6-Oxodendrolasin, 6-hydroxydendrolasin, 9-oxofarnesol and 9-hydroxyfarnesol,stress metabolites of the sweet potato

Four sesquiterpene stress metabolites, 6-oxodendrolasin, 6-hydroxydendrolasin, 9-oxofarnesol, and 9-hydroxyfarnesol have been isolated from mercuric chloride-treated sweet potatoes. The metabolites have been synthesized and feeding studies have been carried out to determine the extent of incorporation of ¹⁴C-labelled 6-oxodendrolasin and 9-hydroxyfarnesol into ipomeamarone.     

In vitro effects of human recombinant alpha-2b interferon on Ph1+ chronic myelogenous leukemia cells maintained in long term marrow cultures: a functional and morphological analysis

In this study in vitro results obtained with hu rec IFN-alpha 2b on Ph1+ stem cells from patients with chronic myelogenous leukemia in chronic phase (CML in CP) will be discussed: cells were incubated with different IFN concentrations (100, 1000, 10000 IU/ml) for different times (24, 96 hrs, 8, 15, days) and maintained in long term marrow cultures (LTMC); CFU-GM assay, cytochemistry and cytogenetic analyses were performed weekly. A high sensitivity of CML cells to the in vitro treatment with IFN was observed. Cell count in LTMC showed a progressive reduction inversely proportional to time of incubation and concentration of IFN; a marked decrease in colony growth was observed at the end of incubations and during the course of LTMC. Low concentrations of IFN permitted a morphological maturation and the expression of alkaline phosphatase. Cytogenetic analyses showed a marked reduction of mytoses in cultures treated with high concentrations of IFN as result of a combined cytostatic and cytolitic effect; the persistance of 100% Ph1+ cells in LTMC and in CFU-GM colonies might be related, as opposed to in vivo results, to different IFN exposure conditions or might be influenced by other factors.     

Binding changes and apparent pK a shifts of bromthymol blue as tools for mitochondrial reactions

The passive interaction of bromthymol blue and other anionic and neutral dyes with mitochondria and particles is accompanied by an increase of the apparent pK_a, is enhanced at higher ionic strengths, is slightly inhibited by neutral dyes, is high only for the acidic, un-ionized form, and is independent of the presence of the sulfonic side chain. The pH dependence for the binding of the dyes follows the ionization of the chromophoric group: the pK of binding, defined as the pH for 50% binding, is identical with the apparent pK_a of the dye.     

Oligosaccharides in Five Different Vicia faba L. Cultivars

The oligosaccharide composition in seeds of some bean cultivars (two of Vicia faba L. and three of V. faba minor) have been examined. The sugars were isolated by solvent extraction and the concentrated extracts were analysed by PC and HPLC analyses. Characterization was also performed by thin-layer chromatography, enzymatic hydrolysis assays, GC determination of alditol acetate derivatives obtained after complete acid hydrolysis and FAB-MS of individual sucrose - galactosides. Although sucrose, raffinose, stachyose and verbascose were always identified, great differences in the total sugar content were found among the five cultivars. Sucrose and verbascose were the major components. Their contents were highest in the Muchamiel cv. (2.30 and 3.05 g/100g of dry bean, respectively). The selection of cultivars is the best way to improve the V. faba quality     

Mitochondrial oscillation and activation of H+/cation exchange

Mitochondria incubated aerobically in the presence of tetrapropylammonium and weak acids and in the presence of trace amounts of tetraphenylboron undergo a series of damped oscillations reflecting cycles of osmotic swelling and shrinkage. The matrix volume changes are consequent to transport of tetrapropylammonium catalytically stimulated by tetraphenylboron. The amplitude and frequency of the oscillations increase with the concentration of tetrapropylammonium, as required for critical rates and extents of ion influx. Addition of bovine serum albumin abolishes both the uptake of tetrapropylammonium and the oscillations. Volume oscillations are paralleled by cyclic activation and depression of the respiratory rate. Two lines of evidence suggest that the train of damped oscillations depends on the cyclic activation of an electroneutral exchange of H⁺ with organic cations rather than on cyclic uncoupling. First, further increase of cation permeability due to a pulse of tetraphenylboron, after initiation of cation efflux, restores cation influx. Second, addition of Mg²⁺, which abolishes the oscillations, has a much more marked inhibitory effect on the process of cation efflux than on cation influx. Conversely, addition of A23187, which removes membrane-bound Mg²⁺, promotes cation efflux and thus the oscillations. It is suggested that, in the present system, stretching of the inner membrane and Mg²⁺ depletion result in activation of an electroneutral H⁺/organic cation exchange, and that cyclic activation of this reaction results in damped oscillations.     

Characterization of a recombinant N-acetylgalactosamine-6-sulfate sulfatase produced in E. coli for enzyme replacement therapy of Morquio A disease

Mucopolysaccharidosis IVA (MPS IVA) is a lysosomal storage disease caused by the deficiency of N-acetylgalactosamine-6-sulfate sulfatase (GALNS) enzyme. Currently, specific therapies are not available for MPS IVA patients. In this study, a biologically active recombinant GALNS enzyme (rGALNS) produced in Escherichia coli was purified through a two-step chromatography process. The effect of temperature and pH on purified rGALNS stability was evaluated, as well as the stability in human serum. Finally, the uptake of rGALNS by HEK 293 cells and MPS IVA fibroblasts was evaluated. The use of a semi-continuous process allowed the production of an active extracellular rGALNS, which was used for protein purification. The purified rGALNS showed a specific activity of 0.29 U mg^?1 and a production yield of 0.78 mg L^?1. The rGALNS presented an optimal pH of 5.5 and was stable for 8 days at 4 °C. In human serum it was stable for up to 6 h. rGALNS was not taken up by the cultured cells, suggesting that N-linked oligosaccharides are not necessary for the production of an active enzyme or enzyme stability but for the cell uptake of protein. This study shows the first characterization of rGALNS produced by E. coli, and provides important information about purification, stability, and glycosylations effect for this type of enzymes.