Untersuchungen über den Vitamin B2-Bedarf der Legehenne

Because of the well established function of carnitine possible effects of carnitine were studied in poultry. In trial I it was investigated if carnitine and its precursors (lysine, methionine) reduce the formation of abdominal fat in broilers. Chickens (10 groups of 10 chickens each) were fed different diets (control, lysine and methionine in excess and deficient, respectively, with or without 5% fat supplement, L‐carnitine and DL‐carnitine supplement, respectively).Performance (body weight gain, feed conversion), amount of abdominal fat and carnitine concentration in blood, muscles (M. sartorius, M.pectoralis superficialis, cardiac), liver and kidney were determined. Performance and abdominal fat were influenced by dietary fat, lysine and methionine as expected and were not altered by carnitine. Excess and deficiency of lysine and methionine did not influence, fat supplement reduced and carnitine supplementation significantly increased tissue concentration of carnitine.

In trial II it was studied if supplementation of a commercial layers’ ration with either 500 mg L‐carnitine or 500 mg nicotinic acid or both per kg reduces the cholesterol concentration in yolk. Influence on body weight, feed intake, laying performance, serum and yolk cholesterol concentration could not be observed, but yolk concentration of carnitine was significantly increased in supplemented groups.

Trial III should clarify if the L‐carnitine content in broiler parentstock ration influences hatchability. Four groups of 1350 hens each were fed a commercial all‐mash supplemented with 0, 20, 50 and 100 mg L‐carnitine, respectively. Hatching rate was increased from 83% to 87% and from 82.4% to 85.3% in groups supplemented with 50 and 100 mg L‐carnitine, respectively, and in randomly sampled eggs of these groups carnitine concentration in yolk was higher.     

Short‐Term Water Deficit Changes Cuticular Sterol Profile in the Eggplant (Solanum melongena)

Crop irrigation uses a majority of a total world water supply, at the same time displaying low efficiency. As the expected, future water requirements are higher than the current ones; there is a risk of a growing deficit of water for the agricultural use. Hence, there is an arising need for better understanding the effects of water deprivation on the crop plants. Eggplant (Solanum melongena L.) is a vegetable crop cultivated in arid and semi‐arid parts of the world. Because of its high water demands, the eggplant is a convenient model organism for studies concerning the effects of water deficit on the plant growth. The objective of the study was to determine the impact of short‐term water deficit on eggplant leaf cuticular waxes and total sterols. Water deprivation did not affect the amount and composition of aliphatic components of cuticular waxes. Significant decrease in the total cuticular sterols and the increase in cuticular cholesterol were observed as an effect of water deficit. In contrast, some of the free internal sterols were more abundant in water‐deprived plants. The possible importance of these observations, including increased biosynthesis of defensive compounds and the need to maintain the cell membrane stability, was discussed.     

Lipid-based transfection reagents can interfere with cholesterol biosynthesis

Lipid-based transfection reagents are widely used for delivery of small interfering RNA into cells. We examined whether the commonly used commercial transfection reagents DharmaFECT-4 and Lipofectamine 2000 can interfere with lipid metabolism by studying cholesterogenesis. Cholesterol de novo synthesis from [¹⁴C]acetate was assessed in human hepatocyte-derived Huh-7 cells. The results revealed that DharmaFECT, but not Lipofectamine, markedly inhibited cholesterol biosynthesis by approximately 70%. Cell viability was not significantly altered. These findings suggest that caution is required in the choice of certain lipid-based transfection reagents for gene silencing experiments, particularly when assessing cholesterol metabolism.     

Immunomodulation of carcinogens-induced steroids-dependent human diseases

The experimental and clinical data about antibodies against environmental chemical carcinogens and endogenous steroids are represented. The conception of immunomodulation of carcinogens- and steroids-dependent human diseases is proposed. It is postulated that antibodies to polycyclic aromatic hydrocarbons and heterocyclic amines in cooperation with antibodies to cholesterol, sex hormones, mineralo- and glucocorticoids stimulate or inhibit cancer, malformation, cardiovascular and autoimmune diseases depending on their personal combination. It is recommended to use immunoassay of these antibodies for the human diseases prediction. The alternative approaches for prevention using the probiotics transformed by anti-carcinogen antibodies are substantiated.     

Feeding in the frequency domain: coarser‐grained environments increase consumer sensitivity to resource variability,covariance and phase

Theory predicts that resource variability hinders consumer performance. How this effect depends on the temporal structure of resource fluctuations encountered by individuals remains poorly understood. Combining modelling and growth experiments with Daphnia magna, we decompose the complexity of resource fluctuations and test the effect of resource variance, supply peak timing (i.e. phase) and co‐limiting resource covariance along a gradient from high to low frequencies reflecting fine‐ to coarse‐grained environments. Our results show that resource storage can buffer growth at high frequencies, but yields a sensitivity of growth to resource peak timing at lower ones. When two resources covary, negative covariance causes stronger growth depression at low frequencies. However, negative covariance might be beneficial at intermediate frequencies, an effect that can be explained by digestive acclimation. Our study provides a mechanistic basis for understanding how alterations of the environmental grain size affect consumers experiencing variable nutritional quality in nature.     

Modifying bio-catalytic properties of enzymes for efficient biocatalysis: a review from immobilization strategies viewpoint

Enzyme-based catalysis has become one of the most important disciplines in organic synthesis and plays a noteworthy role in the establishment of many chemical industries, e.g. fine chemicals, food or energy, textiles, agricultural, cosmeceutical, medicinal and pharmaceutical industries. However, pristine enzymes fail to demonstrate requisite functionalities for an industrial setting where extremely specific and stable catalysts are required. Immobilization enhances the catalytic stability and activity of enzymes and trims the overall cost burden of the enzyme. Therefore, it widely endeavours for proficient, sustainable, and environmentally responsive catalytic processes. Amongst several immobilization strategies, e.g. (1) supports-assisted, i.e. physical or covalent coupling and (2) supports-free techniques, i.e. cross-linked enzyme crystals (CLECs) or aggregates are the most promising ones and widely pursued for enzyme immobilization purposes. This perspective review focuses on up-to-date developments in the area of enzyme immobilization and presents their potentialities to upgrade and/or modify enzyme properties. Both types of immobilization strategies, i.e. supports-assisted and supports-free techniques are discussed with particular reference to CLECs or aggregates and protein-coated microcrystals. Also, several useful traits achieved after immobilization are also discussed in the second half of the review.     

孕烷X受体配体结合结构域蛋白晶体的X线衍射结构解析

目的:利用X线衍射技术解析孕烷X受体(PXR)配体结合结构域(LBD)蛋白晶体的3维结构。方法:对PXR蛋白LBD(130~434氨基酸残基)序列进行密码子优化并化学合成后克隆至pRSFDuet-1表达载体,再将载体导入大肠杆菌BL21(DE3),对PXR-LBD蛋白进行原核表达与分离纯化;采用晶体筛选试剂盒筛选蛋白结晶条件,采用悬滴法获得目标蛋白的晶体;对获得的蛋白晶体进行X线晶体衍射检测,并收集相关数据建立PXR-LBD的三维结构。结果:获得了PXR-LBD的高质量晶体并利用X线衍射解析了该蛋白质晶体的结构数据,使用Phenix.refine软件和COOT软件等对结构进行修正,最终获得了高分辨率的3维结构数据。结论:完成了孕烷X受体配体结合结构域蛋白晶体的X线衍射结构解析,为研究和开发PXR相关药物奠定了基础。     

A cell-based high-throughput screen identifies tyrphostin AG 879 as an inhibitor of animal cell phospholipid and fatty acid biosynthesis

Inhibition of animal cell phospholipid biosynthesis has been proposed for anticancer and antiviral therapies. Using CHOK1 derived cell lines, we have developed and used a cell-based high-throughput procedure to screen a 1280 compound, small molecule library for inhibitors of phospholipid biosynthesis. We identified tyrphostin AG 879 (AG879), which inhibited phospholipid biosynthesis by 85–90% at a concentration of 10 μM, displaying an IC₅₀ of 1–3 μM. The synthesis of all phospholipid head group classes was heavily affected. Fatty acid biosynthesis was also dramatically inhibited (90%). AG879 inhibited phospholipid biosynthesis in all additional cell lines tested, including MDCK, HUH7, Vero, and HeLa cell lines. In CHO cells, AG879 was cytostatic; cells survived for at least four days during exposure and were able to divide following its removal. AG879 is an inhibitor of receptor tyrosine kinases (RTK) and inhibitors of signaling pathways known to be activated by RTK's also inhibited phospholipid biosynthesis. We speculate that inhibition of RTK by AG879 results in an inhibition of fatty acid biosynthesis with a resulting decrease in phospholipid biosynthesis and that AG879's effect on fatty acid synthesis and/or phospholipid biosynthesis may contribute to its known capacity as an effective antiviral/anticancer agent.     

High resolution NMR for studying lipid hydrolysis and esterification in cod (Gadus morhua) gonads

High resolution NMR was applied to study biochemical changes of lipids in cod (Gadus morhua) gonads during 7 days storage at 4 degrees C. Changes were observed in the (13)C and (1)H resonances of cholesterol which were due to esterification of fatty acids at the hydroxyl position in roe and milt. Furthermore, the (13)C NMR spectra showed that the lipolytic changes in milt and roe where different. New resonances appeared during storage, due to formation of specific free fatty acids, with the corresponding changes in resonances of the esterified carbonyls and glycerols. The highly unsaturated n-3 fatty acids were hydrolysed from the sn-1 and sn-2 position of both phosphatidylcholine and phosphatidylethanolamine in milt. The lipolytical changes in roe were less prominent compared to the changes in milt, however significant levels of sn-1-lysophospholipids was detected both in roe and milt. The current data demonstrate that high resolution NMR may be a suitable method to non-destructively study hydrolysis and esterification reactions occurring in heterogeneous marine lipids in a one step procedure.     

Cholesterol depletion alters detergent-specific solubility profiles of selected tight junction proteins and the phosphorylation of occludin

Differential centrifugation of Triton X-100 or CHAPS lysates from control and cholesterol (CH)-depleted MDCK II cells, segregated integral tight junction (TJ) proteins associated with detergent-resistant membranes (DRMs) into two groups. Group A proteins (occludin, claudin-2 and -3) were detected in large, intermediate and small aggregates in both detergents, whereas group B proteins (claudin-1, -4 and -7) were observed in small aggregates in TX-100 and in intermediate and small aggregates in CHAPS. Depletion of CH altered the distribution of group A and B proteins among the three size categories in a detergent-specific manner. In lysates produced with octyl glucoside, a detergent that selectively extracts proteins from DRMs, group A proteins were undetectable in large aggregates and CH depletion did not alter the distribution of either group A or B proteins in intermediate or small aggregates. Neither occludin (group A) nor claudin-1 (group B) was in intimate enough contact with CH to be cross-linked to [(3)H]-photo-cholesterol. However, antibodies to either TJ protein co-immunoprecipitated caveolin-1, a CH-binding protein. Unlike claudins, occludin's presence in TJs and DRMs did not require palmitoylation. Equilibrium density centrifugation on discontinuous OptiPrep gradients revealed detergent-related differences in the densities of TJ-bearing DRMs. There was little or no change in those densities after CH depletion. Removing CH from the plasma membrane increased tyrosine and threonine phosphorylation of occludin, and transepithelial electrical resistance (TER) within 30 min. After 2 h of CH efflux, phospho-occludin levels and TER fell below control values. We conclude that the association of integral TJ proteins with DRMS, pelleted at low speeds, is partially CH-dependent. However, the buoyant density of TJ-associated DRMs is a function of the detergent used and is insensitive to decreases in CH.