Oxidative Damage to DNA and Deoxyribose by β-Lactam Antibiotics in the Presence of Iron and Copper Salts

β-lactam antibiotics in the presence of certain metal ions damage deoxyribose and DNA with the release of thiobarbituric acid-reactive material. This damage can be substantially prevented by catalase, metal chelators and some scavengers of the hydroxyl radical. Ferric salts in the presence of certain β-lactam antibiotics were effective in degrading deoxyribose but they did not appear to damage DNA. In contrast copper salts and p-lactam antibiotics were extremely effective in damaging both DNA and deoxyribose.     

Purification of the Lewis blood-group gene associated α-3/4-fucosyltransferase from human milk: an enzyme transferring fucose primarily to Type 1 and lactose-based oligosaccharide chains

A soluble Lewis blood-group gene associated -3/4-L-fucosyltransferase has been purified from human milk by a series of steps involving hydrophobic chromatography on Phenyl Sepharose 4B, ion exchange chromatography on CM-Sephadex C-50, affinity chromatography on GDP-hexanolamine Sepharose 4B and gel filtration on Sephacryl S-200. The first step separated -3-L-fucosyltransferase activity directed towardsN-acetylglucosamine in Type 2 (Gal1-4GlcNAc-R) acceptors from an -3/4-fucosyltransferase fraction acting on both Type 1 (Gal1-3GlcNAc-R) and Type 2 acceptors. Further purification of this latter fraction on CM-Sephadex and GDP-hexanolamine Sepharose gave a single peak of fucosyltransferase activity that catalysed the addition of fucose toN-acetylglucosamine in both Type 1 and Type 2 acceptors and to theO-3 position of glucose in lactose-based oligosaccharides. The enzyme preparation at this stage resembled previously described -3/4-fucosyltransferase preparations purified from human milk. However, gel filtration of this preparation on Sephacryl S-200 or Sephadex G-150 separated further amounts of -3-fucosyltransferase activity acting solely on Type 2 acceptors and left a residual -3/4-fucosyltransferase that retained strong -4 activity with the Type 1 acceptor, lacto-N-biose 1, and -3 activity with 2-fucosyllactose, but had relatively little -3 activity withN-acetyllactosamine and virtually no capacity to transfer fucose to glycoproteins withN-linked oligosaccharide chains having unsubstituted terminal Type 2 structures.     

Microtubule-associated protein 2 (MAP2)-immunoreactive neurons in the retina of Bufo marinus: colocalisation with tyrosine hydroxylase and serotonin in amacrine cells

Summary Neuron populations in the retina of the toad, Bufo marinus, were labelled with a monoclonal antibody raised against microtubule-associated protein 2 (MAP2). A subpopulation of cones, probably corresponding to the blue-sensitive small single cones, large diameter amacrine cells in the most proximal row of the inner nuclear layer and some large ganglion cells in the ganglion cell layer were labelled. Double labelling experiments were carried out to establish the colocalisation of MAP2 with known putative transmitter substances of the anuran amacrine cells. MAP2 was colocalised in a subpopulation of serotonin-immunoreactive and in all tyrosine hydroxylase-immunoreactive amacrine cells. The results indicate, that the MAP2 content in the neurons of the anuran retina can be correlated with other well-defined neurochemical and/or physiological properties.On leave from Department of Zoology, Attlia József University, Szeged, Hungary     

Target organ-specific inactivation of drug metabolizing enzymes in kidney of hamsters treated with estradiol

Chronic treatment of hamsters with estradiol for several months has previously been shown to decrease the specific content of cytochrome P450 in the kidney, a target of hormonal carcinogenesis, but not in liver. The reason for this decrease in metabolic enzyme activity is unknown and has been examined in this investigation. We now report that the decrease in specific content of renal cytochrome P450 by 73% in response to estradiol was not affected by co-treatment with tamoxifen for 1 month. The subcutaneous infusion of 250 μg/day estradiol for 7 days lowered renal cytochrome P450 by 71% from control values and was therefore used for further mechanistic studies. This treatment decreased renal activities of estradiol 2- or 4-hydroxylase by 77 to 80%, of 7-ethoxycoumarin-O-deethylase by 66% of control values, respectively, and completely eliminated aryl hydrocarbon hydroxylase activities, whereas liver enzymes remained unaffected. After 7 days of infusion of estradiol, fluorescent products of lipid peroxidation were more than doubled in hamster kidney but remained unchanged in liver. The possibility of enzyme destruction by binding of estradiol 2,3-quinone to metabolizing enzymes was investigatedin vitro. In the presence of 2-hydroxyestradiol, cumene hydroperoxide, and microsomes, conditions known to favor the oxidation of the steroid to quinone, the binding of catechol estrogen metabolite to microsomal protein increased 60 fold over control values in the absence of cofactor. Purified rat liver cytochrome P450c also oxidized 2-hydroxyestradiol to 2,3-estradiol quinone. The rate of oxidation was linear for the first 2–3 min, but thereafter decreased with time. Under these incubation conditions, irreversible binding of catechol estrogen metabolite to cytochrome P450c increased for the first 2–3 min and then remained at this plateau level. It was concluded that enzyme destruction by a reactive estrogen metabolite or by lipid peroxides may be a major reason for the organ-specific decrease in cytochrome P450 enzymes in kidneys of estrogen-treated hamsters.     

The “ON”-bipolar agonist,L-2-amino-4-phosphonobutyrate,blocks light-evoked cone contraction in xenopus eye cups

Rhythmic photoreceptor metabolism in relationship to light-dark cycles is now thought be regulated through a retinal feed-back mechanism with dopamine serving as a principal signal initiating light-evoked events. In order to test the hypothesis that depolarizing ON-bipolar neurons participate in the retinal signalling pathway, we determined the effects of L-2-amino-4-phosphonobutyrate (L-APB) on light-evoked cone contraction in eye cups fromXenopus laevis. L-APB blocked the response stereospecifically when applied over a broad concentration range. The high specificity of L-APB in retina suggests that sign-inverting bipolar neurons which depolarize in light are in the signalling pathway. One possibility is that this pathway conveys signals that regulate dopamine release.Special issue dedicated to Dr. Frederick E. Samson.     

Initial reactions in the mineralization of 2-sulfobenzoate by Pseudomonas sp. RW611

Abstract Pseudomonas sp. strain RW611 utilized the ammonium salt of 2-sulfobenzoate as sole source of carbon, sulfur, nitrogen, and energy. The xenobiotic sulfo substituent was dioxygenolytically eliminated as sulfite, which was then slowly oxidized to sulfate. 2,3-Dihydroxybenzoate, which resulted from desulfonation underwent meta -cleavage, mediated by 2,3-dihydroxybenzoate 3,4-dioxygenase activity. This enzyme was inhibited by 3-chlorocatechol and 2,3,4-trihydroxybenzoate.     

Infradian cycles of oxygen consumption in diapausing pupae of the flesh fly, Sarcophaga crassipalpis, monitored by a scanning microrespirographic method.

Fine details of the infradian O2 consumption cycles that characterize pupal diapause in flesh flies have been monitored by a newly designed microrespirographic method coupled with an electronically regulated O2 generator. During the 4-5 days between the peaks of elevated O2 consumption, the diapausing pupae maintained a very low and fairly constant respiratory rate (13 microl O2 x g-1.h-1). During the intercalated peaks of increased respiratory metabolism, which lasted an average of 33.6 h to 24-27 degrees C, the average maximum rate of O2 consumption was 86.9 microl.g-1.h-1, a value of 6.7 times higher than the interpeak values. The respiratory peaks started abruptly in some cases while the decline was consistently gradual. During the periods between the peaks there were no discontinuous bursts of CO2 release, a feature common to diapause in many other insects. Diapause was characteristically terminated during a peak of the O2 consumption cycle. At diapause termination O2 consumption remained at the maximum values of the peak for many hours and then gradually increased to levels characteristic of nondiapause development.     

ATP-dependent Ca2+ transport and Ca2+-ATPase activities in an enriched plasma membrane fraction from gypsy moth larval midgut tissue

A subcellular fraction enriched in plasma membranes was obtained from gypsy moth (Lymantria dispar) larval midgut tissue. Using [⁴⁵Ca]²⁺ as a tracer, Ca²⁺ transport activity by membrane vesicles in the enriched fraction was measured and shown to be ATP-dependent, with a very high affinity for Ca²⁺ (apparent K_m for [Ca²⁺ _free]

1 Abbreviations used: [Ca²⁺free] = concentration of free (unbound) calcium ion;CaM = calmodulin; F = fraction; IOV = inside-out membrane vesicles; W-5 = N-(6-aminohexyl)-1-naphthalenesulfonamide; W-7 = N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide.

= 22 nM). Ca²⁺ transport was abolished upon addition of the calcium ionophore, A23187. Ca²⁺-stimulated, Mg²⁺-dependent ATPase activity peaked between 100 and 200 nM Ca²⁺_free. Ca²⁺-Mg²⁺-ATPase activity was inhibited by vanadate, 2 phenothiazine drugs (trifluoperazine and chlorpromazine), and the naphthalene sulfonamide, W-7; the related compound, W-5, and ouabain had a negligible effect. These results suggest the presence of a high affinity plasma membrane Ca²⁺ pump in gypsy moth larval midgut cells and are discussed in light of earlier work involving calcium transport in isolated midguts of larval Hyalophora cecropia. Ionic and other conditions that characterize the midgut physiology of larval Lepidoptera (e.g., luminal pH; electrochemical gradient for Ca²⁺; effect of certain ions and inhibitors on Ca²⁺ transport) contrast significantly with those found in adult Diptera. The implications that these differences may have for calcium regulation are discussed. © 1992 Wiley-Liss, Inc.