Single‐Molecule Methods to Study Membrane Receptor Oligomerization

Membrane receptors control fundamental cellular processes. Binding of a specific ligand to a receptor initiates communication through the membrane and activation of signaling cascades. This activation process often leads to a spatial rearrangement of receptors in the membrane at the molecular level. Single‐molecule techniques contributed significantly to the understanding of receptor organization and rearrangement in membranes. Here, we review four prominent single‐molecule techniques that have been applied to membrane receptors, namely, stepwise photobleaching, Förster resonance energy transfer, sub‐diffraction localization microscopy and co‐tracking. We discuss the requirements, benefits and limitations of each technique, discuss target labeling, present a selection of applications and results and compare the different methodologies.     

Cellulose Iβ investigated by IR-spectroscopy at low temperatures

Highly crystalline oriented Halocynthia roretzi cellulose Iβ films were investigated by IR-spectroscopy between ?180 and +10 °C. Changes in the IR-spectra induced by temperature were compared to published changes induced by mechanical stretching. This made it possible to conclude that frequency shifts in the O–H stretching region of the IR-spectra due to temperature were not predominantly an indirect effect of thermal expansion leading to greater O–O distances, but were due directly to the effect of temperature on the O–H···O hydrogen bonds. Temperature induced frequency shifts of C–H stretching bands were consistent with the presence of weak inter-sheet C–H···O bonds. Furthermore, no phase transition in cellulose Iβ was found between ?180 and +10 °C.     

The In Vitro Anti-pathogenic Activity of Immunoglobulin Concentrates Extracted from Ovine Blood

An immunoglobulin-rich fraction has been prepared from ovine blood in our laboratory. We have investigated its antibacterial activity and binding activity to pathogenic whole cell antigens, lipopolysaccharide (LPS) and staphylococcal enterotoxin B. Ovine immunoglobulin concentrate (OIC) comprised about 73 ± 2% of IgG and 11 ± 1% of IgM on a protein basis. It inhibited the growth of all 13 strains of pathogens tested, but the inhibitory activity varied according to bacterial strain. The inhibitory activity of OIC was attributed to the high contents of undenatured immunoglobulin present because its inhibitory activity was destroyed by pepsin digestion and heat treatment (65°C for 30 min). OIC bound to all the Gram-positive and Gram-negative pathogens, regardless of cell wall structure. The highest magnitude of crossreactivity to whole cell antigens was against Staphylococcus epidermidis and Shigella soneii strains (p < 0.001). The binding activity of OIC to LPS obtained from Escherichia coli O111:B4 and Salmonella enterica serotype typhimurium was assessed by enzyme-linked immunosorbent assay and lymphoblast K-562 proliferation assay. OIC bound to LPS with a binding activity that was dependent on OIC concentration and saturable, showing typical hyperbolic curves. For toxin-binding activity, an OIC concentration-dependent trend like that for LPS-binding activity was also observed. This preliminary evidence suggests that the OIC used in this study could be a promising supplement for protecting against pathogenic bacteria.     

An EPR, ENDOR and EIE study of gamma-irradiated poly (lactide-co-glycolide) polymers

Gamma radiation of poly (lactide-co-glycolide) raw polymers and processed microspheres under vacuum and at 77 K results in the formation of a series of free radicals. The resulting powder electron paramagnetic resonance (EPR) spectrum contains a distribution of several different radicals, depending on the annealing temperature, and is therefore difficult to interpret. By utilising the selectivity of the electron nuclear DOuble resonance (ENDOR) and associated ENDOR induced EPR (EIE) techniques, a more direct approach for the deconvolution of the EPR spectrum can be achieved. Using this approach, the radiolytically induced CH3 *CHC(O)R- chain scission radical was identified at 120 K by simulation of the EIE spectrum. At elevated temperatures (250 K), this radical decays considerably and the more stable radicals -O*CHC(O)-, CH3 *C(OR)C(O)- and CH3 *C(OH)C(O)- predominate. This work demonstrates the utility of the EIE approach to supplement and aid the interpretation of powder EPR spectra of radicals in a polymer matrix.     

Design of open-framework germanates by combining different building units

Open-framework germanates with extra-large channels are built from two different types of clusters. The two different clusters are connected in such a way to form a pseudo-body-centered cluster aggregate (PBCCA). The different packing of PBCCAs resulted in different 3D open-framework germanates.     

Chemical Probing of the Human Sirtuin 5 Active Site Reveals Its Substrate Acyl Specificity and Peptide‐Based Inhibitors

Sirtuins are NAD⁺‐dependent deacetylases acting as sensors in metabolic pathways and stress response. In mammals there are seven isoforms. The mitochondrial sirtuin 5 is a weak deacetylase but a very efficient demalonylase and desuccinylase; however, its substrate acyl specificity has not been systematically analyzed. Herein, we investigated a carbamoyl phosphate synthetase 1 derived peptide substrate and modified the lysine side chain systematically to determine the acyl specificity of Sirt5. From that point we designed six potent peptide‐based inhibitors that interact with the NAD⁺ binding pocket. To characterize the interaction details causing the different substrate and inhibition properties we report several X‐ray crystal structures of Sirt5 complexed with these peptides. Our results reveal the Sirt5 acyl selectivity and its molecular basis and enable the design of inhibitors for Sirt5.     

Accelerating molecular dynamic simulation on the cell processor and Playstation 3

Implementation of molecular dynamics (MD) calculations on novel architectures will vastly increase its power to calculate the physical properties of complex systems. Herein, we detail algorithmic advances developed to accelerate MD simulations on the Cell processor, a commodity processor found in PlayStation 3 (PS3). In particular, we discuss issues regarding memory access versus computation and the types of calculations which are best suited for streaming processors such as the Cell, focusing on implicit solvation models. We conclude with a comparison of improved performance on the PS3's Cell processor over more traditional processors.     

Molecular Photoacoustic Tomography with Colloidal Nanobeacons

Spotting clots : Vascularly constrained colloidal gold nanobeacons (GNBs; see picture) can be used as exogenous photoacoustic contrast agents for the targeted detection of fibrin, a major biochemical feature of thrombus. Fibrin‐targeted GNBs provide a more than tenfold signal enhancement in photoacoustic tomography in the near‐IR wavelength window, indicating their potential for diagnostic imaging.

     

Cooperative Brønsted acid-type organocatalysis: alcoholysis of styrene oxides

We present a mild and efficient method for the completely regioselective alcoholysis of styrene oxides utilizing a cooperative Br?nsted acid-type organocatalytic system comprised of mandelic acid (1 mol %) and N,N'-bis-[3,5-bis-(trifluoromethyl)phenyl]-thiourea (1 mol %). Various styrene oxides are readily transformed into their corresponding beta-alkoxy alcohols in good to excellent yields at full conversion. Simple aliphatic and sterically demanding, as well as unsaturated and acid-sensitive alcohols can be employed.     

High-birefringence nematic liquid crystal for broadband THz applications

Liquid crystals (LCs) have been studied extensively in the visible range for their dielectric tunability, and the characterisation in the terahertz (THz) range has gained increasing interest due to the need for active THz modulation and switching devices. In this paper, we use THz time-domain spectroscopy to measure the frequency-dependent birefringence and the absorption coefficient of a number of commercial and non-commercial nematic LCs, including E7, BL037, MDA-98-1602, LCMS-107, GT3-23001 and 1825, over a range of bias voltages at room temperature. Furthermore, several basic components of LC mixture are analysed to establish their contributions to birefringence and theoretical model is used to fit the absorption spectra. The large tunability and low loss measured for a range of samples show that the LCs are useful tunable dielectrics for compact, efficient and broadband THz devices.