Direct Observation of Fluorescently Labeled Single-stranded λDNA Molecules in a Micro-Flow Channel

We developed two labeling methods for the direct observation of single-stranded DNA (ssDNA), using a ssDNA binding protein and a ssDNA recognition peptide. The first approach involved protein fusion between the 70-kDa ssDNA-binding domain of replication protein A and enhanced yellow fluorescent protein (RPA-YFP). The second method used the ssDNA binding peptide of Escherichia coli RecA labeled with Atto488 (ssBP-488; Atto488-IRMKIGVMFGNPETTTGGNALKFY). The labeled ssλDNA molecules were visualized over time in micro-flow channels. We report substantially different dynamics between these two labeling methods. When ssλDNA molecules were labeled with RPA-YFP, terminally bound fusion proteins were sheared from the free ends of the ssλDNA molecules unless 25-mer oligonucleotides were annealed to the free ends. RPA-YFP-ssλDNA complexes were dissociated by the addition of 0.2 M NaCl, although complex reassembly was possible with injection of additional RPA-YFP. In contrast to the flexible dynamics of RPA-YFP-ssλDNA complexes, the ssBP-488-ssλDNA complexes behaved as rigid rods and were not dissociated even in 2 M NaCl.     

Where does the metal cation stay in Gd@C2v(9)-C82? A single-crystal X-ray diffraction study

Metal positions in endohedral metallofullerenes (EMFs) are of special importance because their molecular symmetry and intrinsic properties are strongly influenced by the location and motion of the encapsulated metals. X-ray analyses of the cocrystals of Gd@C(2v)(9)-C(82) with nickel(II) octaethylporphyrin [Ni(II)(OEP)] reveal that the Gd(3+) cation is off-center, being located under a hexagonal ring along the 2-fold axis of the C(2v)(9)-C(82) cage. This result is in sharp contrast to that of a previous study, showing that Gd@C(2v)(9)-C(82) has an anomalous endohedral structure, with the metal being positioned over a [6,6] bond, which is opposite to the hexagonal ring along the C(2) axis (Phys. Rev. B 2004, 69, 113412). In agreement with theoretical calculations and related studies, it is conclusive that the single rare-earth metal in M@C(2v)(9)-C(82) always tends to coordinate with the hexagonal ring along the 2-fold axis, instead of interacting with the [6,6] bond on the other end, regardless of the type of metal atom.     

Roots of Ehrhart polynomials arising from graphs

Several polytopes arise from finite graphs. For edge and symmetric edge polytopes, in particular, exhaustive computation of the Ehrhart polynomials not merely supports the conjecture of Beck et al. that all roots α of Ehrhart polynomials of polytopes of dimension D satisfy −D≤Re(α)≤D−1, but also reveals some interesting phenomena for each type of polytope. Here we present two new conjectures: (1) the roots of the Ehrhart polynomial of an edge polytope for a complete multipartite graph of order d lie in the circle |z+\fracd4| ￡ \fracd4|z+\frac{d}{4}| \le \frac{d}{4} or are negative integers, and (2) a Gorenstein Fano polytope of dimension D has the roots of its Ehrhart polynomial in the narrower strip -\fracD2 ￡ Re(a) ￡ \fracD2-1-\frac{D}{2} \leq \mathrm{Re}(\alpha) \leq \frac{D}{2}-1. Some rigorous results to support them are obtained as well as for the original conjecture. The root distribution of Ehrhart polynomials of each type of polytope is plotted in figures.     

Ferromagnetic spin coupling between endohedral metallofullerene La@C82 and a cyclodimeric copper porphyrin upon inclusion

The cyclic host cyclo-[P(Cu)](2) carrying two covalently connected Cu(II) porphyrin units can accommodate La@C(82), a paramagnetic endohedral metallofullerene, in its cavity to form the inclusion complex cyclo-[P(Cu)](2)?La@C(82), which can be transformed into the caged complex cage-[P(Cu)](2)?La@C(82) by ring-closing olefin metathesis of its side-chain olefinic termini. On the basis of electron spin resonance (ESR) and electron spin transient nutation (ESTN) studies, cyclo-[P(Cu)](2)?La@C(82) is the first ferromagnetically coupled inclusion complex featuring La@C(82), whereas cage-[P(Cu)](2)?La@C(82) is ferrimagnetic.     

Synthesis and charge-transfer chemistry of La2@I(h)-C80/Sc3N@I(h)-C80-zinc porphyrin conjugates: impact of endohedral cluster

Two stable electron donor-acceptor conjugates, that is, 3 and 5b, employing La(2)@I(h)-C(80) and Sc(3)N@I(h)-C(80), on one hand, and zinc tetraphenylporphyrin, on the other hand, have been prepared via [1+2] cycloaddition reactions of a diazo precursor. Combined studies of crystallography and NMR suggest a common (6,6)-open addition pattern of 3 and 5b. Still, subtly different conformations, that is, a restricted and a comparatively more flexible topography, emerge for 3 and 5b, respectively. In line with this aforementioned difference are the electrochemical assays, which imply appreciably stronger I(h)-C(80)/ZnP interactions in 3 when compared to those in 5b. Density functional calculations reveal significant attractions between the two entities of these conjugates, as well as their separately localized HOMOs and LUMOs. The geometrical conformations and LUMO distributions of 3 and 5b, at our applied computational level, are slightly varied with their different endohedral clusters. The clusters also exert different impact on the excited state reactivity of the conjugates. For example, 3 undergoes, upon photoexcitation, a fast charge separation process and yields a radical ion pair, whose nature, namely, (La(2)@C(80))(?-)-(ZnP)(?+)) versus (La(2)@C(80))(?+)-(ZnP)(?-)), varies with solvent polarity. 5b, on the other hand, afforded the same (Sc(3)N@C(80))(?-)-(ZnP)(?+)) radical ion pair regardless of the solvent.     

Distribution of chloroquine in ocular tissue of pigmented rat using matrix-assisted laser desorption/ionization imaging quadrupole time-of-flight tandem mass spectrometry

In pharmacology and toxicology, localization of the distribution of a drug molecule in its target tissue provides very important in vivo biological information. Traditionally, this has been examined using autoradiography (ARG). However, there are significant limitations in this application. One is the synthesis and use of radiolabeled compounds, the other is that the image generated expresses an undifferentiated mixture of the parent drug and/or its metabolites. The objective of the study was to define the specific distribution of the parent drug in rat ocular tissue containing melanin (e.g. the retina) using non‐labeled chloroquine by MALDI Imaging tandem mass spectrometry (MS/MS). After single oral administration (at 20 mg/kg) of chloroquine, sections (10 µm) of rat eye tissue were prepared at 24 h. The MS system used was a quadrupole time‐of flight (Q‐TOF) tandem mass spectrometer (MALDI Synapt?, Waters, Milford, MA, USA). Tissue sections were sprayed with CHCA (α‐cyano‐4‐hydroxycinnamic acid, 5 mg/mL) in 80% acetonitrile (ACN) containing 5% formic acid (FA) using either a manual sprayer (airbrush) or an automated sprayer (TM‐Sprayer?, HTX Technologies, Carrboro, NC, USA). Chloroquine was readily detected in the MS/MS mode by monitoring one of its major fragment ions (m/z 247.10) and imaged through the rat eye tissue. The image of the specific distribution within the retina in the rat eye tissue was confirmed, and found to be similar to autoradiograms after oral administration of ¹⁴C‐chloroquine reported previously. Copyright © 2011 John Wiley & Sons, Ltd.     

Synthesis of four diastereomers and structural revision of tetradenolide

Four diastereomers of tetradenolide, a cytotoxic α-pyrone isolated from Tetradenia riparia, were synthesized stereoselectively using the Z-selective Horner–Emmons reaction followed by acid catalyzed lactonization. Making comparison of the ¹H and ¹³C NMR spectral data of the four diastereomers with those of the reported value of natural product did not lead to determine the relative stereochemistry of the natural tetradenolide. Thus detailed investigation of the spectral data of the related compounds led us to revise the structure of tetradenolide as deacetylboronolide.     

Photoresponsive crown ethers. 10. Metal complexation by light-switched crown ethers immobilized in polymer matrices

The crosslinked polystyrene beads which immobilized two terminal azophenoxide groups of an azobenzene-crown(24-crown-8)-azobenzene unit were synthesized. The polymer beads adsorbed Cs⁺ in the dark while they rapidly released Cs⁺ into the solution under UV-light irradiation, and the photoresponsive complexation occurred reversibly. On the other hand, the crosslinked polystyrene beads which immobilized one terminal azophenoxide group of an azobenzene-crown (24-crown-8) unit scarcely (or only to a smaller extent) exhibited such a photoresponsive behavior. Similarly, the polymer beads which immobilized two terminal azophenoxide groups of an azobenzene-crown (18-crown-6) azobenzene unit reversibly adsorbed and desorbed K⁺ in response to UV-light irradiation. The novel photoresponsive behavior is ascribed to a conformational distortion of the crown rings which induced by the photoisomerization of the azobenzene moiety, although the photoinduced polarity change of the polymer beads is not completely ruled out. The results suggest that the polymer support is useful as a “fixed point” to enforce the conformational changes of immobilized functional molecules.