Effect of synthetic lipids on apoptosis and expression of alkaline phosphatase in B-lymphocytes: influence on lipopolysaccharide action

Synthetic lipids were examined for their ability to mimic or to antagonize lipopolysaccharide (LPS) action in murine B-lymphocytes. Several recognized effects of LPS were analyzed: prevention of spontaneous apoptosis, expression of alkaline phosphatase (ALP) and stimulation of proliferation. Three synthetic lipids were used for that purpose: a lipopeptide (compound MTPP) which carries non-hydroxylated fatty acids, and is thus unrelated to LPS, and two glycolipids with hydroxylated fatty acids (compounds D2 and PPDm2-B), structurally related to the lipid A region of enterobacterial and Rhodopseudomonas LPS, respectively. We found that the ability of these lipids to induce LPS-like responses was not correlated with their structural analogy with LPS. Thus, the lipopeptide, MTPP, mimicked LPS in the three activities, whereas the glycolipid, D2, did not. In contrast, the ability of synthetic lipids to block LPS effects was correlated with their structural analogy with LPS. We thus observed that compound D2 selectively blocked LPS-induced ALP expression and that PPDm2-B selectively inhibited LPS-induced prevention of apoptosis. These synthetic lipids could therefore be useful for studying the LPS-mediated signals involved in B-cell activation and apoptosis.     

Alkaline phosphatase at the cell wall of the yeast phase ofParacoccidioides brasiliensis

The activity of alkaline phosphatase demonstrated by histochemical techniques was shown at the cell wall of the yeast form ofParacoccidioides brasiliensis at 3, 6, and 9 days of culture. The results showed a very active deposition at the cell wall as early as 9 days of culture of the fungus which made us think an inactive salt precipitate was also present.     

Rewired phenolic metabolism and improved saccharification efficiency of a Zea mays cinnamyl alcohol dehydrogenase 2 (zmcad2) mutant

Lignocellulosic biomass is an abundant byproduct from cereal crops that can potentially be valorized as a feedstock to produce biomaterials. Zea mays CINNAMYL ALCOHOL DEHYDROGENASE 2 (ZmCAD2) is involved in lignification, and is a promising target to improve the cellulose-to-glucose conversion of maize stover. Here, we analyzed a field-grown zmcad2 Mutator transposon insertional mutant. Zmcad2 mutant plants had an 18% lower Klason lignin content, whereas their cellulose content was similar to that of control lines. The lignin in zmcad2 mutants contained increased levels of hydroxycinnamaldehydes, i.e. the substrates of ZmCAD2, ferulic acid and tricin. Ferulates decorating hemicelluloses were not altered. Phenolic profiling further revealed that hydroxycinnamaldehydes are partly converted into (dihydro)ferulic acid and sinapic acid and their derivatives in zmcad2 mutants. Syringyl lactic acid hexoside, a metabolic sink in CAD-deficient dicot trees, appeared not to be a sink in zmcad2 maize. The enzymatic cellulose-to-glucose conversion efficiency was determined after 10 different thermochemical pre-treatments. Zmcad2 yielded significantly higher conversions compared with controls for almost every pre-treatment. However, the relative increase in glucose yields after alkaline pre-treatment was not higher than the relative increase when no pre-treatment was applied, suggesting that the positive effect of the incorporation of hydroxycinnamaldehydes was leveled off by the negative effect of reduced p-coumarate levels in the cell wall. Taken together, our results reveal how phenolic metabolism is affected in CAD-deficient maize, and further support mutating CAD genes in cereal crops as a promising strategy to improve lignocellulosic biomass for sugar-platform biorefineries.     

Mechanisms of glucagon degradation at alkaline pH

Glucagon is unstable and undergoes degradation and aggregation in aqueous solution. For this reason, its use in portable pumps for closed loop management of diabetes is limited to very short periods. In this study, we sought to identify the degradation mechanisms and the bioactivity of specific degradation products. We studied degradation in the alkaline range, a range at which aggregation is minimized. Native glucagon and analogs identical to glucagon degradation products were synthesized. To quantify biological activity in glucagon and in the degradation peptides, a protein kinase A-based bioassay was used. Aged, fresh, and modified peptides were analyzed by liquid chromatography with mass spectrometry (LCMS). Oxidation of glucagon at the Met residue was common but did not reduce bioactivity. Deamidation and isomerization were also common and were more prevalent at pH 10 than 9. The biological effects of deamidation and isomerization were unpredictable; deamidation at some sites did not reduce bioactivity. Deamidation of Gln 3, isomerization of Asp 9, and deamidation with isomerization at Asn 28 all caused marked potency loss. Studies with molecular-weight-cutoff membranes and LCMS revealed much greater fibrillation at pH 9 than 10. Further work is necessary to determine formulations of glucagon that minimize degradation and fibrillation.     

Production of an alkaline protease by Bacillus cereus MCM B-326 and its application as a dehairing agent

The present investigation describes microbial production of an alkaline protease and its use in dehairing of buffalo hide. Bacillus cereus produced extracellular protease when grown on a medium containing starch, wheat bran and soya flour (SWS). The ammonium sulphate precipitated (ASP) enzyme was applied for dehairing of buffalo hide. Microscopic observation of longitudinal section of buffalo hide revealed that the epidermis was completely removed and hair was uprooted leaving empty follicles in the hide. The ASP enzyme was stable for one month at ambient temperature between 25–35 °C. Enzymatic dehairing may be a promising shift towards an environment-friendly leather processing method.     

Arrowroot (Marantha arundinacea) starch as a new low-cost substrate for alkaline protease production

The feasibility of arrowroot (Marantha arundinacea) starch for alkaline protease production using an alkalophilic Bacillus lentus isolate was evaluated in batch fermentations in shake flasks and in a bioreactor under a range of conditions. A new arrowroot starch-casein medium (pH 10.2) was formulated having a composition (%, w/v): arrowroot starch 1, casein 1, sodium succinate 0.25, NH₄Cl 0.05, NaCl 0.05, KH₂PO₄ 0.04, K₂HPO₄ 0.03, MgCl₂ 0.01, yeast extract 0.01 and Na₂CO₃ 1.05. The isolate produced a maximum protease yield (6754.7 U ml^–1) in this medium when grown for 72 h at 250 rev/min and 37 °C. Scaling-up studies in a bioreactor showed a 5-fold increase in alkaline protease yields (31899 U ml^–1) at a lower production time of 45 h, aeration of 1 v/v/m and agitation of 400 rev/min at 37 °C.     

Seasonal feeding activity and ontogenetic dietary shifts in crucian carp,Carassius carassius

Synopsis A pronounced seasonal cycle in the activity of the intestinal alkaline phosphatase indicated a 6 month pause in feeding by crucian carp in an anoxic pond during winter. Zero enzyme activity was not reached until ice cover of the pond was complete, although even in the preceding two months no food was found in the guts of the fish. The summer diets of four size classes of fish consisted of varying proportions of planktonic and benthic arthropods, including Odonata which were consumed exclusively by the largest size class of crucian carp. Chironomid larvae were consumed even by the smallest fish investigated, although planktonic Cladocera were preferred by this size class. Benthic Cladocera were almost totally absent in the diet of fish> 10 cm. Habitat segregation between size classes developed synchronously with the shift to benthic food. The stunting of crucian carp populations in small ponds is interpreted as resulting from limited resources.     

Stressed Jerusalem artichoke tubers (Helianthus tuberosus L.) excrete a protein fraction with specific cytotoxicity on plant and animal tumour cell

Wounds from Jerusalem artichoke (Helianthus tuberosus L.) tubers excrete bioactive metabolites from a variety of structural classes, including proteins. Here we describe a protein specifically active against tumour cells arising either from human, animal or plant tissues. The non-tumour animal cells or the plant callus cells are not sensitive to these excreta. The active product was only obtained after a wound-drought stress of plant tubers. The cytotoxicity varies according to the tumour cell type. For instance, some human tumour cell lines and especially the human mammary tumour cells MDA-MB-231 were shown to be very susceptible to the active product. The active agent is shown to contain an 18-kDa polypeptide with homology to a superoxide dismutase (SOD). A 28-kDa polypeptide, related to an alkaline phosphatase (AP), was shown to be tightly linked to this 18-kDa polypeptide. The excreted 28-kDa polypeptide also displayed a consensus sequence similar to the group of DING proteins, but with a smaller molecular weight. The superoxide dismutase polypeptide was shown to be involved in the antitumour activity, but the presence of smaller factors (MW < 10 kDa), such as salicylic acid, can enhance this activity.     

Thermotolerant strain of Bacillus licheniformis producing lipase

A thermotolerant variant of Bacillus licheniformis strain H1 (isolated from Jordan valley soil), was highly active in degrading macromolecules, and possessed a lipase activity with a half life of 30 min at 70°C. This activity was produced during exponential growth. The extracellular crude lipase showed maximal activity at pH 10, and retained 65% of its stability at pH 12.The author is with the Department of Biological Sciences, University of Jordan, P.O. Box 2686, Amman 11181, Jordan