Establishment of a microplate-formatted cell-based immunoassay for rapid analysis of nucleotide excision repair ability in human primary cells

DNA photolesions induced by UV, cyclobutane pyrimidine dimer (CPD) and (6-4) photoproduct (6-4PP), are repaired by nucleotide excision repair (NER) in human cells. Various immunoassays using monoclonal antibodies specific for the photolesions have been developed and widely used for the analysis of cellular NER activity. In this study, we have newly developed a microplate-formatted cell-based immunoassay, based on indirect immunofluorescence staining with lesion-specific antibodies combined with an infrared imaging system. Using this assay, we show the repair kinetics of CPD and 6-4PP in various fibroblasts from newborn and adult donors with no age-related difference. Furthermore, epidermal keratinocytes and melanocytes exhibit comparable NER activity, and calcium ion-induced differentiation of keratinocytes has no significant impacts on their NER activity. We also evaluated the effects of a proteasome inhibitor, MG132, and a histone deacetylase inhibitor, sodium butyrate, on NER efficiency using this assay. All these results suggest that the new assay is highly useful for the rapid and quantitative analysis of NER activity in various primary cells with limited growth activity and is applicable to a screening system for drugs affecting NER efficiency.     

Uniform mixing of high-Tc superconductivity and antiferromagnetism on a single CuO2 plane of a Hg-based five-layered cuprate

We report a site selective Cu-NMR study on underdoped Hg-based five-layered high-Tc cuprate HgBa2Ca4CU5O(12+delta) with a Tc = 72 K. Antiferromagnetism (AFM) has been found to take place at T(N) = 290 K, exhibiting a large antiferromagnetic moment of 0.67-0.69 microB at three inner planes (IP). This value is comparable to the values reported for nondoped cuprates, suggesting that the IP may be in a nearly nondoped regime. Most surprisingly, the AFM order is also detected with M(AFM)(OP) = 0.1 microB even at two outer planes (OP) that are responsible for the onset of superconductivity (SC). The high-Tc SC at Tc = 72 K can uniformly coexist on a microscopic level with the AFM at OP's. This is the first microscopic evidence for the uniform mixed phase of AFM and SC on a single CuO2 plane in a simple environment without any vortex lattice and/or stripe order.     

Notable temperature effect on the stereoselectivity in the photochemical [2+2] cycloaddition reaction (Paternò-Büchi reaction) of 2,3-dihydrofuran-3-ol derivatives with benzophenone

A notable temperature effect (nonlinear Eyring plot) on stereoselectivity, trans-configured oxetane 2 versus cis-configured oxetane 2, is reported in the photochemical [2+2] cycloaddition reaction (Paternò-Büchi reaction) of 2,3-dihydrofuran-3-ol derivatives 1 with benzophenone.     

Preferential destruction of metallic single-walled carbon nanotubes by laser irradiation

Upon laser irradiation in air, metallic single-walled carbon nanotubes (SWNTs) in carbon nanotube thin film can be destroyed in preference to their semiconducting counterparts when the wavelength and power intensity of the irradiation are appropriate and the carbon nanotubes are not heavily bundled. Our method takes advantage of these two species' different rates of photolysis-assisted oxidation, creating the possibility of defining the semiconducting portions of carbon nanotube (CNT) networks using optical lithography, particularly when constructing all-CNT FETs (without metal electrodes) in the future.     

Telomerase inhibitors identified by a forward chemical genetics approach using a yeast strain with shortened telomere length

Telomerase has been proposed as a selective target for cancer chemotherapy. We established a forward chemical genetics approach using a yeast strain with shortened telomere length. Since this strain rapidly enters cell senescence in the absence of active telomerase, compounds that induce selective growth defects against telomere-shortened yeast could be candidates for drugs acting on telomeres and telomerase. We screened our microbial products library and identified three structurally unrelated antibiotics, chrolactomycin, UCS1025A, and radicicol, as active compounds. Detailed analysis showed that chrolactomycin inhibited human telomerase in a cell-free assay as well as in a cellular assay. Long-term culture of cancer cells with chrolactomycin revealed population-doubling-dependent antiproliferative activity accompanied by telomere shortening. These results suggest that chrolactomycin is a telomerase inhibitor, and that the yeast-based assay is useful for discovering the small molecules acting on human telomerase.     

Formation, spectral, electrochemical, and photochemical behavior of zinc N-confused porphyrin coordinated to imidazole functionalized fullerene dyads

Donor-acceptor dyads were constructed using zinc N-confused porphyrin (ZnNCP), a structural isomer of zinc tetraphenylporphyrin, as a donor, and fullerene as an electron acceptor. Two derivatives, pyridine-coordinated zinc N-confused porphyrin (Py:ZnNCP) and the zinc N-confused porphyrin dimer (ZnNCP-dimer) were utilized to form the dyads with an imidazole-appended fulleropyrrolidine (C60Im). These porphyrin isomers formed well-defined 1:1 supramolecular dyads (C60Im:ZnNCP) via axial coordination. The dyads were characterized by optical absorption and emission, ESI-mass, 1H NMR, and electrochemical methods. The binding constant, K, was found to be 2.8 x 10(4) M(-1) for C60Im:ZnNCP. The geometric and electronic structure of C60Im:ZnNCP were probed by using DFT B3LYP/3-21G methods. The HOMO was found to be on the ZnNCP entity, while the LUMO was primarily on the fullerene entity. The electrochemical properties of C60Im:ZnNCP was probed using cyclic voltammetry in o-dichlorobenzene, 0.1 n-Bu4NClO4. The Py:ZnNCP was found to be easier to oxidize by over 340 mV compared to Py:ZnTPP. Upon dyad formation via axial coordination, the first oxidation revealed an anodic shift of nearly 90 mV. Evidence of photoinduced charge separation from the singlet excited ZnNCP to the appended fullerene was established from time-resolved emission and nanosecond transient absorption studies.     

Molecular recognition of mono- and disaccharides through interaction with p-iodophenylboronic acid in capillary electrophoresis with a chemiluminescence detection system

Molecular recognition of mono- and disaccharides was performed making use of the interaction between their diol groups and p-iodophenylboronic acid in capillary electrophoresis (CE) with a chemiluminescence (CL) detection system. p-Iodophenylboronic acid acted as an enhancer for luminol-horseradish peroxidase-hydrogen peroxide CL reaction. p-Iodophenylboronic acid was injected as a sample into the present system to give a CL peak on the electropherogram. The CL intensities were examined using running buffers including mono- and disaccharides. The CL intensities with 1-methyl-D-glucoside, D-saccharose, D-maltose, D-glucose, and D-fructose decreased in this order. The decrease in CL intensity was based on the formation by p-iodophenylboronic acid of cyclic esters with mono- and disaccharides, particularly with those including cis-diol groups. That is, the decrease in CL intensity affected the specific complexation between p-iodophenylboronic acids and saccharides, leading to the molecular recognition of saccharides. We also report separation of a mixture of p-iodophenol and p-iodophenylboronic acid as well as estimation of the apparent binding constant between p-iodophenylboronic acid and saccharides taking advantage of their molecular recognition behavior.     

Photoluminescence properties of multilayer oxide films intercalated with rare earth ions by the layer-by-layer technique

Multilayer oxide films consisting of a TiO-Eu3+-TiO-Tb3+-NbO-Tb3+-NbO-Eu3+ unit which was prepared by the layer-by-layer technique, showed photoluminescence with a high intensity containing both red and green lights.     

Can mass dissociation patterns of transition‐metal complexes be predicted from electrochemical data?

The Cooks kinetic method has been very convenient to correlate the relative dissociation rates obtained by collision-induced fragmentation experiments with the energies of two related bonds in molecules and complexes in the gas phase. Reliable bond energy data are, however, not always available, particularly for polynuclear transition-metal complexes, such as the triruthenium acetate clusters of the general formula [Ru(3) (micro(3)-O)(micro-CH(3)COO)(6)(py)(2)(L)](+), where L = ring substituted N-heterocyclic ligands. Accordingly, their gas-phase collision-induced tandem mass spectrometry (CID MS/MS) dissociation patterns have been analyzed pursuing a relationship with the more easily accessible redox potentials (E(1/2)) and Lever's E(L) parameters. In fact, excellent linear correlations of ln(1/2A(L)/A(py)), where A(py) and A(L) are the abundance of the fragments retaining the pyridine (py) and L ligand, respectively, with E(1/2) and E(L) were found. This result shows that those electrochemical parameters are correlated with bond energies and can be used in the analysis of the dissociation data. Such modified Cooks method can be used, for example, to determine the electronic effects of substituents on the metal-ligand bonds for a series of transition-metal complexes.