Browsing by Subject "tuberculosis"
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Item Development of Potent and Selective Inhibitors of Mycobacterium Tuberculosis, Plasmodium Falciparum and Staphylococcus Aureus Dihydrofolate Reductase(2014-12-17) Jung, HunminThe goal of this study was to develop drugs that exclusively affect pathogenic dihydrofolate reductase (DHFR) without causing harm to the human counterpart. To achieve that goal, a well-known dihydrofolate reductase (DHFR) inhibitors, trimethoprim (TMP), methotrexate (MTX) and trimetrexate (TMQ), were modified, tested, and crystallized on Mycobacterium tuberculosis (Mtb) dihydrofolate reductase (DHFR), wild type and quadruple mutant Plasmodium falciparum (Pf) DHFR-thymidylate synthase (TS), Staphylococcus aureus DHFR, and human DHFR. We focused on the drug design to utilize the structural differences between the pathogenic DHFRs and the human DHFR; specifically, we focused on a pocket near the substrate binding site where Asp27 and Gln28 of Mtb DHFR, and Asp54 and Met55 of Pf DHFR-TS are located. The same site is closely packed in human DHFR. From the initial screening and designing process, C-8 benzyl-2,4-diaminoquinazoline TMQ analogs were found to have outstanding selectivity against Mtb and Pf DHFR. Co-crystal structures of C-8 benzyl TMQ analogs with Mtb and Pf DHFR showed that the flexibility of Gln28 in Mtb DHFR, and Met55 in Pf DHFR contributes to extra space and interaction with C-8 benzyl moiety. This flexibility, which is not available in the human DHFR, enables the TMQ analogs to bind exclusively to the pathogenic DHFRs. Our novel C-8 benzyl-2,4-diaminoquinazoline TMQ analogs exhibited great potency and selectivity toward pathogenic DHFRs. In addition, these C-8 benzyl-2,4-diaminoquinazoline TMQ analogs were potent on Staphylococcus aureus DHFR as well and we hypothesize based on our findings that our C-8 benzyl-2,4-diaminoquinazoline TMQ analogs have potential for selective, broad spectrum antimicrobials against whose DHFR share the common structural feature with Mtb or Pf DHFR, an acid residue and a flexible residue next to it.Item Structural Studies of Bacteriophage Lysins and their Implication in Human Diseases(2011-08-08) Sun, QinganStructural biology lays the molecular foundation for the modern field of life sciences. In this thesis, X-ray crystallography is the primary resource for atomic detail structural information and is the major technology employed in our research. Three examples show how structural biology addresses the basic processes of life. Firstly, two crystal structures of R21, corresponding to two biological states, reveal a new activation mechanism of SAR-endolysin, which not only complements the previous model, but is also more generally applicable to the endolysin family. The structural information was further corroborated by NMR data in solution. The second example is the crystal structure of mycobacteriophage lysin B, which identified the function of the protein, and tackles the unique problem of how mycobacteriophage circumvent the mycolic acid-rich outer membrane of mycobacterium. The last example is the homology modeling of the Plasmodium ribosomal L4 protein. The action mode for the drug in Plasmodium was proposed based on that, which accounts for the anti-malaria effect of azithromycin.Item Structural studies of Mycobacterium tuberculosis KatG, an INH drug activator, and Brucella abortus VirB11, an ATPase of type IV translocation system(2009-05-15) Yu, HongCatalase-peroxidase (KatG) of Mycobacterium tuberculosis is a bifunctional heme enzyme that has been shown to play an important role in the activation of a first line drug, isoniazid (INH), used in the treatment of tuberculosis infection. Mutations in the katG gene have been found to be associated with INH resistance. The most commonly encountered mutation is the Ser315Thr point mutation. In this dissertation, the x-ray crystallographic structures of MtbKatG and the mutant enzyme KatG[S315T] are presented to explore the molecular basis of the INH activation and resistance. The structure is dimeric and contains a heme cofactor in each subunit of the dimer. The most important change in KatG[S315T] is due to the presence of the methyl group of the threonine 315 side chain, which is located at the narrowest part of the substrate channel. The protruding methyl group effectively constricts the accessibility to the heme by closing down the dimensions of the channel, constraining the substrate entrance. VirB11 of Brucella abortus is a hexameric ATPase that belongs to the type IV secretion system. The crystal structure of BaVirB11 was found to contain six molecules per asymmetric unit. The Walker A (P loop), His box, and Glu box from the C-terminal domain are located at the interface of the N- and C-terminal domain. A large conformational change was found in the linker region when compared with that of HP0525 structure, the VirB11 analogous from H. pylori. To elucidate the functional role of each domain, seven functional mutations were generated and used for biochemical studies. The GER motif and the linker region were found to be crucial for ATP hydrolysis activity of BaVirB11. Mutations in the GER motif (R101Q) and the linker region (R120E) of BaVirB11 completely abolish the ATP hydrolysis activity of the enzyme. The binding affinities of the two mutants to the ATP; however, are similar to that of the wild-type enzyme, indicating that mutation in the GER motif or the linker region has no effect on ATP binding.Item Structure-based drug mechanism study and inhibitor design targeting tuberculosis(2009-05-15) Wang, FengThe increase of multi-drug resistant and extensively drug resistant tuberculosis (TB) cases makes it urgent to develop a new generation of TB drugs to counter resistance and shorten treatment. Structural biology, which allows us to ?visualize? macromolecules, is now playing a key role in drug discovery. In this work, a structure-based approach was applied to the study of the mode of action of current TB chemotherapies, the identification of potential therapeutic targets, and the design of new inhibitors against TB. Knowledge of the precise mechanisms of action of current TB chemotherapies will provide insights into designing new drugs that can overcome drug-resistant TB cases. Structural biology combined with biochemical and genetic approaches was used to elucidate the mechanisms of actions of isoniazid, ethionamide and prothionamide. The active forms of these anti-TB prodrugs were identified by protein crystallography and the target-inhibitor interactions were revealed by the complex structures. Although these drugs are activated through two completely different routes, they all inhibit InhA, an essential enzyme in mycolic acid biosynthesis, by modification of the enzyme cofactor NAD, which unveils a novel paradigm of drug action. Isoniazid, ethionamide and prothionamide all target InhA, which validates the enzyme as a superb drug target. A structure-based approach was adopted to design new inhibitors targeting InhA, using triclosan as the scaffold. Guided by the InhA-inhibitor complex structures, two groups of triclosan analogs were identified with dramatically increased inhibitory activity against InhA. Structural biology has also provided fundamental knowledge of two potential therapeutic targets, Mtb ?-lactamase (BlaC) and fatty-acyl-CoA thioesterase (FcoT). Mtb ?-lactamase has been proposed to be the most significant reason for mycobacterial resistance to ?-lactam antibiotics. The determination of Mtb BlaC structure not only demonstrates the mechanism of drug resistance but also provides a solid base for the design of new ?-lactamase inhibitors that could be used with ?-lactam antibiotics as a new regimen to treat tuberculosis. The crystal structure of FcoT provided crucial information in identification of the function of this previously hypothetical protein. The characterization of FcoT revealed an important pathway that is critical for Mtb?s survival in host macrophages.Item Tuberculosis: Epidemiology, Diagnosis, Treatment, Prevention and Control in the United States and Worldwide(2013-06-03) Calderon, Veronica Elena; Mayhall, C. Glen; Arcari, Christine; de Boer, MelanieTB is one of the most common infectious diseases worldwide. Approximately one-third of the world’s population is infected with TB. In 2011, there are about 9 million new infections and almost 1.4 million deaths. Furthermore, TB is the leading cause of death in HIV-positive individuals. With the global HIV pandemic and the emergence of MDR- and XDR-TB, new diagnostics and treatments are urgently needed for the control and prevention of TB. Ultimately, the coordinated efforts of international and national government agencies, non-government agencies, healthcare professionals, and the public is needed to ensure the implementation and adherence of control strategies that will lead to the eradication of TB. This capstone will focus on the epidemiological and clinical aspects of TB, and the TB prevention and control measures as recommended by national and international organizations. The objectives of this project were accomplished through the direct observation of TB control and prevention measures in the hospital (UTMB) and public health (Galveston County Health District) settings. In addition, an extensive literature review was performed to gain a complete understanding of TB epidemiology, diagnostics, treatments, and prevention and control strategies.Item Tuberculosis: standards and resources for quality improvement in resource limited settings(2010-01-01) Katherine Louise McQuade; Christine Arcari, PhD MPH; Susan Weller, PhD; Philip Keiser, MDTuberculosis is a significant problem, infecting nearly 9 million new patients per year and killing about 2 million a year. The fight to eradicate TB is located primarily in countries that are resource poor and disproportionately affected by HIV and the development of drug resistant TB. The primary means with which to affect TB globally are to decrease transmission locally, mainly by effective identification, diagnosis, and treatment of TB patients. Providing quality care to TB patients is therefore essential to the global effort to eradicate TB. This review describes the problem of TB, identifies the essential services of TB control programs and standards of care, and reviews the available resources for quality improvement in resource limited settings. Finally, recommendations for a quality improvement plan for a TB control program in resource limited settings are made.