Browsing by Author "Bruce Novak, Committee Member"

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Activators Generated by Electron Transfer for Atom Transfer Radical Polymerization in Polymerization-based DNA Sensing.
(2010-12-16) Qian, Hong; Lin He, Committee Chair; Morteza Khaledi, Committee Member; Bruce Novak, Committee Member; Edmond Bowden, Committee Member
Atom Transfer Radical Polymerization (ATRP)in Amplification-by-Polymerization for DNA Sensing
(2007-08-07) Lou, Xinhui; Bruce Novak, Committee Member; Lin He, Committee Chair; Daniel L. Feldheim, Committee Member; Morteza G. Khaledi, Committee Member
DNA sensing has attracted a lot of attention due to its importance in biological and medical fields. Numerous methods have been developed to amplify DNA hybridization signals to meet different needs. The purpose of this research was to develop a simple yet efficient method to detect sequence specific DNA in a home testing kit format according to the concept of amplification-by-polymerization. This dissertation reports the development of an atom transfer radical polymerization (ATRP)-based DNA detection method. The background knowledge on current DNA sensing methods and the applications of polymers in DNA sensing technologies are described in Chapter 1. Chapter 2 describes the proof-of-concept experiments of this DNA detection method. In this method, DNA hybridization and ligation reactions led to the attachment of ATRP initiators on gold surface where specific DNA sequences located. These initiators subsequently triggered the growth of polymer at the end of DNA molecules. Only the perfectly matched targets were distinctively observed by the naked eye due to the formation of polymer that altered the substrate opacity. The demonstrated capability to detect DNA with direct visualization laid the groundwork for the future development of detector-free testing kits in DNA sensing. Chapter 3 describes the two strategies to further improve the sensitivity of this detection method: by formation of branched polymer through repetitive ATRP and by minimization of the background noise through optimization of the passivation layer in DNA monolayers. Chapter 4 describes the kinetics of DNA⁄polymer formation using different catalyst systems. The effects of the composition and concentration of the catalysts used during DNA-accelerated ATRP reaction were evaluated. The results showed a strong correlation between polymer formation and the reaction conditions. The results also showed that the presence of DNA molecules significantly fastened the growth rates of both PHEMA and POEGMA in ATRP. This accelerating effect was suspected as a combined result of the highly charged DNA backbones and the unique chemical structure of DNA molecules. Chapter 5 describes the two applications of this ATRP-based detector-free DNA detection method: in single nucleotide polymorphism (SNP) detection and in human gender determination. Chapter 6 describes a colorimetric ATRP-based DNA detection method based on the increased stability of polymer coated gold nanoparticles (GNPs). In this method, hybridizations, ligation and ATRP were conducted on ssDNA labeled GNPs. GNPs remained red color or aggregated after ATRP at the presence or absence of complementary target DNA in the hybridization step, respectively. The reported method provides a generic approach for biomolecular hybrid formation on a solid surface and could open up new possibilities in the applications of DNA detection and gene delivery. Chapter 7 describes a direct comparison between CNBr chemical ligation and T4 ligation. Much higher ligation efficiency and specificity of CNBr ligation was found compared to T4 ligation, which renders the potential applications of CNBr ligation in DNA sensing to replace enzymatic ligation.
Control of Biological Processes with Modified Nucleosides, Amino Acids, and Small-Molecule Probes
(2010-07-29) Lusic, Hrvoje; Paul Agris, Committee Member; Tatyana Smirnova, Committee Member; Jonathan Lindsey, Committee Member; Bruce Novak, Committee Member; Alex Deiters, Committee Chair
Several novel caging groups with a wide range of applications were designed. The caging groups were applied to the synthesis of molecular tools for control of gene expression. A number of RNA, DNA, and antisense-oligomer subunits were prepared. The caged nucleosides were site-specifically incorporated into corresponding oligonucleotide strands, using standard automated-synthesis protocols. The caged nucleosides were found to readily photolyse under biologically non-damaging UV-light (>365 nm). Spatio-temporal control over oligomersâ€™ biological and/or catalytic activity was demonstrated with excellent on(or off) photo-triggered switches. Additionally, non-standard nucleosides 5-formylcytidine and N-[(9-D-ribofuranosyl-2-methylthiopurin-6-yl)carbamoyl]threonine were synthesized for incorporation into tRNA to elucidate the importance of the non-standard nucleoside modifications on codon recognition in RNA translation. Furthermore, tools for control over protein functions were designed. A set of novel unnatural amino acids were successfully synthesized and genetically (in vivo) incorporated into proteins using evolved orthogonal tRNA/synthetase pairs. Lastly, several caged protein kinase inhibitors were prepared, and used to study importance of kinase enzymatic activity in embryonic development of Xenopus laevis.
The Design, Synthesis and Biological Examination of Marine Natural Product Inspired Antibiofilm Agents and Antibiotics.
(2010-06-23) Rogers, Steven; Christian Melander, Committee Chair; Daniel Comins, Committee Member; Reza Ghiladi, Committee Member; Frank Edens, Committee Member; Bruce Novak, Committee Member
Design, Synthesis, and Biological Evaluation of Novel Anti-Biofilm Molecules Derived from the Oroidin Alkaloids
(2009-07-07) Richards, Justin James; Bruce Novak, Committee Member; alexander deiters, Committee Member; Christian Melander, Committee Chair; Daniel Comins, Committee Member
Bacterial biofilms are a surface attached community of microorganisms that are protected by an extracellular matrix of biomolecules. Within a biofilm state, bacteria are more resistant to antibiotics and are inherently insensitive to antiseptics and basic host immune responses. Biofilm infections of indwelling medical devices are also of major concern, as once the device is colonized, infection is virtually impossible to eradicate. Given the prominence of biofilms in infectious diseases, there has been an increasing effort toward the development of small molecules that will modulate bacterial biofilm development and maintenance. This, coupled with the spread of multi-drug antibiotic resistance across many of these bacteria, has put a tremendous burden on the medical community to alleviate biofilm related problems. Herein the design, synthesis, and biological evaluation of small molecules derived from the oroidin class of marine natural products that both inhibit and disperse a number of medically relevant bacterial biofilms through a non-microbicidal mechanism is highlighted. This includes improvements to existing methodological approaches aimed toward the synthesis of these molecules in addition to the implementation of novel synthetic pathways which allow for the generation of diverse chemical libraries. Detailed structure-activity relationship (SAR) studies were performed with the ultimate goal of delineating which structural features of the analogues were essential for a biological response within the context of anti-biofilm activity. Preliminary toxicology studies also indicate that many of the analogues display non-cytotoxic properties, thus furthering hope for the use of these molecules in therapeutic biofilm remediation efforts.
Label-free, Multiplexed Bioassay on Au/Ag Striped Nanorods Using Fluorescent Conjugated Polymers.
(2010-08-16) Zheng, Weiming; Lin He, Committee Chair; Edmond Bowden, Committee Member; Bruce Novak, Committee Member; Matthew Breen, Committee Member
Modification of Nylon 6 Structure via Nucleation
(2009-08-12) Mohan, Anushree; Bruce Novak, Committee Member; Jan Genzer, Committee Member; Alan E. Tonelli, Committee Chair; Richard Kotek, Committee Co-Chair
For nearly two decades inclusion compounds (ICs) have been formed by threading polymer chains into the cyclic starches, cyclodextrins (CDs). Non-covalently bonded crystalline ICs have been formed by threading CDs, onto guest nylon-6 (N6) chains. When excess N6 is employed, non-stoichiometric (n-s)-N6-CD-ICs with partially uncovered and dangling N6 chains result. We have been studying the constrained crystallization of the N6 chains dangling from (n-s)-N6-CD-ICs in comparison with bulk N6 samples, as a function of N6 molecular weights, lengths of uncovered N6 chains, and the CD host used. While the crystalline CD lattice is stable to ~ 300Ã‚Â° C, the uncovered and dangling, yet constrained, N6 chains may crystallize below, or be molten above ~225Ã‚Â° C. In the IC channels formed with host ÃŽÂ±- and ÃŽÂ³-CDs containing 6 and 8 glucose units, respectively, single and pairs of side-by-side N6 chains can be threaded and included. In the ÃŽÂ±-CD-ICs the ~ 0.5nm channels are separated by ~ 1.4nm, while in ÃŽÂ³-CD-ICs the ~ 1nm channels are ~ 1.7 nm apart, with each ÃŽÂ³-CD channel including two N6 chains. The constrained dangling chains in the dense (n-s)-N6-CD-IC brushes crystallize faster and to a greater extent than those in bulk N6 melts, and this behavior is enhanced as the molecular weights/chain lengths of N6 are increased. Furthermore, when added at low concentrations (n-s)-N6-CD-ICs serve as effective nucleating agents for the bulk crystallization of N6 from the melt. Because of the biodegradable/bioabsorbable nature of CDs, (n-s)-polymer-CD-ICs can provide environmentally favorable, non-toxic nucleants for enhancing the melt crystallization of polymers and improving their properties.
Sorption to Dissolution: The Reactivity of Small Molecules with Condensed Phase Metal Halide Networks
(2009-11-04) Wilcox, Robert Joseph; Bruce Novak, Committee Member; Paul A. Maggard, Committee Member; Elon Ison, Committee Member; James D. Martin, Committee Chair
Synthesis of Diene and Dienophile Substrates for In-vitro Selection of RNA-hetero-diels-alder Catalysts
(2005-05-02) Latta, Paul Richard III; Jon Lindsey, Committee Member; Bruce Novak, Committee Member; Bruce Eaton, Committee Chair
The hetero-Diels-Alder reaction provides a general and facile entry for the synthesis of six-membered heterocyclic structures, which serve as precursors for a host of biologically active compounds. Although there are a wide variety of heterodienes and heterodienophiles available for synthetic applications, our research focuses on the cycloaddition of activated 1,3-butadienes with an activated aromatic aldehyde dienophile and an unactivated aliphatic aldehyde dienophile. Cycloadditions of this type provide access to an important class of compounds known as dihydropyrans, which are useful intermediates for the preparation of carbohydrates and many other biologically active compounds. Synthesis of pyrans by HDA cycloaddition has been limited to the reaction of either highly electron rich dienes (e.g. Danishefsky's diene) with a broad range of aldehydes or highly activated aldehydes (e.g. glyoxylates) with dienes of varying activity. Currently there are few examples of cycloadditions between unactivated aldehyde dienophiles and dienes that are less activated than Danishefsky's diene. Also, the catalysts currently available for this transformation provide access to only one of two possible regioisomers and provide a single diastereomer almost exclusively. If efficient catalysts for the uncommon HDA regio- and stereoisomers are attainable, then we believe that in vitro selection of these catalysts from a pool of 10∧14 random RNA sequences will provide invaluable clues as to structural properties necessary for such catalysts. To this end, we have prepared novel dienes and dienophiles suitable for in vitro selection of new RNA biocatalysts capable of assembling novel heterocyclic products via a hetero-Diels-Alder cycloaddition.
Synthesis of Substituted Bacteriochlorins
(2010-04-22) Meneely, Kelly Rose; Christian Melander, Committee Member; Jonathan Lindsey, Committee Chair; Bruce Novak, Committee Member
The objective of this work is two-fold: first, to find new conditions to improve the condensation reactions leading to the macrocycles 3,13-dibromo-5-methoxybacteriochlorin (BC-Br3OMe5Br13) and 3,13-dibromobacteriochlorin (BC-Br3Br13), and second, to functionalize the valuable BC-Br3OMe5Br13 building block. This work improves upon past condensation methods, which afforded low yields of bacteriochlorin with an inseparable chlorin product during formation of BC-Br3Br13, and provided no access to BC-Br3OMe5Br13. Through microscale optimization of conditions, solvents, concentrations, Lewis acids, and additives, BC-Br3OMe5Br13 and BC-Br3Br13 were isolated in 42% and 30% yield, respectively. Several derivatizations including diacetylation of BC-Br3OMe5Br13 was achieved in 70% yield, followed by bromination at the Î²-position in 22% yield. Stille coupling of BC-Br3OMe5Br13 to introduce a formyl group was performed in 30% yield. Two TIPS-ethynyl groups were also installed on BC-Br3OMe5Br13 in 4.7% yield.
Synthesis, Characterization, and Reactivity of Monomeric Copper(I) Complexes Possessing pi-Donor Ligands.
(2010-05-14) Delp, Samuel; Thomas Gunnoe, Committee Chair; Bruce Novak, Committee Member; Elon Ison, Committee Member; Wayne Buhler, Committee Member; James Martin, Committee Member