Vectorially Imprinted Hybrid Nanofilm for Acetylcholinesterase Recognition

Effective recognition of enzymatically active tetrameric acetylcholinesterase (AChE) is accomplished by a hybrid nanofilm composed of a propidium‐terminated self‐assembled monolayer (Prop‐SAM) which binds AChE via its peripheral anionic site (PAS) and an ultrathin electrosynthesized molecularly imprinted polymer (MIP) cover layer of a novel carboxylate‐modified derivative of 3,4‐propylenedioxythiophene. The rebinding of the AChE to the MIP/Prop‐SAM nanofilm covered electrode is detected by measuring in situ the enzymatic activity. The oxidative current of the released thiocholine is dependent on the AChE concentration from ≈0.04 × 10^?6 to 0.4 × 10^?6m . An imprinting factor of 9.9 is obtained for the hybrid MIP, which is among the best values reported for protein imprinting. The dissociation constant characterizing the strength of the MIP‐AChE binding is 4.2 × 10^?7m indicating the dominant role of the PAS‐Prop‐SAM interaction, while the benefit of the MIP nanofilm covering the Prop‐SAM layer is the effective suppression of the cross‐reactivity toward competing proteins as compared with the Prop‐SAM. The threefold selectivity gain provided by i) the “shape‐specific” MIP filter, ii) the propidium‐SAM, iii) signal generation only by the AChE bound to the nanofilm shows promise for assessing AChE activity levels in cerebrospinal fluid.     

Studies of histidine as a suitable isoelectric buffer for tryptic digestion and isoelectric trapping fractionation followed by capillary electrophoresis-mass spectrometry for proteomic analysis

The use of histidine as a protein digestion buffer followed by isoelectric trapping separations using "membrane separated wells for isoelectric focusing and trapping" (MSWIFT) and mass spectrometry (MS) analysis is described. Tryptic digestion of bovine serum albumin (BSA) performed in histidine buffered solutions yields similar amino acid sequence coverage values to those obtained using ammonium bicarbonate buffer. Time course studies suggest that histidine buffers provide faster migration of peptides from the loading compartment compared to digestions prepared in ammonium bicarbonate due to differences in conductivities of the two buffers. In addition, this sample preparation method and MSWIFT separations have been coupled with capillary electrophoresis (CE) and matrix-assisted laser desorption ionization-mass spectrometry (MALDI-MS) as an alternative separation approach for proteomic studies. Tryptic peptides of ribosomal proteins in histidine are fractionated using MSWIFT followed by CE-MALDI-MS, which further illustrates the ability to couple fractions from a pI based separation device to CE-MS. Specifically, two-dimensional CE-MS plots provide a direct correlation between the numbers of basic residues within the peptide sequence displayed in charge-state trend lines. Combining MSWIFT and CE-MS provides added information regarding peptide sequence, specifically pI and in-solution charge state. Post-translational modifications can also be identified using this method.     

Synthesis, structure and photocatalytic properties of Fe(III)-doped TiO2 prepared from TiCl3

Iron(III)-doped titanium dioxide photocatalysts were prepared from aqueous titanium(III) chloride solution in the presence of dissolved FeCl₃ (0–10.0 at.% relative to TiCl₃) by co-precipitation method. The precipitate was completely oxidized in the aerated suspension, hydrothermally treated, washed and calcinated. The structure of the powders was characterized by thermoanalysis (TG-DTA), diffuse reflectance spectroscopy (DRS), X-ray diffraction (XRD), nitrogen adsorption and transmission electron microscopy (TEM). The light absorption of the iron-containing powders is red shifted relative to the bare sample. The particle size and anatase content were found to significantly decrease at iron contents ≥6.0 at.% which is accompanied with the increase of their specific surface area. XANES measurements showed that the local structure of iron systematically changes with the variation of the dopant concentrations: at higher Fe-contents, hematite- or goethite-like environments were observed, consistent with the formation of separate X-ray amorphous Fe(III)-containing phases. The local structure of iron gradually transformed with decreasing dopant concentrations, possibly due to substitution of Fe(III) in the titania (TiO₂) crystal lattice. Energy dispersive X-ray analysis (EDX) and chemical analysis was used to characterize the iron content of the samples in the bulk and X-ray photoelectron spectroscopy (XPS) in the surface layer of the particles. The photocatalytic performance of the prepared photocatalysts was compared with the activity of Aldrich anatase under UV–vis and VIS irradiation in two different photoreactors. Maximum photocatalytic performance was found at 3.0 at.% iron concentration for UV–vis and at 1.2 at.% for VIS irradiation. Doping with iron(III) ions increased the photodegradation rate of phenol by a factor of three for UV–vis irradiation and by a factor of two for VIS irradiation, relative to the bare photocatalyst.     

Multilevel Changes in Protein Dynamics upon Complex Formation of the Calcium‐Loaded S100A4 with a Nonmuscle Myosin IIA Tail Fragment

Dysregulation of Ca²⁺‐binding S100 proteins plays important role in various diseases. The asymmetric complex of Ca²⁺‐bound S100A4 with nonmuscle myosin IIA has high stability and highly increased Ca²⁺ affinity. Here we investigated the possible causes of this allosteric effect by NMR spectroscopy. Chemical shift‐based secondary‐structure analysis did not show substantial changes for the complex. Backbone dynamics revealed slow‐timescale local motions in the H1 helices of homodimeric S100A4; these were less pronounced in the complex form and might be accompanied by an increase in dimer stability. Different mobilities in the Ca²⁺‐coordinating EF‐hand sites indicate that they communicate by an allosteric mechanism operating through changes in protein dynamics; this must be responsible for the elevated Ca²⁺ affinity. These multilevel changes in protein dynamics as conformational adaptation allow S100A4 fine‐tuning of its protein–protein interactions inside the cell during Ca²⁺ signaling.     

Short-time effect of heavy metals upon microbial community activity

Microcalorimetry was applied to assess and compare the toxic effect of heavy metals, such as As, Cu, Cd, Cr, Co, Pb and Zn, on the soil microbial activities and community. About 1.0 g soil spiked 5.0 mg glucose and 5.0 mg ammonium sulfate, the microbial activities were recorded as power–time curves, and their indices, microbial growth rate constant k, total heat evolution Q_T, metabolic enthalpy ΔH_met and mass specific heat rate J_Q/S, were calculated. Comparing these thermodynamic parameters associated with growth yield, a general order of toxicity to the soil was found to be Cr > Pb > As > Co > Zn > Cd > Cu. When soil was exposed to heavy metals, the amount of bacteria and fungi decreased with the incubation time, and the bacterial number diminished sharply. It illustrates that fungi are more tolerant, and bacteria–fungi ratio would be altered under metal stress. To determine the status of the glucose consumed, a glucose biosensor with eggshell membrane was used to measure the remaining glucose in soil sample. Results showed that the time at which glucose was consumed completely was agreed with the microcalorimetric time to a large extent, and depended on the toxicity of heavy metals as well.     

Chemoenzymatic Preparation of Enantiopure Homoadamantyl β‐Amino Acid and β‐Lactam Derivatives

Racemic cis‐10‐azatetracyclo[7.2.0.1^2,6.1^4,8]tridecan‐11‐one was prepared from homoadamant‐4‐ene by chlorosulfonyl isocyanate addition. The transformation of the β‐lactam to the corresponding β‐amino ester followed by Candida antarctica lipase A‐catalyzed enantioselective (E>>200) N‐acylation with 2,2,2‐trifluoroethyl butanoate afforded methyl (1R,4R,5S,8S)‐5‐aminotricyclo[4.3.1.1^3,8]undecane‐4‐carboxylate and the (1S,4S,5R,8R)‐butanamide with>99% ee at 50% conversion. Alternatively, transformation of the β‐lactam to the corresponding N‐hydroxymethyl‐β‐lactam and the following Pseudomonas cepacia (currently Burkholderia cepacia) lipase‐catalyzed enantioseletive O‐acylation provided the (1S,4S,6R,9R)‐alcohol (ee=87%) and the corresponding (1R,4R,6S,9S)‐butanoate (ee>99%). In the latter method, competition for the enzyme between the (1R,4R,6S,9S)‐butanoate, 2,2,2‐trifluoroethyl butanoate and the hydrolysis product, butanoic acid, tended to stop the reaction at about 45% conversion and finally gave racemization in the (1S,4S,6R,9R)‐alcohol with time.     

Simplified Model of Voids Able to Mimic Accelerating Expansion at High <Emphasis Type="Italic">z</Emphasis> without Dark Energy

We present a simplified model of voids in the Universe. Using this model, we investigate the effect of inhomogeneities on light propagation. We calculate the corresponding Hubble diagrams and compare them to those expected in different cosmological models. We find that voids in a universe containing only nonrelativistic matter can mimic accelerating expansion as far as Hubble diagrams are concerned.     

Eine rheo-optische Apparatur für die Durchführung homogener,einfacher Dehnungen an Polymerschmelzen mit simultaner Aufzeichnung der Deformation,der mechanischen Spannung und der Doppelbrechung

The present paper describes a rheo-optical apparatus consisting of an elongational rheometer (with rotary clamps) and a device to measure birefringence. This equipment was designed to perform homogeneous deformations on polymer melts in the simple elongational mode. Elongational experiments were performed on polyisobutylene samples at room temperature using different strain rates (from \documentclass{article}\pagestyle{empty}\begin{document}$\dot \varepsilon$\end{document} 0.01 s^?1 to 1 s^?1). In typical experiments homogeneous elongations up to λ ≈ 200 and stresses up to 2 · 10⁶ Pa were achieved. The stress, the birefringence, and the deformation were obtained by measuring the force, the sample thickness, the optical retardation (the frequency of the sampling was 20 Hz) and the width of the sample (the frequency of the sampling was 3 Hz). It was found that the stress-optical coefficient was constant. The homogeneity of the deformations was carefully controlled because inhomogeneities cause serious errors in the measurements of elongational viscosity and birefringence.