Detection and Analysis of Protein Aggregation with Confocal Single Molecule Fluorescence Spectroscopy

The misfolding and aggregation of proteins is a common phenomenon both in the cell, in in vitro protein refolding, and the corresponding biotechnological applications. Most importantly, it is involved in a wide range of diseases, including some of the most prevalent neurodegenerative disorders. However, the range of methods available to analyze this highly heterogeneous process and the resulting aggregate structures has been very limited. Here we present an approach that uses confocal single molecule detection of FRET-labeled samples employing four detection channels to obtain information about diffusivity, anisotropy, fluorescence lifetimes and Förster transfer efficiencies from a single measurement. By combining these observables, this method allows the separation of subpopulations of folded and misfolded proteins in solution with high sensitivity and a differentiation of aggregates generated under different conditions. We demonstrate the versatility of the method with experiments on rhodanese, an aggregation-prone two-domain protein.     

Hydroxyl radical as a probe of the charge distribution in aromatics: phenol

Initial radiation chemical yields of 1.48 (2), 0.24 (2), and 2.01 molecules per 100 eV of absorbed energy are reported for addition of *OH radical to each of the ortho, meta, and para positions of phenol. These yields represent 91% of the yield of 5.96 expected for *OH addition to 5 mM phenol and are in general agreement with other previous measurements. Pulse radiolysis experiments show that phenoxy radical is produced in a yield of approximately 0.42 as a result of addition of *OH at phenol's ipso position. The total of these yields (5.84) accounts for the addition of virtually all of the expected *OH radicals. The relative yields for addition to the ortho, meta, and para positions provide a measure of the charge distribution in phenol that correlates quite well with the unpaired spin distribution in phenoxyl radical. This correlation indicates that the OH substituent similarly affects the charge distribution on the aromatic ring of phenol and the unpaired spin distribution in the phenoxyl radical.     

Mammographic feature enhancement by multiscale analysis

Introduces a novel approach for accomplishing mammographic feature analysis by overcomplete multiresolution representations. The authors show that efficient representations may be identified within a continuum of scale-space and used to enhance features of importance to mammography. Methods of contrast enhancement are described based on three overcomplete multiscale representations: 1) the dyadic wavelet transform (separable), 2) the phi-transform (nonseparable, nonorthogonal), and 3) the hexagonal wavelet transform (nonseparable). Multiscale edges identified within distinct levels of transform space provide local support for image enhancement. Mammograms are reconstructed from wavelet coefficients modified at one or more levels by local and global nonlinear operators. In each case, edges and gain parameters are identified adaptively by a measure of energy within each level of scale-space. The authors show quantitatively that transform coefficients, modified by adaptive nonlinear operators, can make more obvious unseen or barely seen features of mammography without requiring additional radiation. The authors' results are compared with traditional image enhancement techniques by measuring the local contrast of known mammographic features. They demonstrate that features extracted from multiresolution representations can provide an adaptive mechanism for accomplishing local contrast enhancement. By improving the visualization of breast pathology, one can improve chances of early detection while requiring less time to evaluate mammograms for most patients.     

The Synthesis and STM/AFM Imaging of ‘Olympicene’ Benzo[cd]pyrenes

H‐Benzo[cd]pyrene (‘Olympicene′) is a polyaromatic hydrocarbon and non‐Kekulé fragment of graphene. A new synthetic method has been developed for the formation of 6H‐benzo[cd]pyrene and related ketones including the first time isolation of the unstable alcohol 6H‐benzo[cd]pyren‐6‐ol. Molecular imaging of the reaction products with scanning tunnelling microscopy (STM) and non‐contact atomic force microscopy (NC‐AFM) characterised the 6H‐benzo[cd]pyrene as well as the previously intangible and significantly less stable 5H‐benzo[cd]pyrene, the fully conjugated benzo[cd]pyrenyl radical and the ketones as oxidation products.     

Cu2+ in liquid ammonia—The impact of solvent flexibility and electron correlation in ab initio quantum mechanical charge field molecular dynamics

The impact of solvent flexibility and electron correlation on the simulation results of Cu²⁺ in liquid ammonia has been investigated via an ab initio quantum mechanical charge field molecular dynamics (QMCF MD) simulation approach. To achieve this, three different simulation systems were considered in this study, namely Cu²⁺ in rigid and flexible ammonia at Hartree-Fock (HF) level of theory, as well as resolution of identity second order Møller-Plesset (MP2) perturbation theory in the rigid body case. In all cases, a stable octahedral [Cu(NH₃)₆]²⁺ complex subject to dynamic Jahn-Teller distortions without the occurrence of ligand exchange was observed. The Cu²⁺ − NH₃ distance in the first shell agrees well with the experimental and other theoretical data. In all three cases, the structural data shows that the rigid-body ammonia model in conjunction with the HF level of theory provides accurate data for the first solvation shell, while at the same time, the computational demand and thus the achievable simulation time are much more beneficial. The vibrational analysis of the Cu²⁺ − NH₃ interaction yields similar force constants in the three investigated systems indicating that there is no distinct difference on the dynamical properties of the first solvation shell. In addition to the QMCF MD simulations, a number of natural bond orbital (NBO) analyses were carried out, confirming the strong electrostatic character of the Cu²⁺ − NH₃ interaction.     

Measurement and modeling of a new width dependence of nmosfet degradation

NMOS transistors with widths between 1.2μm and 10μm and length of 0.8μm have been stressed for up to 5000 hours. Investigating the threshold voltage shift a new width dependence of degradation has been measured, analysed and modeled by a simple theory. Because of the increasing degradation of NMOSFETs with decreasing width this effect will be more and more important for small-channel LOCOS transistors.     

Generalized Airy Stress Functions

The classical Airy stress function in planar elastostatics cannot, in general, be a smooth function for multiply connected domains. Moreover, if a non-null body force field is active the classical Airy representation for the stress is not complete. Here, a generalized Airy representation for the stress is presented which preserves smoothness and which is complete. The generalized form identifies the explicit additional pieces that are needed for completeness in multiply connected domains and when a body force field is present.     

Shock pulse attenuation in a nonlinear viscoelastic solid

Thin, one-dimensional shock pulses were generated in a nonlinear viscoelastic material (polymethyl methacrylate) by a new experimental technique. The observed pulse attenuation was compared with an approximate theory based on the viscoelastic shock amplitude equation. The central assumption of this approximate theory is that the unloading wave propagates as a simple wave. Given an initial pulse shape it is shown that the attenuation and the pulse shape at any later time are accurately approximated. The calculated attenuation in polymethyl methacrylate agreed well with the experimental results.