DNA Binding Kinetics of Large Antiviral Hairpin Polyamides

While vaccines exist for the some of the most problematic strains of human papillomavirus (HPV), a double stranded DNA virus, there is currently no cure. HPV remains one of the most commonly sexually transmitted infections and is responsible for virtually all cervical cancers and genital warts. Natural products Distamycin A and netropsin have inspired the hairpin Nmethylpyrrole (Py)/N-methylimidazole (Im) polyamides (PAs) studied here. The larger hairpin PAs, designed to bind to sites of 10 or more DNA bp, have been shown to be effective antivirals against oncogenic HPV strains 16, 18, and 31, while smaller hairpin PAs are not. Despite significant differences in potencies among the antiviral PAs tested, the PAs bind to DNA with similar binding affinity (Kd). Ample evidence has shown dissociation rate constants (koff) may be a better indicator of drug efficacy than Kd. While Kd is a function of association rate constant (kon) and koff, respectively (Kd = koff/kon), few studies have focused on the DNA binding kinetics of large hairpin PAs. We are using fluorescence and CD spectroscopy to characterize Kds, obtain DNA binding kinetic rate constants, and determine binding stoichiometries as a function of PA size. K= remains tight (low nM) for all PAs tested (6-20-rings) with our fluorescence assay, which is consistent with what is seen with other methods. The large PAs are characterized by slow DNA dissociation rates with half-lives ranging from 20-30 min, and dissociation slows as the size of PAs increases. Slow dissociation rates are likely the source of the difference in antiviral behavior. Association time courses for a 14-ring PA indicate that \u3e1 equivalent of PA is binding to the DNA. Further supporting this claim, fluorescence and CD spectroscopic experiments indicate that saturation of the DNA does not occur until 2 or more equivalents of PA are added. The slow dissociation of multiple equivalents of PA bound to DNA may cause a large and prolonged disruption in DNA conformation, which in turn elicits or alters the DNA damage response, which is an integral part in the antiviral mechanism of large antiviral hairpin PAs and of the lifecycle of HPV

Создание функциональной керамики на основе оксинитридов алюминия и кремния

Объектом исследования является керамика на основе оксида алюминия, оксида кремния, диоксида циркония, нитрида бора, нитрида кремния. Целью работы является исследование структуры и физико-механических свойств керамики на основе оксида алюминия, диоксида кремния, диоксида циркония, нитрида бора, нитрида кремния, полученной прессованием и спеканием наноструктурных порошков. В процессе исследования изучали технологические свойства оксидных порошков, проводили их механическую активацию, пластификацию, формовали и спекали керамические прессовки, изготавливали микрошлифы спеченных образцов для наноиндентирования и изучения микроструктуры. Были определены плотность прессовок и спечённых образцов, измерены модуль упругости и нанотвердость с использованием методики наноиндентрования.The object of the study is ceramics based on aluminum oxide, silicon dioxide, zirconium dioxide, boron nitride, silicon nitride. The aim of this work is to study the structure and physical and mechanical properties of ceramics based on aluminum oxide, silicon dioxide, zirconium dioxide, boron nitride, silicon nitride obtained by pressing and sintering nanostructured powders. In the process of research, the technological properties of oxide powders were studied, their mechanical activation, plasticization, molding and sintering of ceramic compacts, microsections of sintered samples for nanoindentation and microstructure studying. The density of compacts and sintered samples was determined, the modulus of elasticity and nanohardness were measured using the nanoindentation technique

Numerical Simulation of Turbulent Flow Through a Straight Square Duct

Turbulent duct flows are investigated using large eddy simulation at bulk Reynolds numbers, from 4410 to 250,000. Mean secondary flow is found to reveal the existence of two streamwise counter-rotating vortices in each corner of the duct. Turbulence-driven secondary motions that arise in duct flows act to transfer fluid momentum from the centre of the duct to its corners, thereby causing a bulging of the streamwise velocity contours towards the corners. As Reynolds number increases, the ratio of centreline streamwise velocity to the bulk velocity decreases and all turbulent components increase. In addition, the core of the secondary vortex in the lower corner-bisector tends to approach the wall and the corner with increasing Reynolds number. The turbulence intensity profiles for the low Reynolds number flows are quite different from those for the high Reynolds number flows. Typical turbulence structures in duct flows are found to be responsible for the interactions between ejections from wall and this interaction results in the bending of the ejection stems, which indicates that the existence of streaky wall structures is much like in a channel flow

Investigation of low-dissipation monotonicity-preserving scheme for direct numerical simulation of compressible turbulent flows

© 2014 Elsevier Ltd. The influence of numerical dissipation on direct numerical simulation (DNS) of decaying isotropic turbulence and turbulent channel flow is investigated respectively by using the seventh-order low-dissipation monotonicity-preserving (MP7-LD) scheme with different levels of bandwidth dissipation. It is found that for both benchmark test cases, small-scale turbulence fluctuations can be largely suppressed by high level of scheme dissipation, while the appearance of numerical errors in terms of high-frequency oscillations could destabilize the computation if the dissipation is reduced to a very low level. Numerical studies show that reducing the bandwidth dissipation to 70% of the conventional seventh-order upwind scheme can maximize the efficiency of the MP7-LD scheme in resolving small-scale turbulence fluctuations and, in the meantime preventing the accumulation of non-physical numerical errors. By using the optimized MP7-LD scheme, DNS of an impinging oblique shock-wave interacting with a spatially-developing turbulent boundary layer is conducted at an incoming free-stream Mach number of 2.25 and the shock angle of 33.2°. Simulation results of mean velocity profiles, wall surface pressure, skin friction and Reynolds stresses are validated against available experimental data and other DNS predictions in both the undisturbed equilibrium boundary layer region and the interaction zone, and good agreements are achieved. The turbulence kinetic energy transport equation is also analyzed and the balance of the equation is well preserved in the interaction region. This study demonstrates the capability of the optimized MP7-LD scheme for DNS of complex flow problems of wall-bounded turbulent flow interacting with shock-waves

Turbulent flow through a rectangular channel

Streamwise and normal turbulent velocity measurements were taken at 80 normal locations across a two-inch high rectangular water channel. A two polarization, three beam He-Ne Laser-Doppler Velocimetry system was used. The water channel was specifically designed and constructed to facilitate these optical measurements. It had negligible vibration and flow oscillation. Its length was 200 channel heights, sufficient to produce fully developed turbulent flow. The optical system had a specially designed optical table and traverse; optical glass windows were used as viewing ports.Measurements were made of the first four moments of the velocity fluctuation and the first three moments of Reynolds stress. The turbulence production and the correlation coefficient were calculated. Velocity spectra were measured for both the normal and streamwise velocity fluctuations at four to nine normal locations. Measurements were taken as close as y\sp+ = 0.6 viscous units away from the wall. The experiments were conducted at Reynolds numbers of 2457, 2777, and 18,339. The experimental results were compared to the computer simulation channel flow by Kim, Moin, and Moser. Very good agreement was found.Measurements were also made of the differential surface pressure over two sinusoidal waves. Wave amplitudes of 0.0123 and 0.03125 inches were used. Both waves had two inch wavelengths. Viscous fluid was used to extend the range of the measurements over a much larger range of Reynolds numbers than had been previously reported. Non-linearities were found at low Reynolds numbers in the differential pressure distribution over the 0.03125 inch amplitude wave, indicating a quasi-separated or separated flow. The differential pressure distribution over the 0.0123 inch amplitude wave demonstrated a linear, sinusoidal response. The experimental data were compared to computer models developed in this laboratory and were found to demonstrate only fair agreement. This is not unexpected, since the computer models predict a linear response over all experimental Reynolds numbers examined.U of I OnlyETDs are only available to UIUC Users without author permissio

Interactions of two large antiviral polyamides with the long control region of HPV16.

PA1 and PA25 are large hairpin polyamides that are effective in nearly eliminating HPV16 episomes (DNA) in cell culture, and PA25 has broad spectrum activity against three cancer-causing forms of HPV (Edwards, T. G., Koeller, K. J., Slomczynska, U., Fok, K., Helmus, M., Bashkin, J. K., Fisher, C., Antiviral Res. 91 (2011) 177-186). Described here are the interactions of these PAs with sequences in the long control region (LCR) of HPV16 (7348-122). Using an FeEDTA conjugate of PA1 (designed to recognize 5’-W(2)GW(7)-3’; W = A or T), 34 affinity cleavage (AC) patterns were detected for this fragment. These sites can be rationalized with sequences featuring perfect, single, double, triple and quadruple mismatches. Quantitative DNase I footprinting analysis indicates that perfect sites bind PA1 with K(d)s between 0.7-2.2 nM. K(d)s for single, double, triple and quadruple mismatch sites range from 1-3 nM to 20 nM. Using AC and EDTA conjugates, we report that unlike smaller 8-ring hairpin PAs, introduction of a chiral turn in this large polyamide has no effect on binding orientation (forward vs. reverse). Despite its design to recognize 5’-W(2)GW(5)GW(4)-3’ via two Im residues, a motif not represented in this HPV sequence, a PA25-EDTA conjugate yielded 31 affinity cleavage sites on the region. Low nM K(d)s for PA25 without EDTA indicate a high tolerance for triple and quadruple mismatches. While there is extensive coverage of the sequence examined, AC cleavage patterns for the two PAs show discrete binding events and do not overlap significantly. This indicates that within the context of A/T rich sequences, these PAs do not recognize a simple shared sequence-related feature of the DNA. These insights continue to inform the complex nature of large hairpin PA-DNA interactions and antiviral behavior