As genome sequencing projects begin to come to completion, the challenge becomes one of determining how to understand the information contained within the DNA. DNA is a polymorphic macromolecule; the A- B- and Z-DNA conformations have been observed by a variety of physical techniques. The magnitude of the energetic differences between these conformations suggests that these conformations may be important biologically and thus relevant in the analysis of genomes. A computer program, NASTE, was developed to evaluate the helical parameters of the set of Z-DNA crystal structures in order to determine the true conformation of Z-DNA and to understand the effects of various factors on the observed structure and stability. A thermodynamic method, elucidated in part with a genetic algorithm, was developed to predict the sequence-dependent propensity of DNA sequences for A- versus B-DNA in both the crystal and in natural DNA. Predictions from this method were tested by studying the conformation of short oligonucleotides using circular dichroism spectroscopy. Finally, the thermodynamic method was applied in an algorithm, AHUNT, to identify regions in genomic DNA with a high propensity to form A-DNA. Significant amounts of A-DNA were identified in eukaryotic and archeabacterial genes. E. coli genes have less A-DNA than would be predicted from their (G+C) content. These results are discussed with respect to the intracellular environment of the genomes. / Graduation date: 1998

Graduation date: 1997

Graduation date: 1996

Graduation date: 1994

Graduation date: 1994

Living cells are dependent upon protein synthesis for virtually all cellular functions. The cellular machine responsible for protein synthesis, called the ribosome, is formed through the association of two unequally sized subunits, each composed of RNA and proteins. Proper assembly of each subunit is essential to ribosome function and therefore essential to the cellular life cycle. Previous studies focused on dissecting the assembly of the small ribosomal subunit (30S subunit) from E. coli have shown that 21 proteins sequentially assemble on the 16S rRNA at multiple nucleation sites. For the first time, we are able to monitor changes in the secondary and tertiary structure of the 16S rRNA upon the addition of single proteins during assembly by using time-dependent chemical probing. Results from these studies suggest that protein S17 induces multiple structural changes in 16S rRNA by first binding to helix 11 and then helix 7. S20 also induces changes in the rRNA by interacting with helix 9, 11, 44 and 13 in that order. These structural formations and rearrangements then prepare the binding sites for additional proteins (S12 and S16, respectively). This study demonstrates that time-dependent chemical probing is able to monitor the assembly of the 30S subunit at a level of detail never before seen. These studies also suggest that many motifs in the 16S rRNA structure are formed as a result of the proteins binding, lending evidence to the hypothesis that the function of ribosomal proteins is to shape and/or hold the RNA structure in place.

Detailed geologic mapping of portions of the Banoak, Reepsville, Lincolnton West, and Cherryville 7.5-minute quadrangles has confirmed the easternmost exposure of the Brindle Creek fault, which frames the Newton window. The Brindle Creek fault is a terrane boundary that separates the overlying Siluro-Devonian assemblage of metasedimentary rocks and Devonian-Mississippian anatectic plutons of the Cat Square terrane from the Neoproterozoic(?)-Ordovician metasedimentary and igneous rocks of the Tugaloo terrane. Structures related to six deformational events have been identified in this portion of the Inner Piedmont. The Brindle Creek fault has been folded multiple times, resulting in a sinuous outcrop pattern and the formation of the Newton window and smaller Howards Creek window. Portions of three map-scale sheath folds have been identified by map patterns and orientation of dominant mineral lineations, fold axes, and shear-sense indicators. The discontinuity of map-scale bodies of metagraywacke, mafic complexes, and amphibolite is attributed to extension during sheath fold formation. Dominant foliation, mineral lineation, and fold-axis orientations suggest north-northwest directed flow occurred in this portion of the Inner Piedmont. Zircon geochronology data indicate crystallization of the Vale charnockite at 366.4 ± 3.1 Ma and the enclosing Walker Top Granite at 356.5 ± 5.3 Ma. Zircon saturation thermometry estimates minimum magmatic temperatures for the granitoids at 800-840⁰ C. Whole-rock geochemical and isotopic data indicate the Vale charnockite and Walker Top Granite are genetically related and were derived from deep crustal melting of largely Proterozoic-affinity metasediments in an arc environment. Both granitoids crystallized from the same parent magma; the Vale charnockite is an autolith, or early crystallization of the melt, incorporated into the later crystallizing Walker Top Granite. Geochemical analyses of Cat Square terrane mafic complexes west of the Newton window suggest these bodies represent vestiges of oceanic crust formed in a back-arc basin setting or from both MORB and volcanic-arc sources as characterized by mixed N-MORB and calc-alkaline volcanic-arc signatures. This back-arc basin likely formed from east-dipping subduction during the development of Ordovician volcanic arcs outboard of Laurentia.

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The 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 β-lactamase (BlaC) and fatty-acyl-CoA thioesterase (FcoT). Mtb β-lactamase has been proposed to be the most significant reason for mycobacterial resistance to β-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 β-lactamase inhibitors that could be used with β-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.

The large-scale environment can have a significant impact on subtropical precipitating systems via the baroclinicity of the environment and the associated dynamical forcings. The degree of baroclinicity is examined using National Centers for Environmental Prediction (NCEP) reanalysis temperature and zonal wind fields over a two-year period for Southeast Texas, yielding classifications of barotropic, weakly baroclinic, and strongly baroclinic for the background environment. Weakly baroclinic environments accounted for half of the days throughout the two-year period. Barotropic environments occurred most frequently during summer and strongly baroclinic environments occurred most frequently in winter, although less often than weakly baroclinic environments. A climatology of storm types, based on dynamical forcing (i.e., weak forcing, drylines, cold fronts, warm fronts, stationary fronts and upper level disturbances) and precipitation structure (i.e., isolated, scattered, widespread, linear, unorganized and leading-line/trailing stratiform), was compiled and compared to the baroclinicity designations. Non-frontal storm types (i.e., weak forcing, drylines and upper level disturbances) are typical of barotropic environments, while frontal storm types (i.e.,warm, cold and stationary fronts) are typical of weakly and strongly baroclinic environments. Storm events and drop-size distributions (DSD) were identified from surface rainfall data collected by a Joss-Waldvogel disdrometer located in College Station, Texas. The DSDs were compared by baroclinicity and storm type. The barotropic DSD is weighted towards the largest drops because of the stronger convection and stratiform precipitation in the weak forcing and dryline storm types, while the strongly baroclinic DSD is weighted towards the smallest drops because of the weaker convection from the warm fronts and stationary fronts. The weakly baroclinic DSD is weighted more evenly towards small and large drops than the barotropic and strongly baroclinic DSDs because of the conflicting microphysical processes in the different storm types. The microphysical processes within the storms vary by storm type and baroclinicity regime, such that the large-scale environment modifies the precipitation characteristics of storms in SE Texas.