The variation of a magnetic field in the vicinity of a zero-energy resonance allows highly vibrationally excited molecules (‘Feshbach molecules’) to be produced from an ultracold atomic gas. In this thesis, we study the dynamics of this process. We begin by studying the dissociation of Feshbach molecules, showing that in the limit of a sudden jump the shape of the spectrum of dissociated atoms can act as a probe of the zero-energy resonance. For some resonances, such jumps are within reach of current experiments. We also study the intermediate region between sudden jumps and asymptotically wide, linear ramps. It is shown from a precise derivation how the latter limit leads to a universal spectrum with a shape independent of the implementation of the two-body physics, provided that the near-resonant scattering properties are correctly modelled. We then turn to the dynamics of Feshbach molecule production from thermal and condensed gases. Our microscopic quantum dynamics approach includes the exact twobody evolution as an input to the many-body calculations. We show that in the long-time limit, and the Markov limit for the interactions, the non-Markovian Boltzmann equation (NMBE) we derive for the one-body density matrix reduces to the normal Boltzmann equation. In the limit of short times and small depletion of the atomic gas, the molecule production efficiency can be calculated by thermally averaging the two-body transition probability density. This thermal averaging technique is applied to studies of the formation of Feshbach molecules using a magnetic field modulation that is near-resonant with the molecular bound state energy. The continuum is shown to have a significant effect on both the dynamics and efficiency of this process. We examine the dependence of the molecule production efficiency on the duration, amplitude and frequency of the modulation, as well as the temperature and density of the gas. This method of producing molecules is effective for a wide range of bound state energies, but requires sufficient variation of the two-body energy levels with magnetic field. Lastly, we implement the NMBE for the case of a fast linear ramp across a Feshbach resonance. The solution of this equation is made feasible by including a large part of the required computation in the kernel, which is calculated in advance. The NMBE allows predictions of the molecule production efficiency which go beyond the thermal averaging technique by accounting for the depletion and rethermalisation of the continuum. In the limit of small depletions, the two approaches give the same results. As the depletion increases, the two approaches differ due to many-body effects limiting the maximum possible molecule production efficiency. We have observed this in our simulations by considering higher-density gases. We have therefore shown the suitability and practicability of this beyond mean-field approach for application to further problems in the production of Feshbach molecules from ultracold gases.

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Although dynamic interactions between plant cells and their environment require adhesion between the cell wall (CW) and the plasma membrane (PM), few plant adhesion molecules have been identified. Therefore, the seed coat mucilage secretory cells (MSCs) of Arabidopsis thaliana (which undergo developmentally regulated changes in adhesion) were developed into a novel model system to study PM-CW adhesion. Twenty-seven candidate genes were identified using data from publicly available and seed-specific microarrays. Mutant plants for these genes were screened for defects in adhesion via plasmolysis, and for changes in MSC morphology that may result from defective adhesion (Chapter 1). Two fasciclin-like arabinogalactan proteins were isolated in this screen. One of these, SOS5, was characterized in detail (Chapter 2). sos5 mutants are sensitive to hyperosmotic conditions and show defects in PM-CW adhesion and MSC mucilage structure. Interestingly, these phenotypes may be attributed to defects in adhesion or to defects in cell wall deposition.

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Small robust affinity proteins have shown great potential for use in therapy, in vivo diagnostics, and various biotechnological applications. However, the affinity proteins often need to be modified or functionalized to be successful in many of these applications. The use of chemical synthesis for the production of the proteins can allow for site-directed functionalization not achievable by recombinant routes, including incorporation of unnatural building blocks. This thesis focuses on chemical engineering of Affibody molecules and an albumin binding domain (ABD), which both are three-helix bundle proteins of 58 and 46 amino acids, respectively, possible to synthesize using solid phase peptide synthesis (SPPS). In the first project, an alternative synthetic route for Affibody molecules using a fragment condensation approach was investigated. This was achieved by using native chemical ligation (NCL) for the condensation reaction, yielding a native peptide bond at the site of ligation. The constant third helix of Affibody molecules enables a combinatorial approach for the preparation of a panel of different Affibody molecules, demonstrated by the synthesis of three different Affibody molecules using the same helix 3 (paper I). In the next two projects, an Affibody molecule targeting the amyloid-beta peptide, involved in Alzheimer’s disease, was engineered. Initially the N-terminus of the Affibody molecule was shortened resulting in a considerably higher synthetic yield and higher binding affinity to the target peptide (paper II). This improved variant of the Affibody molecule was then further engineered in the next project, where a fluorescently silent variant was developed and successfully used as a tool to lock the amyloid-beta peptide in a β-hairpin conformation during studies of copper binding using fluorescence spectroscopy (paper III). In the last two projects, synthetic variants of ABD, interesting for use as in vivo half-life extending partners to therapeutic proteins, were engineered. In the first project the possibility to covalently link a bioactive peptide, GLP-1, to the domain was investigated. This was achieved by site-specific thioether bridge-mediated cross-linking of the molecules via a polyethylene glycol (PEG)-based spacer. The conjugate showed retained high binding affinity to human serum albumin (HSA) and a biological activity comparable to a reference GLP-1 peptide (paper IV). In the last project, the possibility to increase the proteolytic stability of ABD through intramolecular cross-linking, to facilitate its use in e.g. oral drug delivery applications, was investigated. A tethered variant of ABD showed increased thermal stability and a considerably higher proteolytic stability towards pepsin, trypsin and chymotrypsin, three important proteases found in the gastrointestinal (GI) tract (paper V). Taken together, the work presented in this thesis illustrates the potential of using chemical synthesis approaches in protein engineering. / <p>QC 20140207</p>

This thesis offers a detailed socio-historical exploration of the practice of molecular modelling in chemistry and allied disciplines. This exploration leads to observations and conclusions that are of relevance to the sociology of science in general and to the issue of representation in scientific practice, in particular. After a general introduction in chapter one, chapter two offers a review of the science studies' literature relevant to the topic, namely the literature on representation and on experimental practices in the sciences. The lack of sources dealing with the issue of physical modelling is highlighted. Chapter three, through specific cases studies, presents the argument that the circulation of structural concepts and accounts in chemistry has been historically associated with the circulation not only of graphical representations, but also of physical models. Implications of this argument for a semiotically-centred view of scientific practice are discussed. Chapter 4 discusses the role of models as a research tool. Through various historical cases, it shows how physical models have been central in the process of defining, exploring and intervening on the domain of molecular-structures as accessed through laboratory manipulations. The work of molecular modelling is presented in particular as the experimental articulation of structural constraints within clearly defined research settings. Chapters five and six analyse the development of three specific modelling systems from the late 1930s to the mid-1960s. These chapters show these developments as contingent processes based on the involvement of elite researchers and both private and public institutions. It shows in particular how these modelling systems were made relevant across individual research sites by providing not only a technical solution to the problem of assembling model structures, but also by incorporating specific structural constraints. Finally, it is argued that these systems are one of

Cancer is the second leading cause of death in the United States (more than half a million people each year), and even with billions of dollars in medical effort patients are rarely cured. This dissertation research is devoted to meeting this medical need by providing new cancer therapeutics that are more potent and safer than current chemotherapies. This is achieved by using two state of the art anticancer “warheads”: 1) gold nanoparticle (AuNP) technology and 2) a new class of epigenetic anticancer small molecules, histone deacetylase inhibitors (HDACi). These warheads are then selectively delivered to cancer cells via “homing devices” targeted to receptors that are overexpressed in the cancers. This work primarily focuses on the androgen receptor (AR) to target prostate cancer. The 1st chapter sets the stage, providing scientific rationale and background for the central hypothesis: small molecules that engage the AR can, upon conjugation to a therapeutic agent, enable selective delivery of that agent to prostate cancer cells. Chapter 2 delves into the structural molecular biology of the androgen receptor. There is a survey of the crystallographic data for all nuclear receptors, providing structural information which is used to build AR homology models for antagonist and inverse agonist modes of ligand binding. These models are used to design AR targeting ligands (Chapters 3, 5, 6 and 7). The application of the targeting technology is illustrated by attaching them to AuNPs for selective delivery to prostate cancer cells (Chapter 3). Next, in order to appreciate the importance of using targeting agents in HDACi cancer therapeutics, we reviewed this recently emerged field in Chapter 4. In this chapter we argue that the failure of HDACi in solid tumors, despite more than 500 clinical trials in the last decade, is primarily due to an inability of these small molecules to accumulate at effective concentrations in the cancer. We provide an analysis of the paradigms being pursued to overcome this barrier, including HDAC isoform selectivity, localized administration, and targeting cap groups to achieve selective tissue and cell type distribution. In Chapter 5, this last approach (targeting cap groups, or a “homing device”) is illustrated with HDACi targeted to prostate cancer via antiandrogens that bind the AR. The second generation of improved “homing devices” is disclosed in Chapter 6 (for both AuNPs and HDACi), in addition to preliminary ADMET data and safety studies in mice. Excitingly, our three dimensional understanding of binding to the AR allowed design and structure-activity-relationship studies that lead to the first reported examples of AR inverse agonists (Chapter 7) Several points of significance: • AuNP targeted to AR ∙ have the strongest binding affinity ever reported (IC50 ~14 picomolar) ∙ are actively recruited to prostate cancer cells ∙ overcome treatment resistance in advanced prostate cancer cells ∙ exhibit nanomolar anticancer potency ∙ resolved the identity of the “membrane AR” as the GPRC6A • HDACi targeted to AR ∙ have HDACi activity and AR binding affinity superior to their clinical precursors ∙ exhibit potent AR antagonist activity ∙ induce AR translocation to the nucleus in a HDACi dependent fashion ∙ selectively and potently kill prostate cancer cells that express AR ∙ are safer than Tylenol®, as tested in small animals • Pure AR binding ligand studies ∙ resulted in the discovery of the first examples of AR inverse agonists, which are vastly more potent that clinically available antiandrogens for prostate cancer ∙ work via a never-before-seen mechanism of action, by localizing to the nucleus and recruiting corepressors to actively shut off AR genes

Bone defects treatment and reconstructive surgery continues to increase at a significant rate. Current bone defect treatments are autotransplantation, allograft, and xenografts create many problems such as, inflammation, infection and chronic pain. Moreover, allografts and xenografts arouse immune rejection. These problems have led to development of controlled release system for use as alternatives to autografts, allografts and xenografts in bone repair. There have been many approaches for sustained drug delivery in local bone regeneration using biodegradable polymers and osteotropic biomolecules. This dissertation presents new approaches that apply intermittent drug delivery for local bone regeneration. In the first, the osteotropic molecules simvastatin (Sim) or parathyroid hormone (PTH) were released with intermittent profiles. In the second, alternating delivery of Sim and PTH as well as alternating release of the antimicrobial agent cecropin B (CB) with Sim or PTH. An association polymer system of cellulose acetate phthalate (CAP) and Pluronic F-127 (PF-127) was used for the delivery vehicle. Each device showed discrete peaks in release profiles and lasted more than 10 days. Release profiles could be controlled by altering surface area exposed to aqueous environment, number of layers, loading, and blending ratios. Cells were cultured with sustained or intermittent exposure to Sim or PTH at various concentrations, and alternating exposure to CB and Sim or PTH and to Sim and PTH at different concentrations. Low dose Sim and PTH treatments stimulated higher osteoblastic activity than observed in control cultures. Furthermore, intermittent delivery was more effective than sustained exposure. In vivo, newly formed bone was found in animals implanted with both blank Sim-loaded devices. However, a greater anabolic effect was seen for Sim release devices. Further, intermittent release devices stimulated the greatest woven bone thickness, total bone area, and lamellar bone area. These results suggest that intermittent release devices containing a single molecule, Sim or PTH, and alternating release devices containing multiple molecules, CB with Sim or PTH, possess promising potential as a treatment for local bone regeneration.

Transient molecules are of great importance due to their critical role as intermediates in the semiconductor industry, in upper atmosphere reactions, and in astrochemistry. In the present work, reactive intermediates were produced in the laboratory by applying an electric discharge through a suitable precursor gas mixture and studied by means of their laser-induced fluorescence and emission spectra. The band systems of and have been studied in detail. The energy levels of both isotopologues were fitted with a Renner-Teller model, and the isotope relations have been used to test the validity of the derived parameters. The A2Πu - X 2Πg electronic transition of jet-cooled has been detected and shown to originate from the Ω=3/2 spin-orbit component of v=0 of the ground state. For the first time, the 0-0 band has been identified and vibrational assignments have been made. Our ab initio studies show that the extensive observed perturbations are due to spin-orbit interaction between A2Πu(3/2) and B2Δu(3/2) states. The experimental data were fitted to an effective Hamiltonian and yielded the spin-orbit coupling term =240 cm-1. LIF and emission spectra of the transition of N2O+ have been recorded. Both spin-orbit components of the band were studied at high resolution and rotationally analyzed, providing precise molecular constants. Emission spectra provided extensive data on the ground state vibrational levels which were fitted to a Renner-Teller model including spin-orbit and Fermi resonance terms. The previously unknown electronic spectrum of the H2PO radical has been identified. Ab initio predictions were used to aid in the analysis of the data. The band system is assigned as the electronic transition. The excited state molecular structure was determined by rotational analysis of high resolution LIF spectra. The band systems of the HBCl and DBCl free radicals have been studied in detail. This electron promotion involves a linear-bent transition between the two Renner-Teller components of what would be a 2Π electronic state at linearity. Ab initio potential energy surface calculations were used to help in assigning the LIF spectra which involve transitions from the ground state zero-point level to high vibrational levels of the excited state.