Production of Acidic Actinomycin Congeners by a Mutant Strain of Streptomyces antibioticus,No. B-1625

Streptomyces sp. No. B-1625, which was identified as a strain of Streptomyces antibioticus, is a typical producer of actinomycin, but also produces minor acidic antibiotic components (FA), besides actinomycins X₂, D and X_0β. The FA-components, which were obtained with a high-producing mutant, 11M-21, showed antibacterial and antitumor activities, and also similar visible and UV absorption spectra to those characteristic of actinomycin. The FA-components were separated into five components, FA₁ FA_2α, FA_2β, FA_3α and FA_3β, on TLC. Among them, one component, FA_3β, isolated in a purified state as an orange powder, has a composition of C, 52.97: H, 6.34: N, 10.48%, and is active against B. subtilis at a MIC of 5mcg/ml. The FA_3β component showed pK_a′ of 5.4 and 12.0 and λ_max at 443, 427 and 233 nm. From these properties, FA_3β is considered to be an acidic actinomycin congener.     

DNA Inversion Regulates Outer Membrane Vesicle Production in Bacteroides fragilis

Phase changes in Bacteroides fragilis, a member of the human colonic microbiota, mediate variations in a vast array of cell surface molecules, such as capsular polysaccharides and outer membrane proteins through DNA inversion. The results of the present study show that outer membrane vesicle (OMV) formation in this anaerobe is also controlled by DNA inversions at two distantly localized promoters, IVp-I and IVp-II that are associated with extracellular polysaccharide biosynthesis and the expression of outer membrane proteins. These promoter inversions are mediated by a single tyrosine recombinase encoded by BF2766 (orthologous to tsr19 in strain NCTC9343) in B. fragilis YCH46, which is located near IVp-I. A series of BF2766 mutants were constructed in which the two promoters were locked in different configurations (IVp-I/IVp-II = ON/ON, OFF/OFF, ON/OFF or OFF/ON). ON/ON B. fragilis mutants exhibited hypervesiculating, whereas the other mutants formed only a trace amount of OMVs. The hypervesiculating ON/ON mutants showed higher resistance to treatment with bile, LL-37, and human β-defensin 2. Incubation of wild-type cells with 5% bile increased the population of cells with the ON/ON genotype. These results indicate that B. fragilis regulates the formation of OMVs through DNA inversions at two distantly related promoter regions in response to membrane stress, although the mechanism underlying the interplay between the two regions controlled by the invertible promoters remains unknown.     

Quantitative examination of a perfusion microscope for the study of osmotic response of cells

The perfusion microscope was developed for the study of the osmotic response of cells. In this microscope, the cells are immobilized in a transparent chamber mounted on the stage and exposed to a variety of milieus by perfusing the chamber with solutions of different concentrations. The concentration of the supplied solution is controlled using two variable-speed syringe pumps, which supply an isotonic solution and a hypertonic solution. Before using this system to characterize the osmotic response of cells, the change in the concentration of NaCl solution flowing through the chamber is examined quantitatively using a laser interferometer and an image processing technique. The NaCl concentration is increased from an isotonic condition to a hypertonic condition abruptly or gradually at a given constant rate, and decreased from a hypertonic condition to an isotonic condition. It is confirmed that the concentration is nearly uniform in the cross direction at the middle of the chamber, and the change in the NaCl concentration is reproducible. The average rate of increase or decrease in the measured concentration agrees fairly well with the given rate when the concentration is changed gradually at a constant rate. The rate of the abrupt change is also determined to be the highest limit achieved by the present method. As the first application of using the perfusion microscope for biological studies, the volume change of cells after exposure to a hypertonic solution is measured. Then, the hydraulic conductivity of the cell membrane is determinedfrom the comparison of the volume change between the experiment and the theoretical estimation for the measured change in the NaCl concentration of the perfused solution.     

Transformation of Aspergillus aculeatus using the drug resistance gene of Aspergillus oryzae and the pyrG gene of Aspergillus nidulans

Transformation systems for Aspergillus aculeatus has been developed, based on the use of the pyrithiamine resistance gene of Aspergillus oryzae and the orotidine-5'-monophosphate decarboxylase gene (pyrG) of Aspergillus nidulans. An A. aculeatus mutant which can be transformed effectively by the A. nidulans pyrG gene was isolated as a transformation host. This is the first report of transformation of A. aculeatus.     

neurotic,a novel maternal neurogenic gene,encodes an O-fucosyltransferase that is essential for Notch-Delta interactions

Notch signalling, which is highly conserved from nematodes to mammals, plays crucial roles in many developmental processes. In the Drosophila embryo, deficiency in Notch signalling results in neural hyperplasia, commonly referred to as the neurogenic phenotype. We identify a novel maternal neurogenic gene, neurotic, and show that it is essential for Notch signalling. neurotic encodes a Drosophila homolog of mammalian GDP-fucose protein O-fucosyltransferase, which adds fucose sugar to epidermal growth factor-like repeats and is known to play a crucial role in Notch signalling. neurotic functions in a cell-autonomous manner, and genetic epistasis tests reveal that Neurotic is required for the activity of the full-length but not an activated form of Notch. Further, we show that neurotic is required for Fringe activity, which encodes a fucose-specific beta1, 3 N-acetylglucosaminyltransferase, previously shown to modulate Notch receptor activity. Finally, Neurotic is essential for the physical interaction of Notch with its ligand Delta, and for the ability of Fringe to modulate this interaction in Drosophila cultured cells. We present an unprecedented example of an absolute requirement of a protein glycosylation event for a ligand-receptor interaction. Our results suggest that O-fucosylation catalysed by Neurotic is also involved in the Fringe-independent activities of Notch and may provide a novel on-off mechanism that regulates ligand-receptor interactions.     

New PEG2 type polyethylene glycol derivatives for protein modification

Although proteins with 2,4-bis (o-methoxypolyethylene glycol)-6-chloro-s-triazine (PEG2-Cl) as a divalent PEG modification have some advantages compared to proteins with the linear PEG modification, PEG2Cl cannot react with amino groups at neutral pH. Therefore, we have prepared new PEG2 derivatives that have an activated ester as the functional group. We confirmed that these derivatives are useful for the divalent modification of proteins, such as bSOD and rhG-CSF. © Rapid Science Ltd. 1998     

Methane production from rice straw pretreated by a mixture of acetic–propionic acid

Rice straw was treated with a mixed solution of acetic acid and propionic acid to enhance its biodegradability. The effect of acid concentration, pretreatment time, and the ratio of solid to liquid on the delignification performance of rice straw were investigated. It was found that the optimal conditions for hydrolysis were 0.75 mol/L acid concentration, 2 h pretreatment time and 1:20 solid to liquid ratio. Batch methane fermentation of untreated rice straw, pretreated rice straw, and the hydrolysates (the liquid fraction) of pretreatment were conducted at 35 °C for 30 days, and the results indicated that methane production of rice straw can be enhanced by dilute organic acid pretreatment. Moreover, most of the acid in hydrolysates can also be converted into methane gas.