive considering the stronger and directional nature of halogen-bonding contacts compared to the non directional, purely dispersion interactions, of (±)-trans-1,2-dimethylcyclohexanes with the host. It is in stark contrast to established concepts for enantioselective complexation of optically pure receptors with chiral guests, where more directional interactions were considered to enhance selectivity. We explained this observation with the much higher polarizability of chlorine and bromine compared to the methyl substituents. Inspired by a crystal structure of the AAC receptor encapsulating one water molecule and two acetonitrile molecules, we expanded our series of guest molecules to cyclic and acyclic alcohols. The alcohol series formed strong directional interactions between the alcohol groups of the guest and the hydrogen-bonding array of the host. Generally, the introduction of an alcohol group increased the binding affinities of the guest to the receptors in solution by ∼3–4 kcal mol–1, resulting in kinetically stable host–guest complexes on the NMR time scale. Solution studies, along with structural information obtained from X-ray co-crystal structures, enabled the conformational analysis of the host-bound guests. Noteworthy was the substantial increase in binding affinity from cycloheptane to endo-tropine with a difference in binding affinitiy of ΔΔG 293 K = –6.1 kcal mol–1 (Ka = 7.0·106 M–1 in n-octane at 293 K), allowing to detect endo-tropine with AACs in the part per billion regime. The directional hydrogen-bonding interactions of the guest to the receptors generated various hydrogen bonding motifs (4-fold to 5-fold and 6-fold), which were strongly dependent on the alcohol guest encapsulated in the interior of the host. The host–guest-complex appeared to retain some directionality of the hydrogen-bonding array, despite the disruptive nature of the directional hydrogen-bonding interactions of the guest with the host. In a collaboration with Dr. Fischer and Prof. Carreira (ETHZ), supported by theoretical studies by T. Husch and Prof. Reiher (ETHZ), we studied the enantioselective binding of various acyclic alkyl and alkyl halide alcohols, undergoing dispersion and halogen-bonding interactions. The formation of diastereoisomeric complexes of the enantiopure hosts with the chiral guests enabled us to assess the enantioselectivity of the receptors towards the guests in solution and in the solid state. X-ray co-crystal structures gave insight into the conformation of the guests complexed to the interior of the host. In the following chapter, we describe the modular synthesis of enantiopure alleno-acetylenic cage receptors with increased surface polarity and solubility in aqueous medium. This new class of receptors revealed conformational switching from an open to a closed form upon guest complexation. The structural similarity of the hydrophobic cavity of the more polar AACs soluble in aqueous medium with the apolar AAC receptors, make them ideal to study the thermodynamic differences of enantioselective complexation in apolar and aqueous solvent systems. The last chapter gives a brief overview on the synthesis and chiroptical properties of covalently capped alleno-acetylenic cage receptors, accessed through intramolecular oxidative dimerization. The covalent AACs showed strong absorption properties towards circulary polarized light, with hardly any temperature dependencies. X-ray co-crystal structure of the covalent (P)4- and (M)4-configured AACs gave insights into the volume of the cavity for molecular recognition studies. Molecular recognition studies on the covalent receptor systems are ongoing.

The cell’s surface is complex and multicomponent in nature. It is the setting where a variety of crucial life processes take place, mediated by the interaction of cell surface receptors with specific molecules outside of the cell. Studying how particular interactions take place in this convoluted environment can be difficult in actual cell membranes. Modeling components of the cell’s surface can be beneficial in studying cellular processes, although mimicking these systems poses a substantial challenge. Synthetic receptors such as calix[n]arenes, cyclophanes, and cyclodextrins are capable of performing molecular recognition of ionic and hydrophobic guests in aqueous environments, however, these synthetic receptors are not useful for molecular recognition studies in membrane environments.This work studies the molecular recognition abilities of water-soluble deep cavitands in membrane environments. The binding behavior of cavitands in membrane-like envrionments was studied using NMR analysis. The differences in binding behavior were elucidated in lipid environments compared to free solution. Supported lipid bilayers have been previously employed to mimic membrane environments, and the binding of trimethylammonium-tagged compounds and cationic native proteins were monitored via surface plasmon resonance (SPR) spectroscopy. By varying the functionality at the cavitand’s rim, a broader range of interactions were explored. The binding of anionic native proteins was possible by positioning a positive charge at the rim of the cavitand and more complex interactions between host and guest were possible by tailoring guests that utilize dual mode binding. In order to imitate larger structures on cell surfaces such as glycopolymers, cavitand-mediated atom transfer radical polymerization (ATRP) was performed in order to create a functional polymer on the SLB surface. Utilizing aminoethyl methacrylate, an amine polymer was formed that can be further derivatized by reacting fluorescent compounds for visualization, or epitopes for the adhesion of larger biomolecules such as avidin. The versatility of the amine-functionalized polymer was demonstrated with its ability to bind large structures such as cells at the interface of SLBs. The diverse interactions of the variably functionalized cavitands also led to their use in indicator displacement assays (IDA) for sensing a variety of analytes ranging from post translational modifications to metals and steroids.

Due to the high cost and environmental instability of antibodies, there is precedent for developing synthetic molecular recognition agents for use in diagnostic sensors. Molecular imprinting was first investigated as a method for improving selectivity of crosslinked polymer hydrogels. However, the lysozyme-imprinted polymers exhibited high cross-reactivity and did not afford the polymer with improved selectivity. Instead, the observed cross-reactivity prompted the investigation of charge-containing, non-imprinted polymers as differential protein receptors. To this end, a set nanogels were synthesized by copolymerizing N-isopropylacrylamide with methacrylic acid, followed by a post-synthesis modification. Specifically, a carbodiimide coupling scheme was used to introduce sulfate, guanidinium, secondary amine, or primary amine groups. As expected, modification of the ionizable groups in the network changed the physicochemical and protein binding properties of the nanogels. For high affinity protein-polymer interactions, turbidity of the nanogel solution increased, while for low affinity interactions minimal change in turbidity was observed. Thus, relative turbidity was used as input for multivariate analysis. Turbidimetric assays were performed in two buffers of different pH (i.e., 7.4 and 5.5), but comparable ionic strength, in order to improve differentiation. Using both buffers, it was possible to achieve 100% classification accuracy of eleven model protein biomarkers with as few as two of the nanogel receptors. Subsequently, these polymeric receptors were synthesized on the surface of silica-gold nanoshells to develop a diagnostic biosensor based on localized surface plasmon resonance (LSPR). In particular, the sensitivity of LSPR to changes in local refractive index was exploited to enable detection of three biomarkers (lysozyme, lactoferrin, and IgG) of Sjögren’s syndrome, an autoimmune disease that primarily affects the exocrine system. Overall, the polymer-gold nanomaterial composites developed in this dissertation provide a promising platform for developing affordable and environmentally robust diagnostic biosensors.

Herein, work on guanidinium-based anion receptors and their anion separation properties are described. First, a novel receptor based on the N,N’-bis(2-pyridyl)guanidinium motif is rationally designed, synthesized, and characterized. In the solid state, X-ray crystallography shows that is has a strong conformational preference for the α,α form of the molecule. This ligand has association constants of 3.78 ± 0.12 and 2.10 ± 0.23 respectively for log K1 and log K2. A lipophilic form of this ligand was synthesized for extraction studies, where it performed better than commercially available Aliquat 336 for the extraction of sulfate into 1,2-DCE from water. Next, a series of bis(imino)guanidinium ligands are created that are capable of forming highly insoluble salts with many oxoanions, the most insoluble being sulfate. This leads to novel methods of separation of highly charged oxoanion species by precipitation followed by simple filtration, and its use was demonstrated on natural seawater. This methodology was adapted to the sequestration of CO2 from ambient air, leading to an effective method for the crystallization of CO2 as its carbonate salt from water as well as its low temperature release. Finally, future work towards the creation of iminoguanidinium-based oxoanion receptors is described.

Molecular recognition is fundamentally important in biological chemistry. Nowadays, with the rapid development of computational technology and algorithm, molecular modeling has become a powerful tool in studying molecular recognition, such as exploring molecular interactions and understanding biological dynamic processes, making significant contributions to modern biology and drug discovery. The state-of-the-art techniques of computational chemistry and molecular modeling can be applied to study a wide range of chemical and biological systems of interest. This enables us to study structural details at the atomic level and obtain chemical/biological information which is not available by experimental measurements. This dissertation project focused on modeling the recognition mechanisms of biomolecules and their conjugated ligands. Multiple computational techniques, such as molecular dynamics simulation, enhanced sampling methods and free energy calculation were applied. The model systems included signaling domains (BRCT domain), kinase (p38 kinase), enzyme system (TRPS) and small biomolecular system (cyclodextrin). The details of protein-ligand interactions, including both enthalpic and entropic contribution within protein domain-phosphopeptide systems were investigated, based on which new inhibitors were proposed. Several enhanced sampling methods like accelerated molecular dynamics simulation, pathway search guided by internal motions (PSIM) and umbrella sampling, were applied to explore the dissociation pathway of kinase-ligand systems and the motions of kinase during dissociation process were studied both thermodynamically and kinetically, protein conformational rearrangement was found to differentiate slow and fast unbinding inhibitors, casting light on high efficacy inhibitor design. Furthermore, using full structural molecular modeling, we explored how the position of a single proton can change the overall protein dynamics and further activate or inactivate enzyme catalysis, elucidating the catalytic mechanism of TRPS. In addition, we performed systematically evaluation to the performance of umbrella sampling, investigated the influence of subtle changes in the dissociation pathways and conformational sampling methods that provide the initial conformations, paving way for future improvement of umbrella sampling. This project studies the details of receptor-ligand interaction and provides a more complete picture of molecular recognition.

The overall picture of molecular recognition covers the thermodynamic and kinetic properties of molecular systems. Regarding thermodynamics, binding affinity or free energy, which can be decomposed into enthalpy and entropy, determines the strength of binding. Binding kinetics, the association and dissociation rate constants, describe the rates for molecular binding and unbinding. In this thesis, I carried out various molecular mechanics modeling tools to understand binding thermodynamics and kinetics of host-guest systems and protein-ligand systems. The work includes both novel method development and applications using existing tools. Here we computed the binding free energy of the surfactant-silver nano-plate complexes using the M2 method and to understand how a capping ligand helps grow shaped nanomaterials. Several factors were determined to be crucial, and our findings allow rational design of capping surfactants in addition to the magic compound citrate. In addition, a molecular dynamics (MD) and docking were applied to find the inhibitor binding site on protein TIM44 to assist cell biology studies. I examined the inhibitor selectivity of wildtype TIM44 and its mutants using energy calculations.The binding/unbinding processes are carefully investigated for β-cyclodextrin and its guests to thoroughly study the thermodynamics and kinetics of binding. I applied MD simulations, post-analyzed the MD trajectories and developed and implemented methods such as the cell method to compute binding enthalpy, solute entropy and solvent entropy. Notably, the latter two entropy components are known to be very challenging to obtain computationally. I also computed the kinetic rate constants and investigated the influence of force fields used. It is the first work that illustrates all entropy and enthalpy components of binding in great details.Realizing how crucial conformational sampling can be in answering fundamental scientific questions and valuable applications, a novel method to accelerate searching molecular conformations and ligand dissociation pathways was developed. The method uses a multi-layer internal coordinate and an internal PCA search algorithm to post-analyze a given MD trajectory and then guide motions of a molecular system. This algorithm is able to find possible dissociation pathways with high efficiency. Five examples are discussed to illustrate the functionality and capacity of this new method.

As chemists characterize molecular systems in greater detail, it becomes clear that some observables can only be properly studied at the macromolecular scale. However, elucidating the physical principles behind such phenomena as molecular recognition or chemical reactivity can be difficult when moving into the macromolecular regime. The objective of this work is to provide insights and predictions to complement experimental undertakings. The work is divided into two categories: 1) modeling molecular recognition through prediction of intermolecular interactions with highly accurate methods and 2) the modeling of chemical reactivity. The separation of N2 and CH4 is particularly pertinent for the natural gas industry, and improved materials for performing this separation would provide an enormous cost savings. This work is focused on the prediction of a new material capable of performing this separation. Through application of multiple tiers of quantum chemical methods, and comparison to similar known experimentally synthesized materials, a novel material is predicted to effect the separation of N2 and CH4 through selective binding of N2 to an open vanadium metal site. A particularly valuable tool for monitoring target delivery or guest encapsulation in macromolecular systems is an easily observed signal that indicates the status of a host-guest complex. Guest complexation can alter the observable properties of the host, including the spin crossover properties of a host macromolecule. Particular care was taken to correlate guest recognition to changes in paramagnetic NMR chemical shifts induced in the host system. The monitoring of subtle changes in a protein's environment is a challenging and complex problem; however, the observation of ligand complexation in biomolecular systems is extremely important in the design of new medicinal therapeutic drugs. This work aims to develop a quantum chemical method to assist experimental assignment of challenging 19F NMR spectra in proteins. The importance of accurately modeling the hydration environment is extremely critical for accurate comparison to experimental measurements. Selective detection of chemical impurities is an attractive capability to have for any chemical process. A key impurity in industrially synthesized explosive TNT is DNT. Given the high prevalence of DNT in TNT, detection of DNT through electrochemistry is a useful sensor for explosives. This work characterizes the mechanism of DNT electrochemical reduction A new material was found to rapidly catalyze the decomposition of extremely toxic chemical warfare agents. The macromolecular metal organic framework NU-1000 was demonstrated to be extremely effective in catalyzing the hydrolysis of phosphoester based chemical warfare agents. Predictive computations were performed on a nerve agent simulant DMNP, and toxic nerve agents GD (Soman) and VX agents, uncovering the key role that the metal nodes of NU-1000 play in activation of the phosphoester bonds for hydrolytic attack

The exploration of molecular recognition with artificial hosts has been centered on mimicking the nature and how natural receptors recognize specific guests. Charles Pedersen thus investigated cyclic polyethers, referred to as crown ethers, and their complexation of cations. Jean-Marie Lehn and Donald Cram further investigated on molecular recognition by using three-dimensional crown ethers and carcerand/hemicarcerand, respectively. The Badjic Group has developed gated molecular baskets and other novel hosts to study molecular encapsulation. The baskets control the rate of trafficking of guests depending on their size, shape, hydrophobicity, and electronic characteristics. In particular, The Badjic Group reported novel dual-cavity baskets, derived from heptiptycene. These hosts are unique in a way that they could express a positive homotropic allosterism. Furthermore, the functionalization of this type of cavitands, with two juxtaposed cavities, could lead to the development of catalysts, chemosensors, drug-delivery systems, or assist the formation of reactive molecules/intermediates. This thesis describes the functionalization of dual-cavity baskets, with polyethylene glycol groups and L-alanines. These functional groups are selected to enhance the solubility of the dual-cavity cavitand in aqueous media and also provide dynamic gates to control the encapsulation.

Organophosphorus (OP) nerve agents are a very toxic class of compounds for which there is a great need of effective therapeutics. OPs are phosphoryl-containing compounds typically used as pesticides, but they have been used as chemical warfare agents. Their toxicity is derived from the inhibition of acetylcholinesterase (AChE), an enzyme found in the human body that regulates levels of the neurotransmitter acetylcholine. OPs inhibit AChE by undergoing nucleophilic attack by the catalytic Ser203 residue. A water molecule is not sufficiently nucleophilic to attack the phosphoryl center to regenerate functional AChE, so addition of a stronger nucleophile, such as an oxime, is utilized for attack at the phosphoryl center and cleavage of the P–O(Ser203) bond to reactivate AChE. However, a second reaction occurs in the inhibition process, called "aging," in which the phosphoryl O-alkyl functionality is dealkylated, creating an anionic species in the active site. There is currently no treatment after aging has occurred, as oximes can no longer attack the phosphoryl center to reactivate AChE. This dissertation encompasses some of the work that has been done on two of the possible routes for treatment after OP exposure. The first route involves utilizing a supramolecular compound that is capable of binding and hydrolyzing the toxic OPs before inhibition of AChE can occur, while the second method attempts to realkylate the aged form of AChE, through use of a selective alkylating agent to replace the departed alkyl group of the OP with a new alkyl group. Gated molecular baskets, with a dynamic mode of action, are offered as models for examining the encapsulation and degradation of toxic OPs. In an effort to understand the binding of OPs to the molecular baskets, we developed a computational protocol for examining the dynamics of the hosts and host-guest interactions through molecular dynamics and docking calculations. We found that modifying the functionality of the baskets can have a profound impact on the ability to trap OP guests. Specifically, the computational results of functionalized molecular baskets correlate with experimental studies in that the size and electronic effects of both the baskets and the guests determine the degree of OP encapsulation, suggesting that these molecular baskets could be utilized for efficient and selective encapsulation, detection, and detoxification of toxic organophosphorus nerve agents.For the realkylation of OP-aged AChE, an efficient alkylating agent must be developed. One such alkylating agent, quinone methides (QMs), could possibly be utilized as they have previously been shown to have highly tunable reactivity and could be administered in an unreactive prodrug form. The naphthalene and quinoline scaffolds were selected for incorporation of the QM moiety and were subjected to a computational study. We found that these naphthyl and quinoline-based QM precursors displayed a variety of reaction energetics in the alkylation of model nucleophiles, with some of the proposed compounds exhibiting particularly encouraging docking results with human AChE. As such, we have selected possible lead compounds from the computational study to be synthesized and investigated experimentally for the goal of realkylating OP-aged AChE.

Ribonuclease P (RNase P) is a divalent metal ion-dependent endonuclease that catalyzes cleavage of the 5’ leader of precursor transfer RNA (pre-tRNA), an essential tRNA processing step in all domains of life. Bacterial RNase P is a potential antibacterial target because it is essential for cell survival and varies in composition from its eukaryotic counterparts. Bacterial RNase P contains a catalytic RNA (P RNA) with one protein subunit while eukaryotic nuclear RNase P has multiple protein subunits with a catalytic RNA core and the human mitochondrial RNase P (mtRNase P) consists solely of protein subunits. In this dissertation research, first a novel fluorescence polarization (FP) assay was developed to facilitate rapid and real-time measurements of RNase P activity. This new FP assay was optimized for high-throughput screening to search for new inhibitors of bacterial RNase P in small molecule (2,880 compounds) and natural product extracts (22,720 samples) libraries. A new RNase P inhibitor was identified from the screen. Second, to test the biochemical role and metal ion binding function of a carbonyl oxygen (oxygen-4, O4) in the universally conserved bulged uridine (U51) of P RNA, RNase P with single atom modifications of 4-thiouridine, 4-deoxyuridine, 3-methyluridine and an abasic site were prepared and analyzed. Binding data demonstrate that the O4 of U51 enhances pre-tRNA affinity in a divalent metal-ion dependent fashion. In addition, kinetic data suggest that U51 enhances pre-tRNA cleavage by interacting with a magnesium ion stabilizing an active enzyme-substrate conformation. Third, a library of human mitochondiral pre-tRNAs and high purity human mtRNase P was prepared for studying the substrate specificity of human mtRNase P. Single-turnover cleavage data demonstrate that the MRPP3 subunit of human mtRNase P is catalytically active alone in vitro and that the MRPP1∙MRPP2 subcomplex increases the cleavage rate and cleavage site fidelity. Overall, the work presented in this dissertation has provided a new real-time and high-throughput methodology to assay RNase P activity in vitro, enhanced our understanding of how bacterial RNase P recognizes inhibitors and metal ions, and laid the foundation for elucidating substrate recognition by the newly identified protein-only human mtRNase P.

Based on its unique elemental properties, fluorine has emerged as an important design element in synthetic pharmaceuticals, but also a difficult substituent to incorporate into small molecules using synthetic chemistry. Synthetic biology approaches represent an attractive alternative that could allow us to harness the catalytic power and specificity of enzymes for fluorine incorporation into complex small molecules. However, fluorinated natural products are rare and derive from a single pathway for biosynthesis of fluoroacetate, an antimetabolite poison. The fluoroacetate-producing bacterium Streptomyces cattleya has evolved a fluoroacetyl-CoA thioesterase (FlK) that is involved in fluoroacetate resistance. Using FlK as a model system, we have focused on understanding enzymatic fluorine selectivity with the goal of defining principles for the design of fluorine-specific enzymes that can be applied in the development of new methods for preparation of fluorinated molecules. We have demonstrated that FlK exhibits a remarkable 106-fold selectivity for hydrolysis of fluoroacetyl-CoA compared to acetyl-CoA, an abundant central metabolite and cellular competitor. This selectivity is based on a decrease in KM (102) and an increase in kcat (104) for the fluorinated substrate. Through x-ray crystallographic studies, we have identified a unique hydrophobic `lid' in our crystal structure of FlK that shields the active site from water. Mutation of a key lid residue (Phe 36) led to a loss of fluorine-based binding specificity, which was correlated with the appearance of additional ordered water molecules at the active site in the mutant crystal structure. A crystal structure of the FlK product complex revealed that Phe 36 is dynamic, undergoing a conformational change when the active site is occupied with product. By studying the binding of a series of non-hydrolyzable substrate analogs, we have shown that the lid confers the fluorinated substrate with an entropic binding advantage that is lost in lid mutants, which may be related to water release from the `polar hydrophobic' C-F unit. Kinetic studies revealed that Phe 36 controls the substrate off rate, providing the fluorinated substrate with a kinetic as well as a thermodynamic advantage. We have also shown that catalytic specificity in FlK is based on utilization of an unusual reaction pathway initiated by deprotonation at the α-carbon for hydrolysis of fluoroacetyl-CoA but not acetyl-CoA. The enolate that is formed can then breakdown through a putative ketene intermediate to give the same product that would be produced by a canonical hydrolysis mechanism. FlK therefore represents a rare example in which the existence of two reaction pathways in the same active site controls substrate selectivity rather than product outcome. Although catalytic fluorine discrimination occurs mainly in one step of the reaction mechanism, we have demonstrated that fluorine is specifically recognized by the enzyme in both the acylation and deacylation steps of the reaction mechanism. These studies have defined the basis of fluorine selectivity in a naturally occurring enzyme-substrate pair, with implications for drug design and development of fluorine-selective biocatalysts.

The studies discussed in this thesis unify experimental and theoretical techniques, both established and novel, in investigating the problem of how a protein that binds specific sites on DNA translocates to, recognizes, and stably binds to its target site or sites. The thesis is organized into two parts. Part I outlines the history of the problem and the theory and experiments that have addressed the problem and presents an apparent incompatibility between efficient search and stable, specific binding. To address this problem, we elaborate a model of protein-DNA interaction in which the protein may bind DNA in either a search (S) mode or a recognition (R) mode. The former is characterized by zero or weak sequence-dependence in the binding energy, while the latter is highly sequence-dependent. The protein undergoes a random walk along the DNA in the S mode, and if it encounters its target site, must undergo a conformational transition into the R mode. The model resolves the apparent paradox, and accounts for the observed speed, specificity, and stability in protein-DNA interactions. The model shows internal agreement as regards theoretical and simulated results, as well as external agreement with experimental measurements. Part II reports on research that has tested the applicability of the two-mode model to the tumor suppressor transcription factor p53. It describes in greater depth the experimental techniques and findings up to the present work, and introduces the techniques and biological system used in our research. We employ single-molecule optical microscopy in two projects to study the diffusional kinetics of p53 on DNA. The first project measures the diffusion coefficient of p53 and determines that the protein satisfies a number of requirements for the validity of the two-mode model and for efficient target localization. The second project examines the sequence-dependence in p53's sliding kinetics, and explicitly models the energy landscape it experiences on DNA and relates features of the landscape to observed local variation in diffusion coefficient. The thesis closes with proposed extensions and complements to the projects, and a discussion of the implications of our work and its relation to recent developments in the field.

Over the past decades, significant efforts have been devoted to explore the use of various nanoparticle-based systems in the field of nanomedicine, including molecular imaging and therapy. Supramolecular synthetic approaches have attracted lots of attention due to their flexibility, convenience, and modularity for producing nanoparticles. In this dissertation, the developmental story of our size-controllable supramolecular nanoparticles (SNPs) will be discussed, as well as their use in specific biomedical applications. To achieve the self-assembly of SNPs, the well-characterized molecular recognition system (i.e., cyclodextrin/adamantane recognition) was employed. The resulting SNPs, which were assembled from three molecular building blocks, possess incredible stability in various physiological conditions, reversible size-controllability and dynamic disassembly that were exploited for various in vitro and in vivo applications. An advantage of using the supramolecular approach is that it enables the convenient incorporation of functional ligands onto SNP surface that confers functionality (e.g., targeting, cell penetration) to SNPs. We utilized SNPs for molecular imaging such as magnetic resonance imaging (MRI) and positron emission tomography (PET) by introducing reporter systems (i.e., radio-isotopes, MR contrast agents, and fluorophores) into SNPs. On the other hand, the incorporation of various payloads, including drugs, genes and proteins, into SNPs showed improved delivery performance and enhanced therapeutic efficacy for these therapeutic agents. Leveraging the powers of (i) a combinatorial synthetic approach based on supramolecular assembly and (ii) a digital microreactor, a rapid developmental pathway was developed that is capable of screening SNP candidates for the ideal structural and functional properties that deliver optimal performance. Moreover, SNP-based theranostic delivery systems that combine reporter systems and therapeutic payloads into a single SNP for both diagnosis and therapy were generated. The results show that this type of theranostic SNPs may have a great contribution in the optimization of therapeutic efficacy for individual patients in clinical translation in the near future. It is anticipated that our supramolecular synthetic approach could be adopted to assemble various SNP-based delivery agents for molecular diagnostics and therapeutics that pave the way toward personalized medicine.

Non-covalent interactions including nucleobase hydrogen bonding and phosphonium/ammonium ionic aggregation were studied in block and random polymers synthesized using controlled radical polymerization techniques such as nitroxide mediated polymerization (NMP) and reversible addition-fragmentation chain transfer polymerization (RAFT). Non-covalent interactions were expected to increase the effective molecular weight of the polymeric precursors through intermolecular associations and to induce microphase separation. The influence of non-covalent association on the structure/property relationships of these materials were studied in terms of physical properties (tensile, DMA, rheology) as well as morphological studies (AFM, SAXS). Ionic interactions, which possess stronger interaction energies than hydrogen bonds (~150 kJ/mol) were studied in the context of phosphonium-containing acrylate triblock (ABA) copolymers and random copolymers. Phosphonium-containing ionic liquid monomers with different alkyl substituent lengths and counterions enabled an investigation of the effects of ionic aggregation of phosphonium cations on the polymer physical properties. The polymerization of styrenic phosphonium-containing ionic liquid monomers using a difunctional alkoxyamine initiator, DEPN2, afforded an ABA triblock copolymer with an n-butyl acrylate soft center block and symmetric phosphonium-containing external reinforcing blocks. Small-angle X-ray scattering (SAXS) and transmission electron microscopy (TEM) of triblock copolymers revealed pronounced microphase separation at the nanoscale. Phosphonium aggregation governed block copolymer flow activation energies. In random copolymers, the phosphonium cations only weakly aggregated, which strongly depended on the length of alkyl substituents and the type of counterions. Acrylate random copolymers consisting of quaternary ammonium functionalities were synthesized using reversible addition-fragmentation chain transfer polymerization (RAFT). The obtained copolymers possessed controlled compositions and narrow molecular weight distributions with molecular weights ranging from Mn =50,000 to 170,000 g/mol. DMA evidenced the weak aggregation of ammonium cations in the solid state. Additionally, this ionomer was salt-responsive in NaCl aqueous solutions. Hydrogen bonding, a dynamic interaction with intermediate enthalpies (10-40 kJ/mol) was introduced through complementary heterocyclic DNA nucleobases such as adenine, thymine and uracil. Our investigations in this field have focused on the use of DNA nucleobase pair interactions to control polymer self-assembly and rheological behavior. Novel acrylic adenine- and thymine-containing monomers were synthesized from aza-Michael addition reaction. The long alkyl spacers between nucleobase and polymer backbone afforded structural flexibility in self-assembly process. Adenine-containing polyacrylates exhibited unique morphologies due to adenine-adenine π-π interactions. The complementary hydrogen bonding of adenine and thymine resulted in disruption of adenine-adenine π-π interactions, leading to lower plateau modulus and lower softening temperatures. Moreover, hydrogen bonding interactions enabled the compatibilization of complementary hydrogen bonding guest molecules such as uracil phosphonium chloride.

This dissertation is composed of all of my efforts devoted into two research projects. The first project (chapter 2-5) includes five designed molecular recognition systems in aqueous solutions. The second project (chapter 6) focuses on using sugar-lipid conjugates to protect liposomes from serum protein binding, and to protect supported lipid bilayer from damage during dehydration and rehydration. At the beginning of the first project, we designed, synthesized and studied a novel monovalent hydrogen bonding phospholipid system, using cyanuric acid (CA) and melamine (M) as the recognition pair. Our study has confirmed that the selective heteromeric membrane apposition was due to the molecular recognition between CA and M groups on the membrane surface. However, the major limitation of this monovalent system is that: the requisite density of CA/M modules for recognition was quite high. Trying to find the minimal valency required for the detectable lipid-lipid binding, we designed, synthesized and studied our second CA/M phospholipid system: the trivalent system. Our studies have revealed that low membrane concentrations of the trivalent CA/M lipids or peptides retained the robust molecular recognition properties found with surfaces composed entirely of monovalent CA/M lipids, although membrane activation upon binding was diminished.The robust molecular recognition of CA/M at the lipid-water interface promotes our efforts to extend the application to other apolar-water interfaces. As the first step, we still focused on the lipid-water interface. Instead of using functionalized lipids, we switched to amphiphilic peptides, specifically magainin. By covalently attaching the CA/M recognition motifs to magainin peptides, we found that these new peptide conjugates bound lipid membrane in a cooperative manner, and also their membrane activity was greatly enhanced. As the second step, we extended our interest further to protein-protein interfaces. We chose streptavidin, a well studied and widely used tetrameric protein. Synthesized biotin derivatives bearing the CA/M recognition motifs, and incorporated them onto streptavidin through biotin-streptavidin binding, we found that these functionalized streptavidin proteins could bind each other in a temperature-dependent and reversible manner.Curious about the mechanism and driving force of the CA-M interaction at various interfaces, we investigated the small molecule CA-M self-assembly in an aqueous solution at no interface. The parent CA-M self-assembly appears to be driven in large part by hydrophobic burial or “base-stacking” effects in water. The examination of aqueous-phase recognition between the trivalent derivatives, TCA and TM, suggests the possibility of designing useful recognition modules that function in water, as trivalent presentation of the heterocycles on a solubilizing molecular scaffold was found to enhance recognition and limit aggregate precipitation. For the second project, we switched our focus to preventing non-specific molecular interaction at the lipid-water interface, especially between serum proteins and liposomes. The non-specific serum protein-lipid binding is the main barrier for creating long-circulating lipid vesicle carrier. We designed and synthesized novel glycolipid polymers that provide vesicle stability in serum as well as unprecedented anhydrobiotic and cryoprotective properties. We anticipate that this glycolipid system will be useful for the stabilization of synthetic membranes for delivery and SLB bioanalytical devices.

Molecular recognition exploiting non-covalent interactions mediates the structure and function of many critical biological and synthetic molecules. There has thus been continuing and intense efforts in the design and synthesis of supramolecular systems with the capability of recognizing specific chemical species. Among various guest species, Prof. Sessler's group has been focused on the study of artificial anion receptors. Calix[4]pyrrole is a tetrapyrrolic macrocycle that is capable of binding anions via concerted and directional hydrogen bonding. Recently, a tetrathiafulvalene (TTF) functionalized calix[4]pyrrole (TTF-C4P) was synthesized and studied as a receptor for various guest species such as anions, electron deficient guest species, and C₆₀. This dissertation focuses on the recent discovery in supramolecular chemistry of TTF-calix[4]pyrrole derivatives. Chapter 1 provides a brief overview of the historical perspective, redox properties, and uses of TTF derivatives as functional building blocks for supramolecular assemblies, as well as previous findings involving the supramolecular chemistry of TTF-C4P. Chapter 2, as the major focus of this dissertation, describes ion mediated supramolecular and reversible electron transfer processes between TTF-C4P and bisimidazolium salts (BIQ²⁺2X⁻). We discovered that the electron transfer processes between these redox couples could be controlled reversibly by ion binding. Specifically, we found that anion binding to the TTF-C4P receptor promotes the forward ET processes. In contrast, cation complexation to the cavity of TTF-C4P causes the reverse ET processes. Such ion mediated ET processes play an essential role in biological ET systems including photosynthesis and respirations. These reversible ET processes were mapped out by spectroscopic (¹H-NMR, UV-Vis NIR titrations, and EPR analysis) and X-ray single crystallographic analyses of both the intermediate and products. Chapter 3 describes the synthesis of aromatic (thiophene and benzene) annulated TTF-calix[4]pyrroles as new and significantly improved receptors for poly-nitroexplosives. The resulting electronic modulations of the parent TTF-pyrrole structure result in significantly enhanced binding affinities for the corresponding TTF-C4Ps toward polynitro-explosives. This is reflected in a high level of positive homotropic allosterism. The degree of the cooperative effect was found to vary depending on the nature of both the receptors and guest species. The origin of the cooperative binding can be explained by conformational locking and an inductive effect of binding the first nitroaromatic guest.