Deoxyhypusine synthase is phosphorylated by protein kinase C in vivo as well as in vitro

Deoxyhypusine synthase catalyzes the first step in the posttranslational synthesis of an unusual amino acid, hypusine, in the eukaryotic translation initiation factor 5A (eIF-5A) precursor protein. We earlier observed that yeast recombinant deoxyhypusine synthase was phosphorylated by protein kinase C (PKC) in vitro (Kang and Chung, 1999) and the phosphorylation rate was synergistically increased to a 3.5-fold following treatment with phosphatidylserine (P.Ser)/diacylglycerol (DAG)/ Ca(2+), suggesting a possible involvement of PKC. We have extended study on the phosphorylation of deoxyhypusine synthase in vivo in different cell lines in order to define its role on the regulation of eIF5A in the cell. Deoxyhypusine synthase was found to be phosphorylated by endogenous kinases in CHO, NIH3T3, and chicken embryonic cells. The highest degree of phosphorylation was found in CHO cells. Moreover, phosphorylation of deoxyhypusine synthase in intact CHO cells was revealed and the expression of phosphorylated deoxyhypusine synthase was significantly diminished by diacyl ethylene glycol (DAEG), a PKC inhibitor, and enhanced by phorbol 12-myristate 13-acetate (PMA) or Ca(2+)/DAG. Endogenous PKC in CHO cell and cell lysate was able to phosphorylate deoxyhypusine synthase and this modification is enhanced by PMA or Ca(2+) plus DAG. Close association of PKC with deoxyhypusine synthase in the CHO cells was evident in the immune coprecipitation and was PMA-, and Ca(2+)/phospholipid dependent. These results suggest that phosphorylation of deoxyhypusine synthase was PKC-dependent cellular event and open a path for possible regulation in the interaction with eIF5A precursor for hypusine synthesis.     

Purification of a primary coolant in a nuclear power plant using a magnetic filter - electrodeionization hybrid separation system

A magnetic filter — continuous electrodeionization (CEDI) hybrid separation system was investigated for the purification of the primary coolant in a nuclear power plant. A magnetic filter system with a 3000 Gauss magnetic field and a CEDI system with a cell consisting of 3 compartments were used for the removal of magnetite and nickel ions, respectively. The hybrid separation system achieved removal rates of 98% for magnetite and 99% for the nickel ions demonstrating its feasibility for the purification of primary coolant.The authors gratefully acknowledge the Basic Atomic Energy Research Institute (BAERI) program at the Korea Institute of Science and Technology Evaluation and Planning (KISTEP) for the financial support to carry out this work.     

Identification of flavonoids using liquid chromatography with electrospray ionization and ion trap tandem mass spectrometry with an MS/MS library

Searchable MS/MS spectra libraries, constructed using the results of liquid chromatography coupled with electrospray ionization (ESI) tandem mass spectrometry (LC/MS/MS) with data-dependent acquisition on an ion trap mass spectrometer, are presented with regard to the identification and confirmation of a variety of closely related flavonoids in a set of biological samples. Flavonoids were found to exhibit a maximum amount of structurally specific MS/MS spectra at 45% of normalized collision energy on the instrument used, without wideband activation. These MS/MS spectra were then searched automatically against a 297-substance MS/MS library that contains many previously acquired spectra of standard flavonoids. The possible applications of this powerful technique to biological samples are also discussed. Daidzein and genistein were identified through the MS/MS spectra library while searching through LC/MS/MS data for plant and microbial extracts. Moreover, these compounds proved completely distinguishable from other flavonoids of closely related structures in the MS/MS spectra library, using the NIST MS search program. The applicability of the library-searchable spectra at low concentrations was demonstrated by successful identification of daidzein and genistein at 0.05 and 0.5 microg/mL, respectively.     

Biocatalytic precipitation induced by an affinity reaction on dendrimer-activated surfaces for the electrochemical signaling from immunosensors

We have developed a strategy of signal generation for immunosensors that transduces biospecific affinity recognition reactions into electrochemical signals. The cyclic voltammetric method, tracking the precipitation of insoluble products onto the sensing surface and the subsequent decrement in the electrode area, was chosen for signal registration. Precipitation of insolubilities was induced by the catalytic reaction of enzymes, which were labeled to the biospecifically attached protein or antibody molecules. As a model system for affinity recognition, we have investigated the functionalization of biotin groups to the sensing monolayer and their biospecific interactions with anti-biotin antibody molecules. The immunosensing interface was developed onto the dendrimer-activated self-assembled monolayers (SAMs), as the base template for the functionalization of the antigen moiety and signal generation. The advantages of using dendrimer-activated SAMs in comparison to the plain modified thiolate SAMs for the sensing surface were shown in terms of sensing performances, and the analytical characteristics of the resulting immunosensor were examined. Additionally, the sensing system was applied for biotin/(strept)avidin couples, extending the applicability of the developed strategy.     

Electrolytic production of metallic uranium from U3O8 in a 20-kg batch scale reactor</p> </p>

Summary The electrolytic reduction of U₃O₈ powder was carried out using LiCl-Li₂O molten salt in a 20-kg U₃O₈ batch cell to verify the feasibility of the process. As the current passes the cell, the decomposition of Li₂O and the reduction of U₃O₈ occur simultaneously in a cathode assembly and oxygen gas evolvs at the anode. The results from a 20-kg U₃O₈ scale cell were compared with data obtained from a bench scale cell. The results suggest a successful demonstration of this process, exhibiting a reduction conversion of U₃O₈ of more than 99% in a batch.</p> </p>     

Efficient excitation energy transfer in long meso-meso linked Zn(II) porphyrin arrays bearing a 5,15-bisphenylethynylated Zn(II) porphyrin acceptor

Electronically coupled porphyrin arrays are suitable for artificial light harvesting antenna in light of a large absorption cross-section and fast excitation energy transfer (EET). Along this line, an artificial energy transfer model system has been synthesized, comprising of an energy donating meso-meso linked Zn(II) porphyrin array and an energy accepting 5,15-bisphenylethynylated Zn(II) porphyrin linked via a 1,4-phenylene spacer. This includes an increasing number of porphyrins in the meso-meso linked Zn(II) porphyrin array, 1, 2, 3, 6, 12, and 24 (Z1A, Z2A, Z3A, Z6A, Z12A, and Z24A). The intramolecular singlet-singlet EET processes have been examined by means of the steady-state and time-resolved spectroscopic techniques. The steady-state fluorescence comes only from the acceptor moiety in Z1A-Z12A, indicating nearly the quantitative EET. In Z24A that has a molecular length of ca. 217 A, the fluorescence comes largely from the acceptor moiety but partly from the long donor array, indicating that the intramolecular EET is not quantitative. The transient absorption spectroscopy has provided the EET rates in real time scale: (2.5 ps)(-1) for Z1A, (3.3 ps)(-1) for Z2A, (5.5 ps)(-1) for Z3A, (21 ps)(-1) for Z6A, (63 ps)(-1) for Z12A, and (108 ps)(-1) for Z24A. These results have been well explained by a revised F?rster equation (Sumi formula), which takes into account an exciton extending coherently over several porphyrin pigments in the donor array, whose length is not much shorter than the average donor-acceptor distance. Advantages of such strongly coupled porphyrin arrays in light harvesting and transmission are emphasized in terms of fast EET and a large absorption cross-section for incident light.     

Synthesis of nanosized TiO2/SiO2 particles using microwave processes and their photocatalytic activity on the decomposition of orange II

The nanosized titania and TiO₂/SiO₂ particles were prepared by the microwave-hydrothermal method. The effect of physical properties TTIP/TEOS ratio and calcination temperature has been investigated. The major phase of the pure TiO₂ particle is of the anatase structure, and a rutile peak was observed above 800°C. In TiO₂/SiO₂ particles, however, no significant rutile phase was observed, although the calcination temperature was 900°C. No peaks for the silica crystal phase were observed at either silica/titania ratio. The crystallite size of TiO₂/SiO₂ particles decreases as compared to pure TiO₂ at high calcination temperatures. The TiO₂/SiO₂ particles show higher activity on the photocatalytic decomposition of orange II as compared to pure TiO₂ particles.     

Self-assembled amphiphilic fluorescent probe: detecting pH-fluctuations within cancer cells and tumour tissues

Abnormal anaerobic metabolism leads to a lowering of the pH of many tumours, both within specific intracellular organelles and in the surrounding extracellular regions. Information relating to pH-fluctuations in cells and tissues could aid in the identification of neoplastic lesions and in understanding the determinants of carcinogenesis. Here we report an amphiphilic fluorescent pH probe (CS-1) that, as a result of its temporal motion, provides pH-related information in cancer cell membranes and selected intracellular organelles without the need for specific tumour targeting. Time-dependent cell imaging studies reveal that CS-1 localizes within the cancer cell-membrane about 20 min post-incubation. This is followed by migration to the lysosomes at 30 min before being taken up in the mitochondria after about 60 min. Probe CS-1 can selectively label cancer cells and 3D cancer spheroids and be readily observed using the green fluorescence channel (λ_em = 532 nm). In contrast, CS-1 only labels normal cells marginally, with relatively low Pearson''s correlation coefficients being found when co-incubated with standard intracellular organelle probes. Both in vivo and ex vivo experiments provide support for the suggestion that CS-1 acts as a fluorescent label for the periphery of tumours, an effect ascribed to proton-induced aggregation. A much lower response is seen for muscle and liver. Based on the present results, we propose that sensors such as CS-1 may have a role to play in the clinical and pathological detection of tumour tissues or serve as guiding aids for surgery.

A self-assembled amphiphilic fluorescent probe allows pH-fluctuations within cancer cells and tumour tissues to be readily detected.     

Photodissociation tandem mass spectrometry at 266 nm of an aliphatic peptide derivatized with phenyl isothiocyanate and 4-sulfophenyl isothiocyanate

Photodissociation (PD) tandem mass spectra have been obtained at 266 nm for the protonated molecules of a tryptic peptide, ASHLGLAR, and of its phenyl isothiocyanate and 4-sulfophenyl isothiocyanate derivatives, generated by matrix-assisted laser desorption/ionization. Derivatization with the aromatic chromophores greatly reduced the intensity of the laser required for efficient PD. Major fragment ions observed in the three spectra are quite similar. General features of the PD tandem mass spectra and their potential utility for analytical purposes are discussed.