'Click chemistry' on polysaccharides: a convenient, general, and monitorable approach to develop (1-->3)-beta-D-glucans with various functional appendages

(1-->3)-beta-D-Glucans having various functional appendages (lactoside, ferrocene, pyrene, and porphyrin) can be prepared in an convenient, quantitative, and regioselective manner through regioselective bromination-azidation of curdlan to afford 6-azido-6-deoxycurdlan followed by chemoselective [3+2]-cycloadditions with various functional modules bearing a terminal alkyne group. The ability to monitor reaction conversions is an additional advantage of this synthetic approach over the conventional direct modifications on polysaccharides; the reaction can be readily monitored based on the intensity of azido peaks in the in situ attenuated total reflection infrared spectra.     

Reduced responsiveness is an essential feature of chronic fatigue syndrome: A fMRI study

Background  

Although the neural mechanism of chronic fatigue syndrome has been investigated by a number of researchers, it remains poorly understood.     

Identification of 8-amino-2,5,7-trihydroxynaphthalene-1,4-dione, a novel intermediate in the biosynthesis of Streptomyces meroterpenoids

Furaquinocin is a natural polyketide-isoprenoid hybrid (meroterpenoid) produced by Streptomyces sp. strain KO-3988. All of the fur genes required for furaquinocin biosynthesis have been cloned, and the heterologous production of furaquinocin has been demonstrated in Streptomyces albus. Here, we report the identification of 8-amino-2,5,7-trihydroxynaphthalene-1,4-dione (8-amino-flaviolin) produced by the S. albus heterologous expression of the three contiguous genes encoding type III polyketide synthase (Fur1), monooxygenase (Fur2), and aminotransferase (Fur3) in the furaquinocin biosynthetic gene cluster. An S. albus transformant (S. albus/pWHM-Fur2_del3) harboring the fur gene cluster and lacking the fur3 gene did not produce furaquinocin, whereas furaquinocin production was restored when 8-amino-flaviolin was added to the culture medium of S. albus/pWHM-Fur2_del3. These results demonstrate that Fur3 aminotransferase is essential for furaquinocin biosynthesis and that 8-amino-flaviolin is an early-stage intermediate in furaquinocin biosynthesis. We propose that the biosynthetic pathway generating 8-amino-flaviolin is the common route for the biosynthesis of Streptomyces meroterpenoids.     

Purification of gangliosides by liquid-liquid partition chromatography

A liquid-liquid partition chromatographic technique was applied to separate amphiphilic glycolipids. A two-phase solvent system composed of n-butanol-t-butyl methyl ether-acetonitrile-water at a volume ratio of 3:1:1:5 was found to be suitable for separating the gangliosides from total lipids extracted from rat brain by liquid-liquid partition chromatographic systems, namely centrifugal partition chromatography (CPC) and high-speed counter-current chromatography. GM1 could be separated rapidly by using the upper phase as stationary phase for both systems. Moreover, various kinds of gangliosides (GM1, GD1a, GD1b, GT1b) could be separated individually by using the lower phase as stationary phase by CPC. The sample can be recovered without loss by these systems.     

Microchamber arrays for the identification of individual cells exposed to an X-ray microbeam

To identify individual cells exposed to a X-ray microbeam in a cell population, we developed a biocompatible microchamber-array chip using UV lithography of photopolymer SU-8. The center-to-center distance between microchambers is 50 μm including a wall of 15 μm height. Using the microchamber-array chip, we performed tracking of individual exposed cells. Sample cells loaded in a microchamber array were selectively irradiated with the X-ray microbeam under microscopic observation. All the irradiated cells were indexed by the array arrangement of the microchambers. For about 24 h of post-irradiation incubation, the irradiated cells were identified successfully by time-lapse observation. In addition, the induction of radiation effects was observed in identified cells using immunofluorescence.     

The speciation of conger eel galectins by rapid adaptive evolution

Many cases of accelerated evolution driven by positive Darwinian selection are identified in the genes of venomous and reproductive proteins. This evolutional phenomenon might have important consequences in their gene-products' functions, such as multiple specific toxins for quick immobilization of the prey and the establishment of barriers to fertilization that might lead to speciation, and in the molecular evolution of novel genes. Recently, we analyzed the molecular evolution of two galectins isolated from the skin mucus of conger eel (Conger myriaster), named congerins I and II, by cDNA cloning and X-ray structural analysis, and we found that they have evolved in the rapid adaptive manner to emergence of a new structure including strand-swapping and a unique new ligand-binding site. In this review article we summarize and discuss the molecular evolution, especially the rapid adaptive evolution, and the structure-function relationships of conger eel galectins. Published in 2004.     

Voxel Based Analysis of Surgical Revascularization for Moyamoya Disease: Pre- and Postoperative SPECT Studies

Moyamoya disease (MMD) is a chronic, progressive, cerebrovascular occlusive disease that causes abnormal enlargement of collateral pathways (moyamoya vessels) in the region of the basal ganglia and thalamus. Cerebral revascularization procedures remain the preferred treatment for patients with MMD, improving the compromised cerebral blood flow (CBF). However, voxel based analysis (VBA) of revascularization surgery for MMD based on data from pre- and postoperative data has not been established. The latest algorithm called as Diffeomorphic Anatomical Registration Through Exponentiated Lie Algebra (DARTEL) has been introduced for VBA as the function of statistical parametric mapping (SPM8), and improved registration has been achieved by SPM8 with DARTEL. In this study, VBA was conducted to evaluate pre- and postoperative single photon emission computed tomography (SPECT) images for MMD by SPM8 with DARTEL algorithm, and the results were compared with those from SPM8 without DARTEL (a conventional method). Thirty-two patients with MMD who underwent superficial temporal artery-middle cerebral artery (STA-MCA) bypass surgery as the first surgery were included and all patients underwent pre- and postoperative 3D T1-weighted imaging and SPECT. Pre- and postoperative SPECT images were registered to 3D T1-weighted images, then VBA was conducted. Postoperative SPECT showed more statistically increased CBF areas in the bypassed side cerebral hemisphere by using SPM8 with DARTEL (58,989 voxels; P<0.001), and increased ratio of CBF after operation was less than 15%. Meanwhile, postoperative SPECT showed less CBF increased areas by SPM8 without DARTEL. In conclusion, VBA was conducted for patients with MMD, and SPM8 with DARTEL revealed that postoperative SPECT showed statistically significant CBF increases over a relatively large area and with at most 15% increase ratio.     

Distinct roles for U‐type proteins in iron–sulfur cluster biosynthesis revealed by genetic analysis of the Bacillus subtilis sufCDSUB operon

The biosynthesis of iron–sulfur (Fe–S) clusters in Bacillus subtilis is mediated by the SUF‐like system composed of the sufCDSUB gene products. This system is unique in that it is a chimeric machinery comprising homologues of E. coli SUF components (SufS, SufB, SufC and SufD) and an ISC component (IscU). B. subtilis SufS cysteine desulfurase transfers persulfide sulfur to SufU (the IscU homologue); however, it has remained controversial whether SufU serves as a scaffold for Fe–S cluster assembly, like IscU, or acts as a sulfur shuttle protein, like E. coli SufE. Here we report that reengineering of the isoprenoid biosynthetic pathway in B. subtilis can offset the indispensability of the sufCDSUB operon, allowing the resultant Δsuf mutants to grow without detectable Fe–S proteins. Heterologous bidirectional complementation studies using B. subtilis and E. coli mutants showed that B. subtilis SufSU is interchangeable with E. coli SufSE but not with IscSU. In addition, functional similarity in SufB, SufC and SufD was observed between B. subtilis and E. coli. Our findings thus indicate that B. subtilis SufU is the protein that transfers sulfur from SufS to SufB, and that the SufBCD complex is the site of Fe–S cluster assembly.     

The giant mycoheterotrophic orchid Erythrorchis altissima is associated mainly with a divergent set of wood‐decaying fungi

The climbing orchid Erythrorchis altissima is the largest mycoheterotroph in the world. Although previous in vitro work suggests that E. altissima has a unique symbiosis with wood‐decaying fungi, little is known about how this giant orchid meets its carbon and nutrient demands exclusively via mycorrhizal fungi. In this study, the mycorrhizal fungi of E. altissima were molecularly identified using root samples from 26 individuals. Furthermore, in vitro symbiotic germination with five fungi and stable isotope compositions in five E. altissima at one site were examined. In total, 37 fungal operational taxonomic units (OTUs) belonging to nine orders in Basidiomycota were identified from the orchid roots. Most of the fungal OTUs were wood‐decaying fungi, but underground roots had ectomycorrhizal Russula. Two fungal isolates from mycorrhizal roots induced seed germination and subsequent seedling development in vitro. Measurement of carbon and nitrogen stable isotope abundances revealed that E. altissima is a full mycoheterotroph whose carbon originates mainly from wood‐decaying fungi. All of the results show that E. altissima is associated with a wide range of wood‐ and soil‐inhabiting fungi, the majority of which are wood‐decaying taxa. This generalist association enables E. altissima to access a large carbon pool in woody debris and has been key to the evolution of such a large mycoheterotroph.