Regioselective glucosidation of <Emphasis Type="Italic">trans</Emphasis>-resveratrol in <Emphasis Type="Italic">Escherichia coli</Emphasis> expressing glucosyltransferase from <Emphasis Type="Italic">Phytolacca americana</Emphasis>

A glucosyltransferase (GT) of Phytolacca americana (PaGT3) was expressed in Escherichia coli and purified for the synthesis of two O-β-glucoside products of trans-resveratrol. The reaction was moderately regioselective with a ratio of 4′-O-β-glucoside: 3-O-β-glucoside at 10:3. We used not only the purified enzyme but also the E. coli cells containing the PaGT3 gene for the synthesis of glycoconjugates. E. coli cell cultures also have other advantages, such as a shorter incubation time compared with cultured plant cells, no need for the addition of exogenous glucosyl donor compounds such as UDP-glucose, and almost complete conversion of the aglycone to the glucoside products. Furthermore, a homology model of PaGT3 and mutagenesis studies suggested that His-20 would be a catalytically important residue.     

Complete Demonstration of the Valence Electronic Structure Inside a Practical Organic Solar Cell Probed by Low Energy Photoemission

The complete electronic structure inside a practical organic photovoltaic (OPV) device consisting of a trilayer structure of copper‐phthalocyanine (CuPc), fullerene (C₆₀), and bathocuproine (BCP) is demonstrated using low‐energy ultraviolet photoelectron spectroscopy (LE‐UPS) and photoelectron yield spectroscopy (PYS). The molecular orbital energy alignment and electrostatic potential distribution throughout the entire device is illustrated based on the LE‐UPS results. A favorable potential gradient to carry the photogenerated holes and electrons is manifested to be built spontaneously in the CuPc and BCP layers, respectively. Furthermore, the ultrahigh sensitivity measurements of LE‐UPS clearly unveil the distributions of faint density‐of‐states in the energy‐gap region in the organic films. Substantially barrierless contacts to both electrodes are fulfilled by the existence of these gap states. The electronic structure under simulated sunlight illumination is examined for the purpose of elucidating the electronic structures inside the working devices in the open‐circuit condition. These results indicate experimentally the electronic functionalities of each organic material, in particular of the BCP buffer layer, on the cell efficiency.     

Synthesis of biomolecules from N2, CO,and H2O by electric discharge

A model primitive gas containing a mixture of N₂, CO and water vapor over a water pool (300 mL, 37 °C) was subjected to electric discharges. The discharge vessel (7 L in volume) was equipped with a CO₂ absorber (The CO₂ being formed during the discharge), thus simulating possible absorption of CO₂ in the primitive ocean. The vessel also has a cold trap ( –15 °C), which protects the primary products against the further decomposition in the discharge phase by enabling these products to adhere to the trap. Since the partial pressures of CO and N₂ decreased at rates of 1.5–1.7 cmHg day^–1 and 0.1–0.2 cmHg day^–1, respectively, the gases were added at regular intervals. The solution was analyzed at regular intervals for HCN, HCHO and urea, and maximum concentrations of about 50, 2, and 140 mM were observed. The discharge phase was continued for 6 months. In the solution, glycine (5.6% yield based on the carbon), glycylglycine (0.64%), orotic acid (0.004%) and small amounts of the other amino acids were found.     

Analysis of two distinct B cell activation pathways mediated by a monoclonal T helper cell. I. MHC-restricted activation of B cells by an IL 2-dependent pathway

The present study was carried out to determine whether the MHC-restricted and MHC-unrestricted B cell activation pathways mediated by a single cloned Th cell are separable, and whether these two pathways are mediated by distinct mechanisms. It was demonstrated that the two B cell activating functions of a single cloned Th cell could be separated by their sensitivity to irradiation. It was shown that MHC-restricted B cell activation is mediated by a radiosensitive Th cell function, whereas MHC-unrestricted B cell activation is mediated by a radioresistant function of the same Th cell. In addition, it was shown that recombinant IL 2 can restore or replace the radiosensitive component of MHC-restricted cognate helper function.     

Gingipains inactivate a cell surface ligand on Porphyromonas gingivalis that induces TLR2-and TLR4-independent signaling

Arginine-specific gingipain and lysine-specific gingipain are two major cysteine proteinases produced by Porphyromonas gingivalis. To clarify the role of gingipains in the interaction between P. gingivalis and the innate immune system, CHO reporter cells expressing TLR2 or TLR4 were stimulated with wildtype or gingipain-deficient P. gingivalis cells and activation of nuclear factor-kappaB in these cells was examined. While CHO/CD14 cells and 7.19 cells, an MD-2-defective mutant derived from CHO/CD14 cells, failed to respond to wild-type P. gingivalis, they responded to gingipain-deficient P. gingivalis. On the other hand, CHO/CD14/TLR2 cells responded to both wild-type and gingipain-deficient P. gingivalis. These results suggested that gingipains have no effects on TLR2-dependent signaling from P. gingivalis but have inhibitory effects on TLR2-and TLR4-independent signaling in CHO cells. Indeed, the activity of gingipain-deficient P. gingivalis to induce the activation of 7.19 cells was diminished after treatment of the bacterial cells with gingipains. We next partially purified bacterial cell components activating 7.19 cells from gingipain-deficient P. gingivalis. The activity of the partially purified components was diminished by treatment with heat or gingipains. It is also noteworthy that anti-CD14 mAb inhibited the activation of 7.19 cells induced by the partially purified components. These results indicated that the components of P. gingivalis that were able to induce TLR2-and TLR4-independent signaling were inactivated by gingipains before being recognized by CD14. The inactivation of the components would be helpful for P. gingivalis to escape from the innate immune system.     

A solid-phase assay to screen monoclonal antibodies against DNA-binding protein.

A method is described for selecting monoclonal antibodies (mAb) against DNA-binding protein. The protocol involves a non-radioactive solid-phase DNA binding assay using a 96-well plate. Because the solid-phase assay is highly specific and sensitive, partially purified antigen is sufficient for the immunization, and mAb screening can be performed with crude cell extract as the antigen. MAbs obtained by this method could supershift the DNA-protein complex in the electromobility shift assay, and were sufficient for immunoscreening of a cDNA expression library.     

Cutting edge: tyk2 is required for the induction and nuclear translocation of Daxx which regulates IFN-alpha-induced suppression of B lymphocyte formation

IFN-alpha inhibits B lymphocyte development, and the nuclear protein Daxx has been reported to be essential for this biological activity. We show in this study that IFN-alpha inhibits the clonal proliferation of B lymphocyte progenitors in response to IL-7 in wild-type, but not in tyk2-deficient, mice. In addition, the IFN-alpha-induced up-regulation and nuclear translocation of Daxx are completely abrogated in the absence of tyk2. Therefore, tyk2 is directly involved in IFN-alpha signaling for the induction and translocation of Daxx, which may result in B lymphocyte growth arrest and/or apoptosis.