Thesis (Ph. D.)--University of Texas at Austin, 2008. / Vita. Includes bibliographical references.

Background: In years 2011-2014, 8.6 million sports-related injuries were reported each year and falls have been identified as a main cause. Purpose: To determine the impact of foot type, quadriceps angle, and Vibram™ footwear on postural stability. Methods: Twentyour males (age 21.38±2.50yr; height 1.74±0.06m; mass 71.24±10.37kg) were categorized as pronated, supinated, and neutral feet using FPI and bilateral quadriceps angles were measured. Participants were tested on barefoot, Vibram™ Bikila and Vibram™ Trek (VT), on stable/unstable, bilateral/unilateral, eyes open/closed conditions. Sway variables were analyzed using 3(foot type) × 3(footwear) repeated measures ANOVA. Pearson product correlation was performed for quadriceps angle with sway variables. Results: Footwear main effect significance was evident in all conditions except stable unilateral eyes open condition, with lower values for barefoot followed by VT. Conclusion: Static balance in BF is superior to shod conditions in all situations except the extremely challenging condition, in which VT showed greater balance.

No description available.

The liver is the largest internal organ and performs a wide variety of functions to maintain organismal homeostasis. While some liver functions are carried out by all hepatocytes, other functions are restricted to certain populations of hepatocytes within the liver. This phenomenon, called zonal gene regulation or liver zonation, controls may metabolic processes within the liver including ammonia detoxification; glucose homeostasis; bile acid and glutamine synthesis; and metabolism of xenobiotics, lipids, and amino acids. The liver also expresses many genes in a developmental or sex-biased manner. Some genes are expressed at higher levels early or late in development, or alternatively, in male or female liver. Several years ago, our lab identified a transcription factor called Zinc finger and homeoboxes 2 (Zhx2) based on its ability to control the silencing of genes that are normally expressed in the fetal liver. Zhx2 belongs to a small gene family that also includes Zhx1 and Zhx3. These four exon genes have a rather unique structure in that their entire protein coding region is located on an unusually large third exon. Preliminary studies indicate that these proteins are found only in vertebrates. I have performed a comprehensive analysis of Zhx proteins across a number of chordate species to determine their relationship throughout chordate evolution. Using multiple sequence alignment and phylogenetic tree-building, my studies have found that the primordial Zhx gene is most related to Zhx3 and that this gene exists in lower chordates including lancelet, sea squirt, and sea lamprey. Additional studies from our lab showed that Zhx2 regulates numerous hepatic genes in the adult liver, including cytochrome p450 (Cyp) genes as well as other genes that exhibit sex-biased expression. Previous studies have demonstrated that female-biased expression of Cyp2a4, is controlled, in part, by Zhx2. I have extended these studies to perform a comprehensive analysis of Cyp2a4 and the highly related Cyp2a5 gene. Despite the high similarity of these two Cyp genes, my data indicate that these genes exhibit different zonal expression patterns and are differentially regulated in the regenerating liver. In the course of these studies, I discovered and characterized antisense transcripts for both Cyp2a4 and Cyp2a5. Both Cyp2a4as and Cyp2a5as have positively correlated expression patterns compared to their sense counterparts. In contrast to Cyp2a4 and Cyp2a5, Cyp2a4as and Cyp2a5as show sex-biased expression patterns earlier in development, suggesting that they might contribute to later sex-biased patterns established for Cyp2a4 and Cyp2a5.

In this thesis I address several topics concerning the interaction of convection and density stratification in oceans and lakes. I present experimental and theoretical investigations of the interaction between a localized buoyancy source and a heat flux through a horizontal boundary, and of the interactions between salt fingers and intermittent turbulence or shear. ¶ An extensive series of laboratory experiments were used to quantify the stratification and circulation that result from the combined presence of a localized buoyancy source and a heat flux through a horizontal boundary. Previous studies found that convection in the form of a turbulent buoyant plume tends to produce a stable density stratification, whereas the distributed flux from a horizontal boundary tends to force vigorous overturning and to produce well-mixed layers. A new result of this thesis is that a steady density profile, consisting of a mixed layer and a stratified layer, can exist when the plume buoyancy flux is greater than the distributed flux. When the two fluxes originate from the same boundary, the steady state involves a balance between the rate at which the mixed layer deepens due to entrainment on the one hand and vertical advection of the stratified water far from the plume (due to the volume flux acquired by entrainment) on the other hand. There is a monotonic relationship between the normalized mixed layer depth and flux ratio R (boundary flux/plume flux) for 0 < R > 1, and the whole tank overturns for R > 1. The stable density gradient in the stratified region is primarily due to the buoyancy from the plume and for R > 0 there is a small increase in the gradient due to entrainment of buoyancy from the mixed layer. For the case of fluxes from a plume located at one boundary and a uniform heat flux from the opposite boundary the shape of the density profile is that given by Baines & Turner (1969), with the gradient reduced by a factor (1 + R) due to the heating. Thus, when R < - 1 there is no stratified region and the whole water column overturns. When 0 > R > - 1, the constant depth of the convecting layer is determined by the Monin-Obukhov scale in the outflow from the plume. ¶ One application of these laboratory experiments is to surface cooling in lakes and reservoirs that have shallow sidearms. During prolonged periods of atmospheric cooling, gravity currents can form in these sidearms and as the currents descend into the deeper waters they are analogous to isolated plumes. This can result in stratification at the base of a lake and an upwelling of cold water. Away from the shallow regions, surface cooling leads to a mixed surface layer. The depth of this layer will be steady when the rate of upwelling balances the rate at which the mixed layer deepens by turbulent entrainment. A series of laboratory experiments designed to model the depth distribution of a lake with a shallow sidearm showed that the steady depth of the mixed layer depended on the ratio of the area of the shallow region to the area of the deep region. Significant stratification resulted only when the reservoir had shallow regions that account for more than 50 % of the surface area. The depth of the surface mixed layer also depended on the ratio of the depths of the shallow and deep regions and no significant stratification forms if this ratio is greater than 0.5. These results are in good agreement with observations of circulation and stratification during long periods of winter cooling from Chaffey reservoir, Australia. Theoretical time scales are also developed to predict the minimum duration of atmospheric cooling that can lead the development of stratification. ¶ In the second part of this thesis, I report a series of laboratory experiments which are designed to investigate the fine structure and buoyancy fluxes that result from salt finger convection in the presence of shear and intermittent turbulence. We find that, when salt finger convection in deep linear gradients is superposed with a depth-dependent spatially periodic shear, variations in the density profile develop on the same wavelength as the shear. The laboratory experiments presented in this thesis were carried out in a continuous density gradient with a spatially periodic shear produced by exciting a low-frequency baroclinic mode of vertical wavelength 60 mm. The density gradient consisted of opposing salt and sugar gradients favourable to salt fingers (an analogue to the oceanic heat/salt system). Where the shearing was large the salt finger buoyancy fluxes were small. Changes in salinity gradient due to the resulting flux divergence were self-amplifying until a steady state was reached in which the spatial variations in the ratio of salt and sugar gradients were such that the flux divergence vanished. Thus, along with reducing the mean salt finger buoyancy flux, a spatially varying shear can also lead to the formation of density structure. ¶ In the ocean intermittent turbulence can occur in isolated patches in salt finger-favourable regions. I present new results from laboratory experiments in which a partially mixed patch was produced in deep linear concentration gradients favourable to salt finger convection. Salt fingers give rise to an “up gradient” flux of buoyancy which can reduce the density gradient in a partially mixed patch. This can then lead to overturning convection of the partially mixed patch if a) the ratio of T and S gradients, R\rho =aTz/_ /betaSz, is near one, b) if turbulence results in a nearly well-mixed patch and c) the patch thickness is large enough that convective eddies are able to transport T and S faster than salt fingers. Once overturning occurs, subsequent turbulent entrainment can lead to growth of the patch thickness. Experimental results agree well with the theoretical prediction that h= \surd 8h B/N2 t, where h is the patch thickness, t is time, h is the mixing efficiency of turbulent entrainment, B is the buoyancy flux of the salt fingers and N is the buoyancy frequency of the ambient gradient region. This thickening is in contrast to the collapse that a partially mixed patch would experience due to lateral intrusion in a very wide tank. In regions of the ocean that contain salt fingers there is the possibility that, after a period of initial collapse, an intrusion could enter a regime where the rate of collapse in the vertical is balanced by the growth rate due to turbulent entrainment from the salt fingers buoyancy flux, thus tending to maintain the rate of lateral spread. ¶ A further series of laboratory experiments quantified the buoyancy fluxes that result from salt fingers and intermittent turbulence. A continuous density gradient, favourable to salt finger convection, was stirred intermittently by an array of vertical rods that move horizontally back and forth along the tank at a constant velocity. Previous experiments had found that continuous turbulence destroys any salt fingers present because the dissipation of turbulent kinetic energy occurs at scales that are generally smaller than salt fingers widths. However, when turbulence is present only intermittently, the salt fingers may have time to grow between turbulent events and so contribute to the vertical diffusivities of heat and salt. We conclude that the vertical buoyancy flux of salt fingers is strongly dependent upon the intermittency of the turbulence, and equilibrium fluxes are only achieved if the time between turbulent events is much greater than the e-folding time of the salt fingers. When these results are applied to an oceanographic setting, the effect of intermittent turbulence, occurring more 5% of the time, is to reduce the effective eddy diffusivity due to salt fingers below equilibrium salt finger values, so that at R\rho > > 2 the eddy diffusivity is due only to turbulence. The time averaged salt fingers fluxes are not significantly reduced by intermittent turbulence when R\rho > 2 or if the intermittence occurs less than 2% of the time, and so may contribute significant diapycnal fluxes in many parts of the ocean.

The plant homeodomain (PHD) is a protein domain of ~45�100 residues characterised by a Cys4-His-Cys3 zinc-binding motif. When we commenced our study of the PHD in 2000, it was clear that the domain was commonly found in proteins involved in transcription. Sequence alignments indicate that while the cysteines, histidine and a few other key residues are strictly conserved, the rest of the domain varies greatly in terms of both amino acid composition and length. However, no structural information was available on the PHD and little was known about its function. We were therefore interested in determining the structure of a PHD in the hope that this might shed some light on its function and molecular mechanism of action. Our work began with the structure determination of a representative PHD, Mi2b-P2, and this work is presented in Chapter 3. Through comparison of this structure with the two other PHD structures that were determined during the course of our work, it became clear that PHDs adopt a well-defined globular fold with a superimposable core region. In addition, PHDs contain two loop regions (termed L1 and L3) that display increased flexibility and overlay less well between the three PHD structures available. These L1 and L3 regions correspond to variable regions identified earlier in PHD sequence alignments, indicating that L1 and L3 are probably not crucial for the PHD fold, but are instead likely to be responsible for imparting function(s) to the PHD. Indeed, numerous recent functional studies of PHDs from different proteins have since demonstrated their ability in binding a range of other proteins. In order to ascertain whether or not L1 and L3 were in fact dispensable for folding, we made extensive mutations (including both insertions and substitutions) in the loop regions of Mi2b-P2 and showed that the structure was maintained. We then went on to illustrate that a new function could be imparted to Mi2b-P2 by inserting a five-residue CtBP-binding motif into the L1 region and showed this chimera could fold and bind CtBP. Having established that the PHD could adopt a new binding function, we next sought to use combinatorial methods to introduce other novel functions into the PHD scaffold. Phage display was selected for this purpose, because it is a well-established technique and has been used successfully to engineer zinc-binding domains by other researchers. However, in order to establish this technique in our laboratory, we first chose a control system in which two partner proteins were already known to interact in vitro. We chose the protein complex formed between the transcriptional regulators LMO2 and ldb1 as a test case. We have examined this interaction in detail in our laboratory, and determined its three-dimensional structure. Furthermore, inappropriate formation of this complex is implicated in the onset of T-cell acute lymphoblastic leukemia. We therefore sought to use phage display to engineer ldb1 mimics that could potentially compete against wild-type ldb1 for LMO2, and this work is described in Chapter 4. Using a phage library containing ~3 x 10 7 variants of the LMO2-binding region of ldb1, we isolated mutants that were able to interact with LMO2 with higher affinity and specificity than wild-type ldb1. These ldb1 mutants represent a first step towards finding potential therapeutics for treating LMO-associated diseases. Having established phage display in our laboratory, we went on to search for PHD mutants that could bind selected target proteins. This work is described in Chapter 5. We created three PHD libraries with eight randomized residues in each of L1, L3 or in both loops of the PHD. These PHD libraries were then screened against four target proteins. After four rounds of selection, we were able to isolate a PHD mutant (dubbed L13-FH6) that could bind our test protein Fli-ets. This result demonstrates that a novel function can be imparted to the PHD using combinatorial methods and opens the way for further work in applying the PHD scaffold to other protein design work. In summary, the work detailed in Chapters 3 and 5 demonstrates that the PHD possesses many of the properties that are desirable for a protein scaffold for molecular recognition, including small size, stability, and a well-characterised structure. Moreover, the PHD motif possesses two loops (L1 and L3) of substantial size that can be remodeled for target binding. This may lead to an enhancement of binding affinities and specificities over other small scaffolds that have only one variable loop. In light of the fact that PHDs are mainly found in nuclear proteins, it is reasonable to expect that engineered PHDs could be expressed and function in an intracellular environment, unlike many other scaffolds that can only function in an oxidizing environment. Therefore, our results together with other currently available genomic and functional information indicate PHD is an excellent candidate for a scaffold that could be used to modify cellular processes. Appendices 1 and 2 describe completed bodies of work on unrelated projects that I have carried out during the course of my PhD candidature. The first comprises the invention and application of DNA sequences that contain all N-base sequences in the minimum possible length. This work is presented as a reprint of our recently published paper in Nucleic Acids Research. The second Appendix describes our structural analysis of an antifreeze protein from the shorthorn sculpin, a fish that lives in the Arctic and Antarctic oceans. This work is presented as a manuscript that is currently under review at the Journal of the American Chemical Society.

Thesis (Ph. D.)--University of Washington, 2000. / Vita. Includes bibliographical references (leaves 70-81).

Thesis (Ph. D.)--University of Washington, 2002. / Vita. Includes bibliographical references (leaves 83-97).

Thesis (M.A.(Biokinetics, Sport and Leisure Sciences))--University of Pretoria, 2002. / Summary in English and Afrikaans. Includes bibliographical references.

abstract / Biochemistry / Doctoral / Doctor of Philosophy