This dissertation explores the structural modification of indigo into its rare cis form by substitution of a short organic bridge between the two indole nitrogen atoms. The synthesized N,N′-tethered cis-indigoids are assessed for their physicochemical properties in order to understand the effect of the organic tether on cis-indigoid systems. Three literature compounds containing “simple” tether structures are synthesized and subjected to complete characterization. Among them, alkyl group based cis-indigos, 2.5 and 2.6 exhibit similar absorption wavelengths as the parent indigo while oxalyl-tethered indigo 2.7 shows a large hypsochromic shift due to the strong electron-withdrawing nature of the oxalyl group. The electronics of the tether also affected the HOMO and LUMO energy levels; the oxalyl tether lowered both energy levels in comparison to the alkyl tethers. However, the indigoid co-planarity was strongly affected by the ring size from the tether rather than electronics. A new tethered cis-indigo structure type was discovered in reactions involving quinones. This new tether type consists of a 2,2′-dihydroindigo unit connected to the cis-indigo backbone through a central C–C bond of the former. Incorporation of the quinone moiety was observed in 3.2 where the final structure is comprised of two molecules of indigo and one naphthoquinone, while structures of 3.3 and 3.4 contained two indigoid units only. Investigation by CV revealed the strength of the quinone altered the early reaction intermediates. These dimeric cis-indigos show a small hypsochromic shift in the absorption compared to the parent indigo and relatively planar cis-indigoid backbone. 3.2 demonstrates rich redox behaviours. 3.3 and 3.4 display dynamic behaviours observed by solution VT 1H NMR spectroscopy. Oxalyl-tethered cis-Nindigo 4.6a and cis-indigo monoimine 4.7a were synthesized and compared with the indigoid counterpart 2.7 to examine the influence of the arylimine groups. Absorption wavelengths of the imine group containing species depended strongly on the electronics of the tether, yet the number of imine groups affected redox potentials. Protonation of 4.6a (to give H+oxalyl Nindigo 4.6aH+) causes a bathochromic shift in the absorption and allows for much easier reduction. The acidity (pKa) of 4.6aH+ is estimated between 3.6 – 4.46 in DMSO. A protonated cis-Nindigo derivative 4.9aH+ was obtained via reduction of the oxalyl group and is compared with 4.6aH+. / Graduate / 2022-08-13

No description available.

Phylum Apicomplexa comprises several thousand parasitic protozoans that cause significant disease in humans and animals worldwide. Of particular relevance to human health are Plasmodium spp., the causative agents of malaria; and Toxoplasma gondii, which infects approximately 30% of all humans on earth, and causes serious disease in immunocompromised individuals and neonatally infected fetuses. Central to the pathogenesis of apicomplexans is a unique form of substrate-dependent locomotion termed “gliding motility”, which is essential for traversing the environment and actively invading host cells. Driving motility is the class-XIV unconventional myosin motor (MyoA), which is notably divergent from canonical myosins in that it lacks a “tail” and conventional sequence motifs in both the neck and motor regions. Thus, the mechanisms that enable MyoA to function with a step size and velocity similar to canonical human myosins are not well understood. Over the past 2 decades, the apicomplexan research community has identified many of the components involved in gliding motility, resulting in a functional model of MyoA and accessory proteins forming the “glideosome” macromolecular complex. However, there was still relatively little known about the unique physical processes that drive force production and transduction in the apicomplexan motor complex. Thus, I set out to use structural and biophysical methods to interrogate this divergent molecular motor, and provide the first high-resolution model of apicomplexan motility. Towards this goal, I first used structural and biophysical methods to establish the most complete model to date of class-XIV motor complex assembly, answering key questions about the interface between MyoA and its accessory proteins. To understand the unique molecular basis of force production in apicomplexan motors, I then solved the first ever crystal structure of a class-XIV myosin, MyoA from T. gondii. Supplementing this structure with further biophysical data, I was able to determine the functional consequences of class-defining sequence polymorphisms, and elucidate the basis of phosphorylation-dependent motor regulation. The systematic dissection of apicomplexan motor complexes described herein provides crucial insight into a fundamental biological process, and may help overcome existing barriers for targeted therapeutic development. / Graduate

Vitamin B1 is an indispensable co-factor for various enzymes inter alia in the Krebs cycle, pentose phosphate pathway, nucleotide and amino acid synthesis. Due to its importance in metabolism, proteins involved in the synthesis of vitamin B1 have been identified as potential drug targets. Thiamine monophosphate kinase (ThiL), catalyses the last reaction in the pathway, the ATP dependent phosphorylation of thiamine monophosphate (TMP) producing thiamine pyrophosphate (TPP) the active and co-factor form of vitamin B1. In this study, thiamine monophosphate kinase from Mycobacterium tuberculosis (MtbThiL, ~36 kDa) was produced as an N-terminally His6-tagged fusion protein, purified by affinity and size exclusion chromatography, and crystallised. Hexagonal MtbThiL crystals belonged to space group P6122. Molecular replacement revealed a symmetric homodimer with a single monomer occupying the asymmetric unit. Analysis of the structure showed that each subunit of MtbThiL has an ATP and TMP binding site and is structurally related to other ThiL enzymes. Ten lead compounds were identified from compound databases as potential ThiL inhibitors, and oxythiamine was chosen for further study. The binding affinities of oxythiamine and TMP to MtbThiL were determined by isothermal titration calorimetry and a pyruvate kinase-lactate dehydrogenase enzyme assay, which revealed that the binding affinity for oxythiamine by MtbThiL is lower than the substrate TMP. / Dissertation (MSc)--University of Pretoria, 2020. / Biochemistry / MSc / Unrestricted

The main focus of this thesis is on the phase retrieval problem. This problem has a broad range of applications in advanced imaging systems, such as X-ray crystallography, coherent diffraction imaging, and astrophotography. Thanks to its broad applications and its mathematical elegance and sophistication, phase retrieval has attracted researchers with diverse backgrounds. Formally, phase retrieval is the problem of recovering a signal 𝔁 ∈ ℂⁿ from its phaseless linear measurements of the form |𝛼ᵢ∗𝔁| + 𝜖ᵢ where sensing vectors 𝛼ᵢ, 𝑖 = 1, 2, ..., 𝓶, are in the same vector space as 𝔁 and 𝜖ᵢ denotes the measurement noise. Finding an effective recovery method in a practical setup, analyzing the required sample complexity and convergence rate of a solution, and discussing the optimality of a proposed solution are some of the major mathematical challenges that researchers have tried to address in the last few years. In this thesis, our aim is to shed some light on some of these challenges and propose new ways to improve the imaging systems that have this problem at their core. Toward this goal, we focus on the high-dimensional setting where the ratio of the number of measurements to the ambient dimension of the signal remains bounded. This regime differs from the classical asymptotic regime in which the signal's dimension is fixed and the number of measurements is increasing. We obtain sharp results regarding the performance of the existing algorithms and the algorithms that are introduced in this thesis. To achieve this goal, we first develop a few sharp concentration inequalities. These inequalities enable us to obtain sharp bounds on the performance of our algorithms. We believe such results can be useful for researchers who work in other research areas as well. Second, we study the spectrum of some of the random matrices that play important roles in the phase retrieval problem, and use our tools to study the performance of some of the popular phase retrieval recovery schemes. Finally, we revisit the problem of structured signal recovery from phaseless measurements. We propose an iterative recovery method that can take advantage of any prior knowledge about the signal that is given as a compression code to efficiently solve the problem. We rigorously analyze the performance of our proposed method and provide extensive simulations to demonstrate its state-of-the-art performance.

Thesis (Nucl.E.)--Massachusetts Institute of Technology, Dept. of Nuclear Engineering, 1981. / MICROFICHE COPY AVAILABLE IN ARCHIVES AND SCIENCE. / Includes bibliographical references. / by El-Khider Si Ahmed. / Nucl.E.

Indiana University-Purdue University Indianapolis (IUPUI) / The human aldehyde dehydrogenase superfamily consists of 19 distinct genetic loci that play key roles in both health and disease. Aldehyde dehydrogenases are primarily involved in the metabolism of reactive aldehyde substrates; the ALDH1A subfamily, in particular, metabolizes retinaldehyde and is involved in a pathway regulating tissue differentiation, cell proliferation, and apoptosis. Recently, ALDH1 isoenzymes have been implicated as significant elements in cancer progression. ALDH1 activity has been used as a marker of cancer stem cells, a subpopulation of cancer stem cells with high drug resistance, proliferative potential, and ability to differentiate into multiple cell types. In accordance with this, ALDH1 activity and expression has been shown to correlate with lower survival, increased chemoresistance, and increased chance of relapse in multiple solid cancer types, including breast, ovarian, lung, and colorectal. Despite the clear relevance of ALDH1 enzymes in cancer, the specific roles of individual isoenzymes are unclear. Isoenzyme-selective small molecule modulators of the ALDH1A subfamily would allow the probing of the function of individual isoenzymes in healthy and disease states. Two ALDH1A1 inhibitors, CM38 and C10, were previously identified in a high-throughput screen. In this study, CM38, an ALDH1A1-selective inhibitor, and CM10, an ALDH1A inhibitor, were characterized using kinetic assays, structural biology, and cell culture experiments. A structure-activity relationship was built for each series, and an X-ray crystallography structure was used to determine the binding mode. These approaches allowed the investigation of the ALDH1A active site and identification of structural features that can be used to design and improve selective modulators of this subfamily. CM38 and CM10 were also tested in a breast cancer cell line to determine their efficacy in a cellular environment. While the CM38 series showed warning signs of potential off-target toxicity, members of the CM10 compound series showed excellent initial characteristics as potential chemical tools. The results of this study may be useful in the design of new chemical tools to delineate the functions of individual ALDH1 isoenzymes in cancer biology, as well as in the development of drugs to selectively target cancer stem cells.

Indiana University-Purdue University Indianapolis (IUPUI) / The over-accumulation of glycogen appears as a hallmark in various glycogen storage diseases (GSDs), including Pompe, Cori, Andersen, and Lafora disease. Glycogen synthase (GS) is the rate-limiting enzyme for glycogen synthesis. Recent evidence suggests that suppression of glycogen accumulation represents a potential therapeutic approach for treating these diseases. Herein, we describe the discovery, characterization, and development of small molecule inhibitors of GS through a multicomponent study including biochemical, biophysical, and cellular assays. Adopting an affinity-based fluorescence polarization assay, we identified a substituted imidazole molecule (H23), as a first-in-class inhibitor of yeast glycogen synthase 2 (yGsy2) from the 50,000 ChemBridge DIVERSet library. Structural data derived from X-ray crystallography at 2.85 Å, and enzyme kinetic data, revealed that H23 bound within the uridine diphosphate glucose binding pocket of yGsy2. Medicinal chemistry efforts examining over 500 H23 analogs produced structure-activity relationship (SAR) profiles that led to the identification of potent pyrazole and isoflavone compounds with low micromolar potency against human glycogen synthase 1 (hGYS1). Notably, several of the isoflavones demonstrated cellular efficacy toward suppressing glycogen accumulation. In an alternative effort to screen inhibitors directly against human GS, an activity-based assay was designed using a two-step colorimetric approach. This assay led to the identification of compounds with submicromolar potency to hGYS1 from a chemical library comprised of 10,000 compounds. One of the hit molecules, hexachlorophene, was crystallized bound to the active site of yGsy2. The structure was determined to 3.15 Å. Additional kinetic, mutagenic, and SAR studies validated the binding of hexachlorophene in the catalytic pocket and its non-competitive mode of inhibition. In summary, these two novel assays provided feasible biochemical platforms for large-scale screening of small molecule modulators of GS. The newly-developed, potent analogs possess diverse promising scaffolds for drug development efforts targeting GS activity in GSDs associated with excess glycogen accumulation. / 2021-07-01

京都大学 / 0048 / 新制・課程博士 / 博士(農学) / 甲第19195号 / 農博第2134号 / 新制||農||1034(附属図書館) / 学位論文||H27||N4941(農学部図書室) / 32187 / 京都大学大学院農学研究科食品生物科学専攻 / (主査)教授 谷 史人, 教授 保川 清, 准教授 橋本 渉 / 学位規則第4条第1項該当 / Doctor of Agricultural Science / Kyoto University / DFAM

京都大学 / 0048 / 新制・課程博士 / 博士(工学) / 甲第20398号 / 工博第4335号 / 新制||工||1672(附属図書館) / 京都大学大学院工学研究科分子工学専攻 / (主査)教授 関 修平, 教授 今堀 博, 准教授 伊藤 彰浩, 教授 白川 昌宏 / 学位規則第4条第1項該当 / Doctor of Philosophy (Engineering) / Kyoto University / DGAM