Diamidines are a class of compounds that target the minor groove of DNA and have antiparasitic and antimicrobial properties. Their mechanism of action has not been fully elucidated, but may include changes in DNA topology. In this study we have investigated such changes using methods of gel electrophoresis including ligation ladders and cyclization assays. We found that topology changes were sequence dependent. Compounds typically caused non-anomalously migrating ATATA sequences to migrate as if they were bent, while A5 sequences that normally migrated anomalously became less so in the presence of certain diamidines. Select compounds induced changes in cyclization efficiency that were also sequence dependent; DB75 significantly abolished cyclization in A5 containing sequences but enhanced it in sequences containing ATATA sites.

SHAPE-DEPENDENT MOLECULAR RECOGNITION OF SPECIFIC SEQUENCES OF DNA BY HETEROCYCLIC CATIONS by YI MIAO Under the Direction of Dr. W. David Wilson ABSTRACT DB921 and DB911 are biphenyl-benzimidazole-diamidine isomers with a central para- and meta-substituted phenyl group, respectively. Unexpectedly, linear DB921 has much stronger binding affinity with DNA than its curved isomer, DB911. This is quite surprising and intriguing since DB911 has the classical curved shape generally required for strong minor groove binding while DB921 clearly does not match the groove shape. Several biophysical techniques including thermal melting (Tm), circular dichroism (CD), biosensor-surface plasmon resonance (SPR), and isothermal titration calorimetry (ITC) have been utilized to investigate the interactions between these compounds and DNA. The structure of the DB921-DNA complex reveals that DB921 binds to DNA with a reduced twist of the biphenyl for better fit of DB921 into the minor groove. A bound water molecule complements the curvature of DB921 and contributes for tight binding by forming H-bonds with both DNA and DB921. Structure-affinity relationship studies of a series of DB921 analogs show that the benzimidazole group is one of the key groups of DB921 for its strong binding to the minor groove. Thermodynamic studies show that the stronger binding of DB921 is due to a more favorable binding enthalpy compared to DB911 even though the complex formation with DNA for these compounds are all predominantly entropically driven. DB921 also has more negative heat capacity change than DB911. The initial studies of inhibition of the interaction between an AT hook peptide of HMGA proteins and its target DNA by a set of diamidine AT-minor groove binders using biosensor-SPR technique show that the inhibitory ranking order is consistent with that of binding affinity and linear-shaped DB921 still has excellent inhibitory effects. These heterocyclic cations rapidly inhibit the binding of DBD2 peptide to the DNA and may only block the specific AT binding of the peptide without hindering the non-specific binding interaction. The results of this project have shown that DB921 represents a new novel effective minor groove binder that does not fit the traditional model and is a potential inhibitor for DNA/protein complexes. INDEX WORDS: Molecular recognition, DNA binding, Minor groove binding, Linear shape, Compound curvature, Binding affinity, Binding kinetics, Thermodynamics, Surface plasmon resonance, Isothermal titration calorimetry, Inhibition

Small molecule mediated chemical intervention of biological processes using nucleic acid targets has proven extremely successful and is continually providing exciting new avenues for the development of anti-cancer agents and molecular probes. Among the alternative DNA confrormations, G-quadruplexes has certainly garnered much recognition due to increase in evidences supporting their involvement in diverse biological process. The grooves of the quadruplexes offer an alternate recognition site for ligand interactions with potentially higher selectivity than the traditional terminal stacking sites. DB832, a bifuryl-phenyl diamidine, was recently reported to selectively recognize human telomeric G-quadruplex, as a stacked species, with significant selectivity over duplex sequences. A series of biophysical studies were conducted to test the groove-binding mode of DB832, along with the selectivity for diverse quadruplex forming sequences. To gain better understanding of quadruplex groove-recognition by DB832, a series of structurally similar heterocyclic diamidines were also evaluated. The unique binding mode of DB832 may allow it to serve as a paradigm for the design of new class of highly selective quadruplex groove-binding molecules. Beyond the alternative secondary structures, it is also becoming increasingly apparent that the structure and dynamics of the canonical Watson–Crick DNA double helix play pivotal roles in diverse biological functions. DB1878, a phenyl-furan-indole diamidine, was shown to recognize a mixed GC/AT motif as a stacked antiparallel dimer, and a detailed structural analysis is reported here. Interestingly, the DNA recognition is completely different from the reported molecules in literature, and represents an entirely new motif for DNA minor groove recognition.

DNA minor groove binding compounds have had limited therapeutic uses, in part due to problems with sequence specificity. A two-site model has been developed to enhance specificity, in which compounds bind to two short AT sites separated by one or two GC base pairs. Using thermal melting, heterocyclic dications with this capability were tested with various oligonucleotides for binding affinity and specificity. Compounds of interest were further probed using circular dichroism, mass spectrometry, biosensor-SPR, and molecular modeling. Several compounds were found to “jump” a GC base pair, binding to AT sites in the minor groove of DNA with a two-site recognition mode. One compound was also found to recognize a single intervening GC base pair. Compounds with terminal, non-polar amidine extensions were found to have increased DNA binding compared to analogs with terminal amidines. This unique, two-site DNA recognition mode offers novel design principles to recognize entirely new DNA motifs.

Utilizing bulk micromachining technology, this thesis aimed to develop a flexural plate-wave(FPW) device with novel groove reflection microstructure for high-sensitivity and low insertion-loss biomedical microsystem applications. The influences of the amount and depth of the groove and the distance between the groove and the boundary of ZnO piezoelectric thin-film (DGB) on the reduction of insertion-loss and the enhancement of quality factor (Q) and electromechanical coupling coefficient (K2) were investigated. Three critical technology modules established in this thesis are including the development of (1) a sputtering deposition process of high C-axis (002) orientation ZnO piezoelectric thin-film, (2) an electrochemical etch-stop technique of silicon anisotropic etching and (3) an integration process of FPW device. Firstly, under the optimized conditions of the sputtering deposition process (300¢J substrate temperature, 200 W radio-frequency (RF) power and 30/70 Ar/O2 gas flow ratio), a high C-axis (002) orientated ZnO piezoelectric thin-film with a high X-ray diffraction (XRD) intensity (50,799 a.u.) and narrow full width at half maximum (FWHM = 0.383¢X) can be demonstrated. The peak of XRD intensity of the standard ZnO film occurs at diffraction angle 2£c = 34.422¢X, which matches well with our results (2£c = 34.357¢X). Secondary, an electrochemical etch-stop system with three electrode configuration has been established in this research and the etching accuracy can be controlled to less than 1%. Thirdly, this thesis has successfully integrated the main fabrication processes for developing the FPW device which are including six thin-film deposition processes and six photolithography processes. The implemented FPW device with RIE etched groove reflection microstructure presents a low insertion-loss of -12.646 dB, center frequency of 114.7 MHz, Q factor of 12.76 and K2 value of 0.1876%.

In this study, the PDMS with microgroove structure was used controlling pretilt anlge of liquid crystal. Polydimethylsiloxane, also called PDMS, is one transparent, flexible, and stable material. It was usually fabricated the flexible display and so on. Based on Groove Theory, we can create the microgroove structure with the different groove depths and the width of the lines on PDMS by Replica Molding Method, in order to controlling the pretilt anlge of liquid crystal. We used the photoresist with different thickness to developing, and then the groove will get with different depth of groove. The PDMS was injected to the surface of groove with slow motion. When the liquid-like PDMS was curing, the PDMS can readily convert into solid elastomers by cross-linking. Finally, The microgrooved PDMS structure will obtain.

This thesis provides experimental static and dynamic results for four pairs of seals, including a pair of smooth seals and three pairs of centrally grooved seals. The grooved seals have groove depth to clearance ratios (Dg/C) of 5, 10, and 15. The radial clearance of each test seal is 0.0891 mm. Test conditions include three shaft rotational speeds from 4000 to 10000 rpm, three inlet oil pressures from 24 to 70 bars, and seal dimensionless eccentricities from 0 (centered) to 0.7. For each pair of test seals, dynamic results include stiffness and damping coefficients; static results include stator position, attitude angles, and seal leakage. Stiffness, damping, and leakage are compared among the seal pairs with various groove depths. Results show that all rotordynamic coefficients consistently decrease with increasing seal groove depths, and seal leakage remains constant through varying groove depths. Additionally, a comparison is made between experimental results of all test seals and XLLubeGT. XLLubeGT is a computer model developed at Texas A&M University, which uses a Reynolds equation + energy equation model to predict dynamic performance of a grooved seal. It operates on the assumption that the groove is large enough to create separate lands within the seal, creating a zero or negligible pressure perturbation across the seal. A comparison with XLLubeGT shows that even the seal with the largest groove depth tested is not deep enough to agree with XLLubeGT predictions.

Between 1985 and 1997 a hip hop culture emerged in Halifax, Nova Scotia through rap music artists' live performances at various venues, and by releasing original music on commercial and non-commercial cassette tapes and compact discs. This thesis examines the evolution of this grassroots musical culture through the lenses of Halifax's geography, innovative musical and technological trends, ever-present racial politics, and a strong "do-it-yourself" ethic. This thesis argues that hip hop in Halifax during these years can be divided into two eras distinguished by dynamic racial and stylistic changes. While the 1980s saw a predominantly Black hip hop community take root around Uniacke Square and Gottingen Street, the 1990s saw a geographic and demographic shift as the rap music scene expanded and competed with the mainstream music scene of the city. In doing so, the integrated downtown hip hop community produced a significant amount of work, overcoming institutional opposition

The field of popular music studies currently lacks effective and extensive discourse on drumming and rhythmic parameters in general. Some important preliminary work exists primarily due to significant contributions by relatively few authors. This thesis serves to expand this literature by providing a detailed explanation of many of the primary elements involved in the intricate practice of rock drumming. Additionally, it expands the literature on the music of Jimi Hendrix by thoroughly exploring the musical contributions Mitch Mitchell made as the drummer for the Jimi Hendrix Experience. Mitchell's stylistic approach to rock drumming is illuminated through analysis of the drum parts in four of the group's songs. An explanation of rock drumming in general and one effective individual approach are present within the work.

PURPOSE: This phase I dose-escalation study was undertaken to establish the maximum tolerated dose of the sequence-selective minor groove DNA binding agent SJG-136 in patients with advanced solid tumors. The study also investigated antitumor activity and provided pharmacokinetic and pharmacodynamic data. EXPERIMENTAL DESIGN: Sixteen patients were assigned sequentially to escalating doses of SJG-136 (15-240 microg/m(2)) given as a 10-minute i.v. infusion every 21 days. The dose was subsequently reduced in incremental steps to 45 microg/m(2) due to unexpected toxicity. RESULTS: The maximum tolerated dose of SJG-136 was 45 microg/m(2). The main drug-related adverse event was vascular leak syndrome (VLS) characterized by hypoalbuminemia, pleural effusions, ascites, and peripheral edema. Other unexpected adverse events included elevated liver function tests and fatigue. The VLS and liver toxicity had delayed onset and increased in severity with subsequent cycles. Disease stabilization was achieved for >6 weeks in 10 patients; in 2 patients this was maintained for >12 weeks. There was no evidence of DNA interstrand cross-linking in human blood lymphocytes with the use of the comet assay. Evidence of DNA interaction in lymphocytes and tumor cells was shown through a sensitive gamma-H2AX assay. SJG-136 had linear pharmacokinetics across the dose range tested. CONCLUSIONS: SJG-136 was associated with dose-limiting VLS and hepatotoxicity when administered by short injection every 21 days. DNA damage was noted, at all dose levels studied, in circulating lymphocytes. The etiology of the observed toxicities is unclear and is the subject of further preclinical research. Alternative clinical dosing strategies are being evaluated.