Structure-based discovery of human L-xylulose reductase inhibitors from database screening and molecular docking

Human L-xylulose reductase (XR) is an enzyme of the glucuronic acid/uronate cycle of glucose metabolism and is a possible target for treatment of the long-term complications of diabetes. In this study we utilised the molecular modelling program DOCK to analyse the 249,071 compounds of the National Cancer Institute Database and retrieved those compounds with high predicted affinity for XR. Several carboxylic acid-based compounds were tested and shown to inhibit XR. These included nicotinic acid (IC50=100 microM), benzoic acid (IC50=29 microM) and their derivatives. These results extend and improve upon the activities of known, commercially available inhibitors of XR such as the aliphatic fatty acid n-butyric acid (IC50=64 microM). To optimise the interaction between the inhibitor and the holoenzyme, the program GRID was used to design de novo compounds based on the inhibitor benzoic acid. The inclusion of a hydroxy-phenyl group and a phosphate to the benzoic acid molecule increased the net binding energy by 1.3- and 2.4-fold, respectively. The resultant compounds may produce inhibitors with improved specificity for XR.     

Echocardiographic assessment of LV hypertrophy and function in aortic-banded mice: necropsy validation

We characterized the time course of the left ventricular (LV) geometric and functional changes after aortic banding, validated them by necropsy, and investigated the sensitivity of echocardiographic findings on LV hypertrophy. C57BL/6 mice were subjected to transverse aortic constriction (TAC) or sham operation; echocardiographic assessments were performed before or at 2, 4, 6, and 11 wk after surgery; and some of the mice were euthanized at the corresponding time points. There was a progressive increase in diastolic posterior wall thickness and LV systolic dimension; the percentage of LV fractional shortening (LV%FS) decreased progressively at 4 wk, whereas these parameters remained stable in sham-operated mice. Echo LV mass and LV%FS correlated well with actual whole heart mass and ratio of lung weight to body weight, respectively (r = 0.765 and -0.749, respectively; P < 0.0001). These results suggest that the development of myocardial hypertrophy and systolic dysfunction is a time-dependent process. Echocardiographic assessment of myocardial hypertrophy and functional changes correlate well with the actual heart mass and lung mass. Echocardiography is sensitive enough to assess myocardial hypertrophy and heart functional changes induced by pressure overload in mice.     

An investigation of the intracellular site of anthocyanoplasts using isolated protoplasts and vacuoles

Microscopic observations made during preparation of protoplasts and vacuoles from red radish seedling hypocotyl (Raphanus sativus L.) show that anthocyanoplasts, the strongly pigmented bodies present in the pigmented cells of the hypodermis, begin as apparently membranous vesicles in the cytoplasm made visible by the deposition and accumulation of anthocyanins, but only rarely appear in the isolated vacuole. Isolation of protoplasts and vacuoles was also achieved from mung bean seedling hypocotyl (Vigna radiata L Wilczek), red cabbage leaf (Brassica oleracea L.) and Prunus x yedoensis Matsum callus. Anthocyanoplasts were usually in the vacuole, although sometimes in the cytoplasm, of the mung bean and cabbage, but were never seen in vacuoles of Prunus callus.     

A mutation in the gene for trigger factor suppresses the defect in cell division in the divE42 mutant of Escherichia coli K12

A total of sixteen spontaneously generated, independent suppressor mutants was isolated from a mutant (divE42) of Escherichia coli K12 that is defective in cell division. One of the suppressor mutants, designated TR4, had a novel phenotype: it was able to grow at 42° C but not at 32° C. The Kohara genomic library was screened for complementing clones. Clone 148 was able to complement the mutation responsible for the cold-sensitive phenotype, and the gene for trigger factor (tig), which encodes a ribosome-associated peptidyl-prolyl cis/trans isomerase, was identified as the mutated gene by deletion analysis with the insert DNA from clone 148. DNA sequencing revealed that the mutation in the tig gene of the TR4 suppressor mutant was a single nucleotide insertion (+A) at a distance of 834 nucleotides from the initiation codon for this enzyme. When the wild-type tig gene was introduced into the TR4 suppressor mutant, the bacteria were able to grow at 32° C but not at 42° C, an indication that the intergenic suppressor mutation was recessive to the wild-type allele. A model is proposed that accounts for the phenotypes of the divE42 mutant and the TR4 suppressor mutant. Received: 3 March 1998 / Accepted: 14 July 1998     

Ecological Studies on Reovirus Pollution of Rivers in Toyama Prefecture. II. Molecular Epidemiological Study of Reoviruses Isolated from River Water

In order to clarify the source of reovirus pollution in river water, comparative surveys have been carried out between reovirus isolates from river water and those from sewage, human or animal, by making use of the analysis of genomic RNA-migration pattern of reovirus in polyacrylamide gel electrophoresis (electropherotype). The strains of reovirus serotype 1 and 2 isolated from river water were classified into 3 and 9 electropherotypes, respectively, and 8 out of these 12 types were also found among strains isolated from sewage or human. When the monthly distribution of the river isolates classified by electropherotypes was compared with that of the sewage isolates, there were cases in which strains of the same electropherotype were simultaneously isolated from both sources. The electropherotypes of 3 isolates from pig and field rodents were different from those of the other isolates. The electropherotype of an oyster isolate coincided with that of some of the isolates from humans and river water. These results indicate that the major sources of reoviruses polluting river water may be the human excretion.     

Isolation of a new flavonol glycoside and its effects on the blue color of seed coats ofOphiopogon jaburan

From the blue seed coats ofOphiopogon jaburan, a new flavonol glycoside was isolated as needles and determined to be kaempferol 3-O-β-d-galactoside-4′-O-β-d-glucoside (OK-2) by UV and NMR spectral analyses. OK-2 and kaempfrol 3, 4′-di-O-β-d-glucoside (OK-1), which was detected previously, in the blue seed coat were present in a molar ratio of about 13:7. OK-2 was newly found as a factor causing the blueing effects on ophionin which is a main anthocyanin in the blue seed coats. The mixture of 4.8×10⁻³ M OK-2 and 2.5×10⁻³ M ophionin in Mcllvaine's buffer solution (pH 5.6) showed stable blue color, and the absorption spectrum of the mixture showed two absorption peaks and a shoulder in visible reasion, coinciding with that of the fresh blue seed coat. The effect of ophionin and OK-2 co-pigmentation on the blue color of seed coat ofO. jaburan was discussed.     

Characterization of two different factors chemotactic for cancer cells from tumor tissue.

Two different factors chemotactic for cancer cells were extracted in pseudoglobulin fraction of rat ascites hepatoma transplanted tissue. After chromatography on Sephadex G-50 and CM-sephadex, these factors were separated by gel filtration on Sephadex G-100. The factor a was further fractionated by immunoadsorbent chromatography with goat antirat gamma-globulin antibody and then with rabbit antirat hemoglobin antibody; it was a protein with a molecular weight of about 78,000, resembling a chemotactic factor previously reported, and its activity was thermolabile. The previously undescribed factor b was also a protein with a molecular weight of about 14,000 and its activity was thermostable. Intradermal injection of these factors at low concentrations induced an extravascular migration of circulating tumor cells and formation of metastatic secondary tumors; and little difference in the in vivo effect between these factors was observed. It was thus assumed that the combined action of these two factors may be involved in malignant invasion.     

The nucleotide sequence of Bacillus subtilis tRNA genes

Clones carring Bacillus subtilis tRNA genes were isolated from a lambda 816 library. A recombinant phage lambda 816-BS83 which was hybridized effectively with unfractionated tRNA probes contained a 3-kb fragment. By a Southern's blot analysis, it was found that tRNA genes were located in Eco RI-Hinc II region of this fragment. Sequence determination revealed the presence of a cluster of four tRNA genes in this region. The gene organization was as follows: tDNALys-9bp-tDNAGlu-81bp-tDNAAsp-30bp-tDNAPhe. The RNA sequences expected from tDNALys and tDNAPhe were identical with the reported RNA sequences. Two tRNA genes, tDNALys and tDNAAsp encoded the CCA sequence of 3'-terminal region, but the other two, tDNAGlu and tDNAPhe did not. A promoter-like sequence which corresponds to the sigma 55-recognition site was found in a region about 100bp upstream from tDNALys.