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Biomedical subjects

T Chojnacki

Publications and source records attributed to T Chojnacki.

At least 19 recordsLinked to original sources

Regulatory aspects of coenzyme Q metabolism.

A number of factors are involved in the regulation of the amount and distribution of coenzyme Q in cells and tissues. These factors modify preferentially the biosynthetic mechanism in order to keep up an optimal tissue concentration of the lipid. The amount of substrate provided by the mevalonate pathway is able to both up- and down-regulate the velocity of synthesis. At the translation level, regulation occurs by receptor-mediated ligand binding and appears most clearly upon treatment with hormones and peroxisomal inducers. There are a number of pathophysiological conditions when these mechanisms of regulation are modified and explain the decreased coenzyme Q tissue concentrations. It is of considerable interest to establish appropriate physiological, hormonal and drug-mediated conditions in order to counteract disturbed cellular functions caused by coenzyme Q deficiency

Aging↗

Search for polyprenols in leaves of evergreen and deciduous Ericaceae plants.

Various species and cultivars of Ericaceae family were checked for the presence of long-chain polyprenols in their leaves. In the genus Rhododendron no polyprenols were found in the ever-green species, while they were present in the deciduous type. The polyprenols were of chain-length of 14-20 isoprene residues and they occurred in the form of acetic acid esters. The polyprenol accumulation is discussed with respect to senescence of leaves.

Chromatography, High Pressure Liquid↗

Modulation of properties of phospholipid membranes by the long-chain polyprenol (C(160)).

The electrical measurements of phospholipid bilayers and the studies of phospholipid vesicles by using the transmission electron microscopy (TEM) showed that dotriacontaprenol (C(160)) isolated from leaves of Spermatophyta influences some properties of membranes. The current-voltage characteristics, the membrane conductance-temperature relationships, the membrane breakdown voltage and the membrane capacitance have been measured for different mixtures of C(160)/DOPC. The membrane conductance, the activation energy of ion migration across the membrane and the membrane thickness were determined. Dotriacontaprenol decreases the membrane breakdown voltage, the activation energy and the membrane capacitance, and increases the membrane conductance and the membrane hydrophobic thickness. The analysis of TEM micrographs shows several characteristic structures, which have been described. The results indicate that dotriacontaprenol increases the membrane elasticity and modulates the surface curvature of the membranes by the formation of fluid microdomains. We suggest that the long polyprenols facilitate the formation of transmembrane, ions-conductive pores.

Alkenes↗

Single polyprenol and dolichol isolation by semipreparative high-performance liquid chromatography technique.

A new method of separation of single polyprenols (or dolichols) from a mixture of isoprenoid alcohols is described. Application of a high-performance liquid chromatography (HPLC) apparatus equipped with a semipreparative ODS column resulted in preparation of long-chain (dihydro)polyprenols of high purity (>95%). This approach substantially decreases the time scale of the conventional chromatographical preparative procedure. The method can be widely used in chemical and biochemical projects, where single polyprenols or dolichols are required.

Chromatography, High Pressure Liquid↗

Polyprenols in hairy roots of Coluria geoides.

Long-chain polyisoprenoid alcohols built from several up to more than 100 isoprenoid units are common constituents of all living organisms. They were found mostly in plants, bacteria, yeasts and mammalian cells. In vitro hairy root culture of Coluria geoides was obtained from plants transformed with Agrobacterium rhizogenes. Growth was optimal at 0.75% (w/v) glucose and at 22 degrees C. Dry samples of roots were extracted and lipid content was analysed by HPLC. According to our estimation, polyprenols are accumulated in roots of C. geoides cultivated in vitro as a mixture of several prenologues with the dominating prenol composed of 16 isoprenoid units. The content of polyprenols in tissue was approx. 300 microg/g of dry weight.

Culture Techniques↗

Alterations in the biosynthesis of cholesterol, dolichol and dolichyl-P in the genetic cholesterol homeostasis disorder, Niemann-Pick type C disease.

The biosynthesis of cholesterol, dolichol and dolichyl-P were investigated in a murine model of Niemann-Pick type C disease using both in vitro and in vivo systems. In vivo incorporation of [3H]mevalonate into squalene, dolichol and dolichyl-P decreased. The amount of dolichyl-P was elevated due to a decrease in the rate of degradation. Labeling of squalene and cholesterol of liver homogenates in vitro was decreased in the diseased mice and a lowering of microsomal activities of both HMG-CoA reductase and squalene synthase were also observed. In experiments with brain homogenate, decreased [3H]mevalonate labeling of squalene, cholesterol and dolichol was found in vitro. The decreases in cis-prenyltransferase and squalene synthase activities were observed at a very early phase of the disease. In contrast to the decreased biosynthesis of cholesterol observed in vitro, the labeling of total liver cholesterol was found to be increased in Niemann-Pick type C liver upon in vivo investigation, possibly due to the accumulation of this lipid as a result of a deficient transport process. In the brain, where in vivo labeling reflects only biosynthesis, a decreased rate of cholesterol synthesis was demonstrated.

Animals↗

Dolichyl sulphate and H-phosphonate: enzymatic reactions with activated sugars.

Two phosphate-modified analogues of dolichyl phosphate were evaluated as substrates or inhibitors of the reactions catalyzed by mammalian microsomal enzymes. Dolichyl H-phosphonate could serve as an efficient acceptor for mannosyl and glucosyl transfer. The reaction products were chromatographically different from those formed from dolichyl phosphate. Lower activity of the H-phosphonate was observed for the reaction of N-acetylglucosaminyl phosphate transfer from UDP-GlcNAc. Dolichyl sulphate was shown not to serve as a substrate for the transfer of mannosyl (from GDP-Man), glucosyl (from UDP-Glc) or N-acetylglucosaminyl phosphate (from UDP-GlcNAc) residues in the presence of rat liver microsomes. Weak inhibitory properties of this analogue were demonstrated.

Animals↗

The diversity of polyprenol pattern in leaves of fruit trees belonging to Rosaceae and Cornaceae.

The polyprenol pattern in leaves of fruit trees belonging to the Rosaceae (genera: Prunus, Malus) and Cornaceae (genus: Cornus) families is presented. The content of polyprenyl acetates varied within plant species between 10-50 mg per gram of dry weight. In genus Prunus, Cornus and in representatives of species Malus domestica, a mixture of polyprenols composed of 18, 19, 20, 21 isoprene units was found. In six species of genus Prunus (sour-cherry): P. serrulata-spontanea, P. yedoensis, P. fruticosa. P. kurilensis, P. subhirtella and P. incisa the presence of a second polyprenol family, i.e. the group of prenologues consisting of prenol -35, -36, -37, etc. up to -42 was detected.

Chromatography, High Pressure Liquid↗

On the specific pattern of long chain polyprenols in green needles of Pinus mugo Turra.

In green needles of Pinus mugo the most abundant polyprenols occur as a mixture of prenologues in which the dominant alcohol is built of 16 isoprene units. The characteristic spectrum of polyprenols (prenol-15, -16 and -17) was the same irrespective of the location of plant and of distinct morphological differences observed in the various selected forms of this species. The constant pattern of the polyprenols spectrum was preserved throughout the 2-year life span of needles, although the level of polyprenols was increased 2-3-fold. The polyprenol pattern in Pinaceae family differs from species to species, thus it may serve as chemotaxonomic criterion within this systematic group.

Alcohols↗

Polyprenols in leaves of fruit-trees of Rosaceae family.

In leaves of various species of fruit-trees belonging to the Rosaceae family, large amounts of polyprenyl acetates (0.5-5.0% of dry weight) were found. Discrete constant differences of polyprenol spectrum characteristic of each genus studied: Malus, Prunus and Pyrus were observed. In each species poly-cis-prenols composed of 19 and 20 isoprene units were predominating. In one of the 23 studied species (Prunus incisa) a fraction of long-chain polyprenols composed of 35-45 isoprene units was also present. It seems that this type of unusually long-chain polyprenols could occur also in other plants of the Rosaceae family.

Chromatography, Thin Layer↗

The occurrence of long-chain polyprenols in leaves of plants of Combretaceae family.

The presence of poly-cis-prenols of chain length 20-60 isoprene units or longer in leaves of plants belonging to Combretaceae family was shown to be a common feature in this group of plants. The polyprenols of this type were found in half of the 20 species studied. In most cases the polyprenols occurred in the form of fatty acid esters. Only in one species--Combretum molle, the polyprenols were found in the form of free alcohols. The amount of long-chain polyprenols varied with the plant species; the richest source was C. molle (about 4% of dry mass of leaves). Polyprenol groups characteristic of other systematic families of plants were not found in the Combretaceae studied.

Chromatography, High Pressure Liquid↗

Distribution of branch point prenyltransferases in regions of bovine brain.

Bovine brains contain large amounts of isoprenoid compounds and the enzymes involved in their biosynthesis were investigated. Ten different regions were dissected from fresh bovine brains and, in addition, fractions from cerebellum, spinal cord, and hypophysis were obtained. The cholesterol concentration was found to be approximately 8 mg/g in the cortex regions and three times higher in the pons, medulla oblongata, and white matter. Dolichol concentration varied between 8 and 40 micrograms/g in the different tissues, and ubiquinone was found at a lower level, which varied between 3 and 25 micrograms/g. Farnesylpyrophosphate synthase activity in cytosolic fractions from various regions exhibited only a twofold variation, whereas geranylgeranyl pyrophosphate synthase displayed larger differences, being particularly rich in the pons, medulla oblongata, white matter, and spinal cord. Squalene synthase activity was lowest in the thalamus and threefold higher in the pons. Determination of specific activity based on cholesterol content revealed that enzyme activities in various regions are not related to the actual lipid amount present. Both cis- and trans-prenyltransferases exhibited similarities in their regional distribution showing up to 20-fold differences in activity. Thus, it appears that the mevalonate pathway lipids and the various branch point enzymes involved in their syntheses vary greatly in different brain regions and are subjected to separate regulation.

Animals↗

Recognition of the lipid intermediate for arabinogalactan/arabinomannan biosynthesis and its relation to the mode of action of ethambutol on mycobacteria.

Despite major advances in our understanding of the structure of mycobacterial cell walls, little is known of their biogenesis, and yet they are the site of action of many anti-tuberculosis drugs and implicated in much of the pathology of tuberculosis and leprosy. A family of monoglycosyl polyprenylphosphates was isolated from Mycobacterium smegmatis, containing arabinose, ribose, and mannose. The isoprenoid nature of the lipid components was established by 1H NMR, and fast atom bombardment mass spectroscopy (FAB-MS) demonstrated the presence of C50 decaprenyl-P derivatives and smaller amounts of the C35 octahydroheptaprenyl-P products. The configuration of the mycobacterial decaprenol was established as mono-trans, octa-cis, pointing to carriers of unusual structure. Combined gas chromatography (GC)/MS, FAB-MS/MS, and 1H NMR allowed characterization of one of the primary components as beta-D-arabinofuranosyl-1-monophosphodecaprenol. Pulse-chase metabolic labeling of cells with D-[14C]glucose indicated that the decaprenyl-P-arabinose is an active intermediate in the biosynthesis of the arabinan of cell wall arabinogalactan and arabinomannan. The identification of polyprenyl-P-ribose suggests the existence of ribose-containing polysaccharides in the cell walls of M. smegmatis or/and of a novel epimerase in the D-arabinose biosynthetic pathway. Ethambutol, a powerful anti-tuberculosis drug known to inhibit arabinogalactan and arabinomannan biosynthesis, results in the rapid accumulation of decaprenyl-P-arabinose, indicating that the drug interferes with either the transfer of arabinose from the donor or, alternatively, the synthesis of the arabinose acceptor itself.

Arabinose↗

Modulations in hepatic branch-point enzymes involved in isoprenoid biosynthesis upon dietary and drug treatments of rats.

Three branch-point enzymes of the mevalonate pathway, farnesyl pyrophosphate synthase, cis-prenyltransferase and squalene synthase were characterized in rat hepatic cytosol, microsomes and peroxisomes isolated from rats after treatment with peroxisome proliferators, inducers of the endoplasmic reticulum or modulators of lipid metabolism. Cholestyramine and phenobarbital induced primarily the cytosolic farnesyl pyrophosphate synthase, whereas clofibrate and phthalates elevated the corresponding peroxisomal activity. cis-Prenyltransferase activities in microsomes were induced 4-5-fold after clofibrate, phthalate and phenobarbital administration, but these same treatments affected the peroxisomal activity to only a limited extent. Squalene synthase activity in microsomes was completely abolished, but the peroxisomal activity was unaffected after administration of cholesterol. On the other hand, clofibrate and phthalate induced only the microsomal activities. Mevinolin treatment greatly increased peroxisomal and cytosolic farnesyl pyrophosphate synthase activities, but not the mitochondrial activity, and the cis-prenyltransferase activities were elevated in peroxisomes, but not in microsomes. These results demonstrate that the branch-point enzymes in cholesterol and dolichol biosynthesis at various cellular locations are regulated differentially and that the capacities of peroxisomes and the endoplasmic reticulum to participate in the synthesis of polyisoprenoid lipids is affected profoundly by treatment with different xenobiotics.

Animals↗

Isoprenoid biosynthesis in rat liver mitochondria. Studies on farnesyl pyrophosphate synthase and trans-prenyltransferase.

Mevalonate pathway enzyme activities in rat liver mitochondria were investigated, and it was found that isopentenyl pyrophosphate can be utilized for the synthesis of all-trans-polyprenyl pyrophosphates in vitro. In this reaction sequence intermediate formation of farnesyl pyrophosphate (FPP) predominates, and the FPP synthase activity was studied in more detail. The mitochondrial activity constitutes 13% of the total hepatic capacity for FPP synthesis, exceeding the corresponding microsomal, nuclear, and peroxisomal activities by 10-fold. Mitochondrial FPP synthase exhibits trypsin sensitivity only after sonication of intact mitochondria and upon subfractionation the activity is found localized in the matrix. FPP synthase activities at different locations responded distinctly when rats were treated with a diet enriched in cholesterol or containing mevinolin or cholestyramine. With the high cholesterol diet, mitochondrial FPP synthase activity increased 2-fold, while the cytosolic activity was slightly decreased. Both mevinolin and cholestyramine treatment resulted in 3-fold increases in cytosolic FPP synthase activities, without altering the mitochondrial activity. FPP was utilized as substrate for trans-prenyltransferase activity in the inner mitochondrial membrane. The products formed in this reaction were identified as nona- and decaprenyl-PP, and the reaction was influenced by changes in both substrate and Mg2+ concentration, giving more decaprenyl-PP when the concentrations of these substances were increased. These results demonstrate that mitochondria utilize endogenously produced FPP for isoprenoid biosynthesis and that the biosynthetic steps in mitochondria are regulated independently from those occurring in other subcellular compartments.

Animals↗

The effect of undecaprenol on bilayer lipid membranes.

The influence of undecaprenol on phosphatidylcholine macrovesicular bilayer lipid membranes has been studied by electrophysiological techniques. The current-voltage characteristics, ionic transference numbers, the membrane conductance-temperature relationships and the membrane breakdown voltage were measured. The permeability coefficients for Na+ and Cl- ions, the activation energy of ion migration across the membrane, the membrane hydrophobic thickness and the membrane Young's modulus were determined. Undecaprenol increases membrane conductance, membrane capacitance, membrane ionic permeability and membrane elastic deformability, decreases the activation energy, membrane hydrophobic thickness and membrane electromechanical stability, and does not change membrane selectivity. The formation by undecaprenyl molecules of fluid microdomains modulating membrane hydrophobic thickness is postulated. The data suggest that the behaviour of undecaprenol in membranes is regulated by transmembrane electrical potential.

Electric Conductivity↗