Preparation of methyl glycosides of di- and higher oligosaccharides from glycosaminoglycuronans by solvolysis with dimethyl sulfoxide containing methanol

Solvolysis of chondroitin 4- or 6-sulfate (pyridinium salt) with dimethyl sulfoxide containing 10% of methanol for 18 h at 95° resulted in the cleavage of the 2-amino-2-deoxy-D-glucoside bonds together with initial desulfation to give methyl β-glycosides of N-acetylchondrosine as a main product and, in addition, higher oligosaccharides, without any loss of uronic acid. Dermatan sulfate was also depolymerized to yield methyl glycosides of di- and higher oligosaccharides under the same conditions. Hyaluronic acid (free acid) was depolymerized by the same solvent in the presence of an equimolar amount of pyridine-sulfur trioxide or pyridinium sulfate per disaccharide unit to give methyl glycosides of di- and higher oligosaccharides. In contrast N-desulfated, N-acetylated heparin was stable under these solvolytic conditions and did not yield heparin oligosaccharides.     

Substitution of leucine for tryptophan 412 does not abolish cytochalasin B labeling but markedly decreases the intrinsic activity of GLUT1 glucose transporter

GLUT1 glucose transporter cDNA was modified to introduce a single amino acid substitution of leucine for tryptophan 412, a putative cytochalasin B photo-affinity labeling site. Although the mutated transporter was expressed into plasma membranes of Chinese hamster ovary cells, glucose transport activity of the mutated transporter was observed to be only 15-30% of that of the wild-type GLUT1 when glucose transport activity was assessed by 2-deoxyglucose uptake at 0.1-10 mM concentrations. Analysis of glucose uptake kinetics depict that a mutation induced a 3-fold decrease in turnover number and a 2.5-fold increase in Km compared with the wild-type GLUT1. Importantly, cytochalasin B labeling was not abolished but decreased by 40%, and cytochalasin B binding was also decreased. In addition, the results obtained with side-specific glucose analogs suggested that the outer glucose binding site of the mutant appeared intact but the inner binding site was modulated. These results indicate 1) tryptophan 412 is not a cytochalasin B labeling site(s), although this residue is located in or close to the inner glucose binding site of the GLUT1 glucose transporter, 2) substitution of leucine for tryptophan 412 decreases the intrinsic activity of GLUT1 glucose transporter, which is definable as the turnover number/Km, to approximately 15% of that of the wild-type.     

Molecular structure of the Japanese hepatitis C viral genome

The amino acid sequence of the polyprotein deduced from the nucleotide sequence of the Japanese hepatitis C virus genome (N. Kato et. al. (1990) Proc. Natl. Acad. Sci. USA 87, 9524–9528)indicated that this virus is a member of a new class of positive-stranded RNA viruses. Several domains of this polyprotein also showed weak homology with those of flaviviruses and pestiviruses including the chymotrypsin-like serine proteinase. NTPase and RNA-dependent RNA polymerase     

Ultrastructure of pinealocytes of the cotton rat,Sigmodon hispidus

Summary Fine structural features of pinealocytes of cotton rats (Sigmodon hispidus) were examined. Golgi complexes, mitochondria, endoplasmic reticulum and polysomes are usual organelles seen in the perikaryonal cytoplasm of pinealocytes. Many non-granulated vesicles (40 to 80 nm in diameter) and a few granulated vesicles (about 100 nm in diameter) are associated with the Golgi cisternae. Occasionally, the cisternae contain granular materials. The perikaryonal cytoplasm of pinealocytes is characterized by the presence of inclusion bodies. These bodies are usually round in shape, not bounded by a limiting membrane and composed of fine granular or filamentous materials of high electron-opacity, which are similar in appearance to the substance seen in the nucleolonema. Pinealocyte processes, filled with abundant non-granulated vesicles and some granulated vesicles, are mainly found within the parenchyma and occasionally in perivascular spaces.Supported in part by NSF grant no. PCM 77-05734 and NIH grant no. HD-10202 (Morphology Core)     

Sulfonamide-based non-alkyne LpxC inhibitors as Gram-negative antibacterial agents

We attempted to optimize sulfonamide-based non-alkyne LpxC inhibitors by focusing on improvements in enzyme inhibitory and antibacterial activity. It was discovered that inhibitors possessing 2-aryl benzofuran as a hydrophobe exhibited good activity. In particular, compound 21 displayed impressive antibacterial activity (E. coli MIC = 0.063 μg/mL, K. pneumoniae MIC = 0.5 μg/mL, and P. aeruginosa MIC = 0.5 μg/mL), and is a promising lead for further exploration as an antibacterial agent.     

Structural Change of DNA Induced by Nucleoid Proteins: Growth Phase-Specific Fis and Stationary Phase-Specific Dps

The effects of nucleoid proteins Fis and Dps of Escherichia coli on the higher order structure of a giant DNA were studied, in which Fis and Dps are known to be expressed mainly in the exponential growth phase and stationary phase, respectively. Fis causes loose shrinking of the higher order structure of a genome-sized DNA, T4 DNA (166 kbp), in a cooperative manner, that is, the DNA conformational transition proceeds through the appearance of a bimodal size distribution or the coexistence of elongated coil and shrunken globular states. The effective volume of the loosely shrunken state induced by Fis is 30–60 times larger than that of the compact state induced by spermidine, suggesting that cellular enzymes can access for DNA with the shrunken state but cannot for the compact state. Interestingly, Dps tends to inhibit the Fis-induced shrinkage of DNA, but promotes DNA compaction in the presence of spermidine. These characteristic effects of nucleotide proteins on a giant DNA are discussed by adopting a simple theoretical model with a mean-field approximation.     

Establishment of a mouse model for post‐inflammatory hyperpigmentation

Post‐inflammatory hyperpigmentation (PIH) is a common cutaneous condition that can cause a disfigured appearance. However, the pathophysiology of PIH remains poorly understood, at least in part, because an appropriate animal model for research has not been established. In order to analyze the pathomechanism of PIH, we successfully induced PIH in a hairless version of transgenic mice (hk14‐SCF Tg/HRM) that have a human‐type epidermis containing melanin by repeated hapten application of 2,4‐dinitrofluorobenzene. Histopathologic observation showed epidermal hyperplasia, predominant infiltrations of inflammatory cells, and melanin‐containing cells in the dermis just after elicitation of the atopic dermatitis‐like condition. At week 2, the findings were similar to the characteristics of PIH, that is, an increase of melanin without spongiosis or liquid degeneration in the epidermis and an increase in dermal melanophages. Dynamic analysis of melanin showed that the melanin in the dermis remained for a longer duration than in the epidermis. Furthermore, immunohistochemical staining revealed that the majority of cells containing melanin were positive for the anti‐CD68 antibody, but negative for the anti‐F4/80 antibody. These data suggest that novel treatments of PIH should be targeted against macrophages and should eventually lead to the development of new treatment modalities.