[Campylobacteriosis, a zooanthroponosis as an important cause of human diarrheal diseases].
Explore the source record for details and available documents.
Animals↗
Biomedical subjects
H Kühn
Publications and source records attributed to H Kühn.
Hydroperoxyfatty acids inactivate the reticulocyte lipoxygenase independently of a hydroperoxidase reaction.
From a comparison of 9Ds-HPODE and 13Ls-HPODE and their methyl esters as substrates and inactivating agents of reticulocyte lipoxygenase it is concluded that the compounds inactivate the enzyme independently of any hydroperoxidase reaction. The protective effect of 4-nitrocatechol indicates the formation of Fe(III) complexes of the enzyme with the hydroperoxyfatty acid compounds prior to inactivation.
Anaerobiosis↗
The stoichiometry of oxygen uptake and conjugated diene formation during the dioxygenation of linoleic acid by the pure reticulocyte lipoxygenase. Evidence for aerobic hydroperoxidase activity.
Simultaneous measurements of oxygen uptake and conjugated diene formation (increase in the absorbance at 234 nm) during the dioxygenation of linoleic acid by the pure reticulocyte lipoxygenase gave a nearly theoretical stoichiometry of 1.1 in a temperature range from 5 to 30 degrees C and a wide range of concentrations of both oxygen and linoleic acid. At low concentrations of either oxygen or linoleic acid or both, secondary processes occurred such as linoleic acid-supported lipohydroperoxidase reactions leading to the disappearance of conjugated dienes and to the formation of oxodienes, linoleic acid dimers and epoxyhydroxy derivatives. Under these conditions marked deviations of the stoichiometry between oxygen uptake and conjugated diene formation appeared. The formation of conjugated oxodienoic fatty acids absorbing at 285 nm occurred only under conditions of high concentrations of linoleic acid and limiting oxygen supply. The results indicate that lipohydroperoxidase reactions catalyzed by the pure reticulocyte lipoxygenase do not only take place under strictly anaerobic conditions but also under conditions of limiting concentrations of either linoleic acid or oxygen or both.
Aerobiosis↗
Enzymology and physiology of reticulocyte lipoxygenase: comparison with other lipoxygenases.
Explore the source record for details and available documents.
Animals↗
The stereochemistry of the reactions of lipoxygenases and their metabolites. Proposed nomenclature of lipoxygenases and related enzymes.
Explore the source record for details and available documents.
Animals↗
Activation of retinal rod cyclic GMP-phosphodiesterase by transducin: characterization of the complex formed by phosphodiesterase inhibitor and transducin alpha-subunit.
The GTP-binding subunit of transducin (T alpha) activates the cGMP phosphodiesterase (PDE) of bovine retinal rods by relieving the constraint imposed by the inhibitory subunit PDE gamma. We have isolated and characterized the complex T alpha.GTP gamma S-PDE gamma formed when T alpha is activated by the nonhydrolyzable analog GTP gamma S. Sedimentation and light-scattering techniques demonstrate that, in contrast to free T alpha.GTP gamma S, which is soluble, the T alpha.GTP gamma S-PDE gamma complex, as well as T alpha.GTP-PDE gamma, is membrane bound at cytosolic ionic strength. It is eluted from the membrane at low ionic strength as a monomeric and 1:1 stoichiometric complex. The relative affinities of PDE gamma for PDE alpha beta and for T alpha.GTP are discussed.
3',5'-Cyclic-GMP Phosphodiesterases↗
Positional specificity of lipoxygenases and their suitability for testing potential drugs.
Comparative studies on isolated lipoxygenases revealed the following features: variability of their positional specificity, diversity of enzymatic properties including sensitivity to inhibitors, a common general mechanism of action. The molecular basis of the positional specificity of lipoxygenases is discussed. The lipoxygenase of reticulocytes proved to be an excellent model in the search for inhibitors of the arachidonate 5-lipoxygenase. Reticulocyte lipoxygenase may form a wide spectrum of products belonging to all important classes of primary lipoxygenase metabolites hitherto known (HPETE's, diHPETE's, lipoxins, epoxyleukotrienes and others). This enzyme was used as basal test in a screening hierarchy for lipoxygenase-inhibitory antiasthmatic drugs and led to the discovery of new orally acting potential antiasthmatics. There was a high correlation between inhibition of reticulocyte lipoxygenase and antibronchoconstrictory activity with airway preparations in vitro and in vivo.
Animals↗
Phosphodiesterase activation by photoexcited rhodopsin is quenched when rhodopsin is phosphorylated and binds the intrinsic 48-kDa protein of rod outer segments.
Each photoexcited rhodopsin (R*) molecule catalyzes binding of GTP to many copies of the guanine nucleotide-binding protein transducin, which, in its GTP-binding form, then activates cGMP phosphodiesterase (PDEase). Subsequent deactivation of this light-activated enzyme cascade involves hydrolysis of the GTP bound to transducin, as well as decay of the activating capacity of R*. We report here that deactivation of PDEase in rod outer segment suspensions is highly enhanced by addition of ATP and purified 48-kDa protein, which is an intrinsic rod outer segment protein that is soluble in the dark but binds to photolyzed rhodopsin that has been phosphorylated by rhodopsin kinase and ATP [Kühn, H., Hall, S.W. & Wilden, U. (1984) FEBS Lett. 176, 473-478]. To analyze the mechanism by which ATP and 48-kDa protein deactivate PDEase, we used an ATP-free system consisting of thoroughly washed disk membranes, whose rhodopsin had been previously phosphorylated and chromophore-regenerated, and to which purified PDEase and transducin were reassociated. Such phosphorylated membranes exhibited a significantly lower (by a factor less than or equal to 5) light-induced PDEase-activating capacity than unphosphorylated controls. Addition of purified 48-kDa protein to phosphorylated membranes further suppressed their PDEase-activating capacity; suppression could be as high as 98% (as compared to unphosphorylated membranes), depending on the amount of 48-kDa protein and the flash intensity. Addition of ATP had little further effect. In contrast, PDEase activation or deactivation with unphosphorylated control membranes was not influenced by 48-kDa protein, even in the presence of ATP, provided rhodopsin kinase was absent. Our data suggest that 48-kDa protein binds to phosphorylated R* and thereby quenches its capacity to activate transducin and PDEase.
3',5'-Cyclic-GMP Phosphodiesterases↗
The occurrence of a lipoxygenase pathway in reticulocytes of various species.
Reticulocytes from various species (rat, mouse, rhesus monkey) obtained by phenylhydrazine treatment of the animals metabolized polyenoic fatty acids via a lipoxygenase pathway. Linoleic acid was converted to 13-hydro(pero)xy-9,11(Z,E)octadecadienoic acid [13-H(P)ODE] and 9-hydro(pero)xy-10,12(E,Z)octadecadienoic acid [9-H(P)ODE], whereas arachidonic acid was oxygenated to 15-hydroxy-5,8,11,13(Z,Z,Z,E)eicosatetraenoic acid (15-HETE) as shown by straight-phase high-pressure liquid chromatography (SP-HPLC). Addition of calcium and ionophore A 23,187 strongly enhanced the formation of lipoxygenase products, whereas 5,8,11,14eicosatetraenoic acid (ETYA) completely inhibited their formation. Estimates of the specific radioactivities of the lipoxygenase products indicate differences in the metabolization of externally added and endogenously released polyenoic fatty acids. These results strongly suggest that lipoxygenases generally occur in immature red blood cells.
5,8,11,14-Eicosatetraynoic Acid↗
Pure enzymes in a test hierarchy for antibronchoconstrictory lipoxygenase inhibitors.
A test hierarchy for potential antibronchoconstrictive drugs acting as lipoxygenase inhibitors was developed which includes a purified mammalian lipoxygenase/leukotriene A4 synthase, purified cyclooxygenase, arachidonic acid metabolism of polymorphonuclear leukocytes, arachidonic acid-induced contraction of lung strips, isolated human bronchi, and ovalbumin-induced bronchoconstriction with sensitized guinea pigs. The lipoxygenase from rabbit reticulocytes turned out to be superior to a variety of other animal and plant lipoxygenases. Among the various test systems with respiratory tract preparations, the arachidonic acid-induced contraction of guinea pig lung strips gave the most reliable results in the search for new antiasthmatic drugs. The validity of the test hierarchy was ascertained in screening investigations. Novel classes of antibronchoconstrictory lipoxygenase inhibitors were found.
Animals↗
Metabolism of polyunsaturated fatty acids by rabbit reticulocytes.
Rabbit reticulocytes obtained by repeated bleeding metabolize exogenous [1-14C]linoleic acid and [1-14C]arachidonic acid by three different pathways. 1. Incorporation into cellular lipids: 50% of the fatty acids metabolized are incorporated into phospholipids, mainly phosphatidylcholine (32.8%) but also into phosphatidylethanolamine (12%), whereas about 10% of the radioactivity was found in the neutral lipids (mono- di- and triacylglycerols, but not cholesterol esters). 2. Formation of lipoxygenase products: 30% of the fatty acids metabolized are converted via the lipoxygenase pathway mainly to hydroxy fatty acids. Their formation is strongly inhibited by lipoxygenase inhibitors such as 5,8,11,14-eicosatetraynoic acid or nordihydroguaiaretic acid. Inhibition of the lipoxygenase pathway results in an increase of the incorporation of the fatty acids into cellular lipids. 15-Hydroxy-5,8,11,13(Z,Z,Z,E)eicosatetraenoic acid and 13-hydroxy-9,11(Z,E)-octadecadienoic acid are incorporated by reticulocytes into cellular lipids and also are metabolized via beta-oxidation. The metabolism of arachidonic acid and linoleic acid is very similar except for a higher incorporation of linoleic acid into neutral lipids. 3. beta-Oxidation of the exogenous fatty acids: about 10% of the polyenoic fatty acids are metabolized via beta-oxidation to 14CO2. Addition of 5,8,11,14-eicosatetraynoic acid strongly increased the 14CO2 formation from the polyenoic fatty acids whereas antimycin A completely abolished beta-oxidation. Erythrocytes show very little incorporation of unsaturated fatty acids into phospholipids and neutral lipids. Without addition of calcium and ionophore A23187 lipoxygenase metabolites could not be detected.
Animals↗
Retinal S antigen identified as the 48K protein regulating light-dependent phosphodiesterase in rods.
Retinal S antigen chromatographically purified from whole retina, induces experimental autoimmune uveoretinitis in laboratory animals. The 48K protein, a soluble protein found in rod outer segments, is purified through its specific binding to photoexcited rhodopsin and is involved in the quenching of light-induced guanosine 3',5'-monophosphate-phosphodiesterase activity. Biochemical, immunological, functional, and pathological tests showed that retinal S antigen and the 48K protein are identical.
3',5'-Cyclic-GMP Phosphodiesterases↗
Can an osteoblastoma become malignant?
A rare case of an osteoblastoma in the clavicle is reported, that became malignant within 3 1/2 years, after two recurrences. The clinical course of the disease and a comparison of tissue sections removed during a total of four operations point to a secondary malignant transformation of an osteoblastoma, such as has been described by Schajowicz, among others. Six years after the first diagnosis was reached and 3 1/2 years after the last resection of the clavicle and the removal of a tumour from definitely healthy surrounding tissue, and after long-term chemotherapy, the now 17-year-old youth is free of metastases and there is no sign of a recurrence of the tumour. The problem of differentiating this kind of tumour from other types is discussed in light of international literature on the subject, and therapeutic recommendations are made.
Bone Neoplasms↗
Metabolism of [1-14C]-arachidonic acid by cultured calf aortic endothelial cells: evidence for the presence of a lipoxygenase pathway.
Intact cultured calf aortic endothelial cells from a 10th-14th subculture rapidly metabolize exogenous [1-14C]-arachidonic acid by three different routes: i) incorporation into triglycerides and phosphilipids in a ratio of about 2:1, ii) formation of lipoxygenase metabolites (12-hydroxy-5,8,10,14-eicosatetraenoic acid) and iii) formation of cyclooxygenase metabolites (6-keto-PG F1 alpha and PG F2 alpha). From analyses by thin-layer chromatography and high-pressure liquid chromatography it was established that the main lipoxygenase metabolites in intact cells are 12-hydroxy-5,8,10,14-eicosatetraenoic acid and a compound proposed to be (a) dihydroxyeicosatetraenoic acid(s). In frozen and thawed cells the incorporation of arachidonic acid into cellular lipids is abolished, whereas the lipoxygenase pathway is strongly enhanced. Under these conditions the cells produce predominantly 15-hydroxy-5,8,11,13-eicosatetraenoic acid in addition to 12-hydroxy-5,8,10,14-eicosatetraenoic acid. The formation of lipoxygenase products was inhibited by heating the cells or by preincubation with nordihydroguaiaretic acid or BW 755 degrees C, whereas indomethacin was without effect. The formation of 12-hydroxy-5,8,10,14-eicosatetraenoic acid by intact cells was inhibited by 5,8,11-eicosatriynoic acid. Indomethacin and acetylsalicylic acid inhibited the formation of cyclooxygenase metabolites. 15Ls-hydroxy-5,8,11,13-eicosatetraenoic acid was incorporated into cellular lipids, but not dihydroxyeicosatetraenoic acids. Exogenous [3H]-labelled prostacyclin and TxB2 were not incorporated but were metabolized to less polar products.
Animals↗
On the nature of the stimulation of the lipoxygenase from rabbit reticulocytes by biological membranes.
The oxygenation of concentrated emulsions (about 260 microM) of arachidonic acid or linoleic acid catalyzed by the purified lipoxygenase from rabbit reticulocytes is strongly stimulated in the presence of low concentrations of beef heart submitochondrial particles or other membrane preparations. Maximal stimulation was observed at a proportion of about 5 mumoles polyenoic acid per mg of membrane protein. Whereas at a constant ratio of arachidonic acid and membranes the reaction rate obeyed the Michaelis-Menten kinetics with an apparent Km value of 75 microM for arachidonic acid, sigmoid kinetics was observed under conditions of constant concentration of membranes and, consequently, varying proportions of arachidonate and membranes. Under conditions of maximal stimulation binding of reticulocyte lipoxygenase to the membranes was insignificant. Moreover, one-third of the arachidonic acid sedimented during ultracentrifugation of the membranes as judged from experiments with [14C] arachidonic acid. From other sedimentation experiments and from the comparison with soybean lipoxygenase which is not stimulated by membranes it is concluded that the emulsion droplets interacting with the membranes are the substrate of the stimulated reaction. The stimulation may be brought about by facilitating the susceptibility of the fatty acids to reticulocyte lipoxygenase. The membrane-stimulated reaction of reticulocyte lipoxygenase was not inhibited by the antioxidant 2,6-di-t-butyl-4-hydroxytoluene (BHT) excluding the participation of free radicals. The stimulatory effect of membranes seems not to be related to their susceptibility towards lipoxygenase attack which varied strongly in order mitochondrialmembranes greater than endoplasmic membranes greater than erythrocyte ghosts. The stimulation by membranes resembles that produced by the detergent sodium cholate at concentrations near to the critical micellar concentration. Moreover, the stimulations by membranes and by cholate do not behave synergistically or abolish each other. The stimulatory effect of membranes did not occur at a proportion of 200 nmoles per mg of membrane protein.
Animals↗
[A clinical case of an unusual metastasis of cervical carcinoma].
Explore the source record for details and available documents.
Aged↗
The formation of products containing a conjugated tetraenoic system by pure reticulocyte lipoxygenase.
The pure lipoxygenase from reticulocytes converts 5,15-di-HETE at 2 degrees C to product(s) showing a characteristic UV spectrum with maxima of strong absorbance at 300 and 316 nm and shoulders at 285 and 350 nm. Their formation was completely prevented by the lipoxygenase inhibitors, ETYA and NDGA, by heating of the enzyme and by anaerobiosis. At 35 degrees C the products were formed only initially and in small amounts. The reaction products were purified by SP-HPLC and shown to migrate in the region of tri-HETEs in thin-layer chromatograms. The lipoxygenase from soybeans also converts 5,15-di-HETE to these product(s) with a comparable initial rate but different kinetics. These data suggest that 5,15-di-HETE is converted via a lipoxygenase reaction to 5,6,15-trihydroxy-7,9,11,13-(e,e,c,e)- and/or 5,14,15-trihydroxy-6,8,10,12-(e,c,e,e)-eicosatetraenoic acid, both of which contain a conjugated tetraene system.
Anaerobiosis↗
Light-induced binding of 48-kDa protein to photoreceptor membranes is highly enhanced by phosphorylation of rhodopsin.
The 48-kDa protein, a major protein of rod photoreceptor cells, is soluble in the dark but associates with the disk membranes when some (5-10%) of their rhodopsin has absorbed light and if this rhodopsin is additionally phosphorylated by ATP and rhodopsin kinase. If rhodopsin has been phosphorylated and regenerated prior to the protein binding experiment, the binding of 48-kDa protein depends on light but no longer on the presence of ATP. Another photoreceptor protein, GTP-binding protein, associates with both phosphorylated and unphosphorylated rhodopsin upon illumination. Excess GTP-binding protein thereby displaces 48-kDa protein from phosphorylated disks; this indicates competition between these two proteins for binding sites on illuminated phosphorylated rhodopsin molecules.
Adenosine Triphosphate↗