Monitoring Alzheimer Amyloid Peptide Aggregation by EPR

Plaques containing the aggregated β-Amyloid (Aβ) peptide in the brain are the main indicators of Alzheimer’s disease. Fibrils, the building blocks of plaques, can also be produced in vitro and consist of a regular arrangement of the peptide. The initial steps of fibril formation are not well understood and could involve smaller aggregates (oligomers) of Aβ. Such oligomers have even been implicated as the toxic agents. Here, a method to study oligomers on the time scale of aggregation is suggested. We have labeled the 40 residue Aβ peptide variant containing an N-terminal cysteine (cys-Aβ) with the MTSL [1-oxyl-2,2,5,5-tetramethyl-Δ-pyrroline-3-methyl] methanethiosulfonate spin label (SL-Aβ). Fibril formation in solutions of pure SL-Aβ and of SL-Aβ mixed with Aβ was shown by Congo-red binding and electron microscopy. Continuous-wave 9 GHz electron paramagnetic resonance reveals three fractions of different spin-label mobility: one attributed to monomeric Aβ, one to a multimer (8–15 monomers), and the last one to larger aggregates or fibrils. The approach, in principle, allows detection of oligomers on the time scale of aggregation.     

Catalytically Active Cu(II)-Pybox Complexes: Insights by EPR Spectroscopy and DFT Computations

This paper addresses the geometry of the catalytic Cu(II) complexes formed in the course of stereoselective Diels–Alder reaction. The parent catalyst is [Cu(II)-isopropyl-pybox](OTf)₂. Ethyl glyoxylate serves as dienophile and it reacts with 1,3-cyclohexadiene. The reaction is followed by CW EPR, HYSCORE, and (pulsed) ENDOR spectroscopy and the experimental results are supported by DFT computations. It is shown that during the catalytic process, the Cu(II) complex is pentacoordinated and one of the triflate counterions is coordinated to the Cu(II) center in axial position.     

Solvation and Rotational Diffusion of Solutes in Room Temperature Ionic Liquids as Studied by EPR Spectroscopy with Nitroxide Spin Probing Method

In this article, we briefly introduced our studies on solvation and rotational diffusion of solutes in room temperature ionic liquids (RTILs) by electron paramagnetic resonance with nitroxide spin probing method. Most of the rotational correlation times for the nitroxide radicals are within the range calculated on the basis of Stokes–Einstein–Debye hydrodynamic theory with stick and slip boundary conditions or Gierer–Wirtz theory except for smaller solutes in some RTILs with smaller BF₄ and PF₆ anions. In RTILs with 1-butyl-3-methylimidazolium as cation and BF₄ or PF₆ as anion, nitroxide radicals undergo rotational diffusion like supercooled liquids and nitroxide radical with smaller volume rotationally slips.     

Photo-Induced Electron Spin Polarization in Chemical and Biological Reactions: Probing Structure and Dynamics of Transient Intermediates by Multifrequency EPR Spectroscopy

In this minireview, modern multifrequency electron paramagnetic resonance (EPR) spectroscopy, in particular, at high magnetic fields, is shown to provide detailed information about structure, motional dynamics and spin chemistry of transient radicals and radical pairs occurring in photochemical reactions. Examples discussed comprise spin-polarized radicals and radical pairs in disordered systems, such as ultraviolet-irradiated quinone and ketone compounds in fluid alcohol solutions, green-light initiated electron transfer in biomimetic porphyrin?Cquinone donor?Cacceptor model systems in frozen solution, aiming at artificial photosynthesis, and red-light initiated electron transfer in natural photosynthetic reaction-center protein complexes. The transient paramagnetic states exhibit characteristic electron polarization (CIDEP) effects originating from a triplet mechanism, a radical-pair mechanism or a correlated coupled radical-pair mechanism. They contain valuable information about structure and dynamics of the short-lived reaction intermediates. Moreover, the CIDEP effects can be exploited for signal enhancement. Continuous-wave and pulsed versions of time-resolved high-field EPR spectroscopy, such as transient EPR and electron spin-echo experiments, are compared with respect to their advantages and limitations for the specific photoreaction under study. Furthermore, orientation resolving W-band pulsed electron-electron double resonance (PELDOR) experiments on the spin-correlated coupled radical pair $ {\text{P}}_{865}^{ \cdot + } $ $ {\text{Q}}_{\text{A}}^{ \cdot - } $ in frozen solution reaction centers from the purple photosynthetic bacterium Rb. sphaeroides reveal details of distance and orientation of the pair partners in their charge-separated transient state. The results are compared with those of the ground-state P₈₆₅Q_A. In conjunction with Q-band proton electron-nuclear double resonance (ENDOR) experiments the W-band PELDOR results provide decisive evidence that the local structure of the Q_A binding site does not change under photoreduction of the quinone??in agreement with earlier FTIR studies. The examples given demonstrate that multifrequency EPR experiments on disordered systems add heavily to the capabilities of ??classical?? spectroscopic and diffraction techniques for determining structure?Cdynamics?Cfunction relations of biochemical processes, since short-lived intermediates can be observed in real time while staying in their working states at biologically relevant time scales.     

Study of Stable L-Alanine Radicals by W-Band EPR Spectroscopy

Stable L-alanine radicals, SAR1 and SAR2, induced by γ-irradiation of the L-alanine crystal have been investigated by electron paramagnetic resonance (EPR) technique at W-band (94 GHz) frequency. The study provides assignment of radical centers detected by continuous-wave EPR, saturation transfer mode and echo-detected field-swept EPR at W-band frequencies. The phase memory time, T _m, which was measured simultaneously at X-band (9.5 GHz) and W-band frequencies for different spectral components has been employed to estimate rotation correlation times of CH₃ protons and an effective correlation time related to the local dynamics of the entire SAR1 center at room temperature.     

Radiostability of Allantoin by Using Electron Spin Resonance Spectroscopy

The radiosterilization potential and dosimetric feature of allantoin were investigated through the molecular degradations produced after ultraviolet and gamma irradiation using electron spin resonance, infrared, and ultraviolet spectroscopies and thermal measurement techniques (differential thermal analysis and the glass transition temperature). Although ultraviolet-irradiated allantoin presents no electron spin resonance signal, gamma irradiation exhibited an electron spin resonance signal of triplet appearance. Room temperature and high-temperature line intensity and spectrum area data and their variations with applied microwave power, storage time, annealing time, annealing temperature, and applied radiation dose were analyzed by assuming the production of two different types of radicals having different spectroscopic and decay characteristics. Based on its relatively stable nature toward gamma and ultraviolet radiations, it was concluded that allantoin itself and the products containing it can be sterilized by gamma and/or ultraviolet radiations without creating a great loss in its beneficial effects in the allowed radiosterilization dose limits.     

Spin dynamics and internal motion in magnetically dilute manganites probed by EPR

EPR spectra of a series of single crystal samples of LaGa_1?xMn_xO₃ manganites were studied in a broad range of temperatures (20–300 K) and compositions (0.02≤x≤1). Evolution of the spectra under the action of exchange interactions of Mn³⁺ ions leads to the formation of a single Lorentzian line for x≥0.2. The results confirm the transition from antiferromagnetic to ferromagnetic spin ordering during diamagnetic dilution of the manganite with gallium, beginning with x=0.8. For x=0.1, the EPR spectra exhibit an anomalous broadening and splitting on cooling of the samples, which is interpreted as manifestations of a thermoactivated internal motion with a characteristic energy of about 50 meV. The results are compared with published data on the nuclear spin-lattice relaxation of gallium. The model of thermoactivated reorientations of the electron orbitals of Mn³⁺ ions subject to the Jahn-Teller effect is discussed.     

Kinetics of Surfactant-induced Aggregation of Lysozyme Studied by Fluorescence Spectroscopy

The study of protein conformational changes in the presence of surfactants and lipids is important in the context of protein folding and misfolding. In the present study, we have investigated the mechanism of the protein conformational change coupled with aggregation leading to size growth of Hen Egg White Lysozyme (HEWL) in the presence of an anionic detergent such as sodium dodecyl sulphate (SDS) in alkaline pH. We have utilized intrinsic protein fluorescence (tryptophan) and extrinsic fluorescent reporters such as 8-anilinonaphthalene-1-sulfonic acid (ANS), dansyl and fluorescein to follow the protein conformational change in real-time. By analyzing the kinetics of fluorescence intensity and anisotropy of multiple fluorescent reporters, we have been able to delineate the mechanism of surfactant-induced aggregation of lysozyme. The kinetic parameters reveal that aggregation proceeds with an initial fast-phase (conformational change) followed by a slow-phase (self-assembly). Our results indicate that SDS, below critical micelle concentration, induces conformational expansion that triggers the aggregation process at a micromolar protein concentration range.     

X-Ray Photoelectron Spectroscopy of Nanodiamonds Obtained by Grinding and Detonation Synthesis

Technical Physics - The initial composition and chemical state of nanopowder primary particles obtained by grinding natural diamond and detonation synthesis have been studied by X-ray photoelectron...     

EPR Spectroscopy Using Magnetic Field Gradient Modulated by a Triangular Wave

Magnetic field gradient modulation is one of the techniques to obtain an electron paramagnetic resonance (EPR) spectrum in a selected region of the sample. In this study, the magnetic field gradient modulation using a triangular wave was performed to overcome a problem during the sine wave modulation. Plastic materials were used for the bobbins and cases of the electromagnet to reduce the eddy current loss and drive the gradient coils in three-dimensional directions at a frequency of over 160 Hz. While the EPR signal splitting in a nonselected region, which is a problem in spectral analysis, was observed during the simulation and the actual measurement with the sine wave gradient modulation, the EPR signal broadening without splitting was observed in those with the triangular wave modulation. Thus, it is postulated that the triangular wave is more suitable than the sine one for the field gradient modulation. The spatial resolution was determined to be about 4 or 2 mm at the field gradient of 1 or 2 mT/cm, respectively. The separation of the EPR spectra of two types of radicals was also made by the triangular wave gradient modulation. Authors' address: Hidekatsu Yokoyama, Department of Pharmaceutical Sciences, International University of Health and Welfare, 2600-1, Kitakanemaru, Ohtawara 324-8501, Japan     

Spin polarization transfer during intramolecular triplet-triplet energy transfer as studied by time resolved EPR spectroscopy

Spin polarization conservation during intramolecular triplet-triplet energy transfer was studied for the phthalimide derivatives. It is shown that spin polarization transfer is useful for determining the conformation and the nature of the lowest triple (T₁) states of the donor and acceptor moieties. The polarization pattern of the acceptor triplet state was well reproduced taking into account the polarization of the donor and the mutual orientation of the donor and acceptor moieties. This technique clarified the order of the triplet sublevels in energy of the phthalimide chromophore. The T₁ states of stilbene derivatives, which have a low quantum yield of intersystem crossing under direct excitation, were also detected.     

EPR和NMR自旋标记法研究部分折叠蛋白质构象

部分折叠蛋白质的特征是在溶液中有部分二级结构,但是三级结构接触比较松散或者较少,其意义可能是蛋白质折叠过程的中间态,或者有重要的生理功能. 利用EPR和NMR波谱可以表征其构象特征,从而构建其二级结构、三级结构、四级结构以及构象变化的结构模型. 利用分子生物学的点突变技术可以在蛋白质主链上插入1个或2个半胱氨酸残基,然后把顺磁自旋标记物特异地共价结合在半胱氨酸的侧链巯基上来制备自旋标记样品. 位点特异性自旋标记电子顺磁共振（SDSL-EPR）波谱是通过测定2个自旋标记间的偶极相互作用,从而推测2个硝基氧自由基间的距离. 核磁共振（NMR）波谱则是通过测定单个自旋标记中心对周围原子核驰豫效应增强（PRE）的效应,推测出顺磁中心相对于周围原子核的距离. 连续波EPR和NMR自旋标记方法可以测定2.5 nm左右的偶极相互作用距离,属于长程的距离约束,这对于确定部分折叠蛋白质的构象至关重要. 该文将就蛋白质部分折叠态研究的生物学意义,自旋标记方法以及EPR和NMR方法研究其构象特征举例描述.     

Drying Characteristics-Property Correlationships in Gas-Fired Infrared Drying of Paper Obtained by Diffuse Reflectance FT-IR Spectroscopy

Introduction

Infrared (IR) drying of paper in a pilot scale has been investigated by several researchers in the past [1–3]. Both high-intensity electric heating and medium-intensity gas heating are now being used industrially either to preheat wet paper before conventional steam drying [1] or as a moisture-profile corrector just before paper sizing press [2]. Our own investigation [3] on the use of radiant energy from a gas-fired IR radiator have revealed that the drying efficiency of IR radiation is influenced by the nature of association of water molecules with cellulose in wet paper. Depending on the total moisture content of a paper, the free and bound moisture contents of paper will vary in wet paper sheet. Our previous study [3] has also confirmed that it is possible to remove free moisture more efficiently using gas-fired IR emitter operated at 1373 K rather than as 1223 K. It is reported that besides emitter temperature, the efficiency of drying also depends on the location of the emitter. In general, the efficiency is high if the emitter is installed near the preheating zone and the efficiency changes in the following order: falling rate < constant rate < preheating. Pikulik [4] compared the efficiency of hot gas impinging drying with conventional steam drying emphasizing the paper properties. The properties of a high-intensity hot gas-dried paper has been reported to be better than those of conventionally steam dried paper. An immediate quality improvement in terms of reduced moisture steaks and superior resistance to paper embrittleness was also observed when installing a high-intensity infrared drying system [5,6].     

利用双色激光感生光栅光谱法测定^127I的核自旋

本文利用ＮｄＹＡＧ激光倍频光５３２ｎｍ激光分成强度大致相等的两束激光作为泵浦光以２．７３°交叉于碘分子样品室中，两束泵浦光发生干涉在碘分子中选择激发形成了空间正弦分布的激光态分子光栅和基态耗尽型光栅。另一束窄线宽染料激光作为探索光射入到激光感生光栅上，沿着布拉格衍射的方向接收信号光。利用泵浦共振为３２－０Ｒ（５５），探索共振为１３－０Ｐ（５５），Ｒ（５５）和泵浦共振为３２－０Ｐ（５２），探索共振为１３－０Ｐ（５２），Ｒ（５２）的碘分子双色激光感生光栅光谱谱线强度测定了１２７Ｉ的核自旋，测定值为５／２。     

Influence of C60 Aggregation on Pseudorotation in the Excited Triplet State Probed by Multifrequency Time-Resolved EPR

The influence of C₆₀ aggregation on time-resolved (TR) electron paramagnetic resonance (EPR) of C₆₀ in the excited triplet state was investigated by multifrequency EPR techniques. Temperature-independent X-band (9.7 GHz) TR-EPR spectra were observed in a fresh toluene solution, while temperature-dependent ones were reported in literatures. The experimental spectra in this study indicated that the pseudorotation of pristine C₆₀ in frozen toluene solution is not frozen out even at lower temperatures. Careful investigations of TR-EPR and its decay kinetics demonstrated that the pseudorotation can be affected by C₆₀ aggregation. A comparison between X- and W-band (94.9 GHz) results indicated that the aggregation can be accelerated by a capillary effect. Three decay constants were extracted from the analysis of the decay kinetics. The fastest component was ascribed to the pseudorotation, which was independent of temperature in the range of 10–40 K. The temperature dependences of the decay kinetics showed that the pseudorotation is not affected by C₆₀ aggregation at higher temperatures.     

Features of the Distribution of Spectral Parameters of Intermolecular Vibrations in Water Obtained by Raman Spectroscopy

Optics and Spectroscopy - Using Raman scattering, we have studied the spectral characteristics of intermolecular vibrations of water that are associated with low- (about 50 cm–1 in the IR...     

共振拉曼光谱研究卟啉二酸H_4~(2+)TSPP的J-聚焦

用共振拉曼光谱和ＵＶ Ｖｉｓ吸收光谱考察了离子强度对四一（对一磺酸基）苯基卟啉二酸（Ｈ２＋４ＴＳＰＰ）Ｊ 聚集体形成的影响。实验表明加入少量的ＮａＣｌＯ４有助于聚集体的形成,而Ｎａ ＣｌＯ４的浓度过高则倾向于形成单体。ＮａＣｌＯ４的加入不改变聚集体中分子的构型和排列方式,但高浓度的ＮａＣｌＯ４可能在聚集体中引入某种缺陷,使激子态发生局域化。我们还得到了具有宏观尺寸的纤维状Ｈ２＋４ＴＳＰＰ沉淀,并研究了其固体显微拉曼谱。     

Low-Temperature Molecular Motions in Phospholipid Bilayers in Presence of Glycerol as Studied by Spin-Echo EPR of Spin Labels

Glycerol is used as a cryoprotective agent to protect biological systems under freezing conditions. Electron spin echo (ESE) spectroscopy, a pulsed version of EPR, is capable of studying low-temperature molecular motions of nitroxide spin labels. ESE technique was applied to study molecular motions in phospholipid bilayers prepared from 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) with added spin-labeled lipids 1-palmitoyl-2-stearoyl-(n-DOXYL)-sn-glycero-3-phosphocholine (n-PCSL, n was optionally 5 or 16). Bilayers were hydrated (solvated) either in pure water or in a 1:1 v/v water–glycerol mixture. In the used ESE approach, there were studied stochastic (or diffusive) orientational vibrations of the molecule as a whole (i.e., stochastic molecular librations). The anisotropic contribution to the echo decay rate, W _anis, was measured, which is proportional, according to theory, to the product of the mean-squared angular amplitude \(\langle \alpha^{ 2} \rangle\) and the correlation time τ _c. W _anis was found to be small below and to sharply increase above 200 K, for the both types of solvents and the both label positions. As compared with hydration by pure water, in presence of glycerol W _anis was larger for the 5th label position while for the 16th one it did not change. Also, for the 5th label position W _anis values were found to be nearly the same as those for a polar spin probe 3,4-dicarboxy-PROXYL which was separately added to the bilayer as a reference and which is assumed to be partitioned only into the solvating shell. These results indicate that motions at the surface of bilayer are governed by the motion of solvating shell while motions in the bilayer interior occur independently. The relation of the obtained data with the dynamical transition phenomenon that is known for biological substances near 200 K from neutron scattering and Mössbauer absorption is discussed.