Eicosanoids in breast cancer patients before and after mastectomy.

In 19 patients with a malignant breast tumor, tumor tissue and blood were taken to determine the eicosanoid profile and platelet aggregation. Values were compared with those of patients with benign tumors (n = 4), or undergoing a mammary reduction (n = 7). Postoperatively, blood was taken as well in order to compare pre- and postoperative values. Eicosanoids were measured in peripheral blood monocytes and mammary tissue by means of HPLC; furthermore, TXA2, 6-keto-PGF1 alpha, and PGE2 were determined by RIA. Differences in pre- and postoperative values of cancer patients were seen in plasma RIA values: PGE2 and 6-k-PGF1 alpha were significantly higher preoperatively when compared with postoperatively, however, such differences were seen in the control groups as well. Compared to benign tumor or mammary reduction test material the eicosanoid profile of tissue obtained from malignant mammary tumors showed important differences. Except for PGF2 alpha, HHT and 15-HETE no detectable quantities of eicosanoids were found in the non-tumor material, whereas in the malignant tumor material substantial quantities of a number of eicosanoid metabolites were present. Statistically significant correlations could be established between patient/histopathology data and the results of the platelet aggregation assays, e.g. between menopausal status and ADP aggregation; oestrogen receptor (+/-) and collagen and arachidonic acid aggregation, inflammatory cell infiltration score and arachidonic acid aggregation and fibrosis score and ADP aggregation. The results show that eicosanoid synthesis in material from mammary cancer patients is different from that in benign mammary tissue. The implications, in particular, in relation to future prognosis of the patient, remain obscure.     

Localization and quantification of extracellular polymers in methanogenic granular sludge

Summary Extracellular polymers were localized and quantitatively analysed in methanogenic granular sludge cultivated on either propionate or ethanol in laboratory upflow anaerobic sludge-blanket (UASB) reactors. Electron microscopical analysis of ultrathin sections of the two sludge types stained with ruthenium red revealed the presence of extracellular polymers with different densities and structures. For quantification, granular sludge from a large-scale UASB reactor at a liquid sugar plant was also included in this study. A three-step physical disintegration procedure was used to extract water-soluble extracellular material from the granules. After each disintegration step the extracts were analysed for polysaccharides and proteins. Cell damage and thus the contribution of intracellular proteins and polysaccharides was estimated simultaneously by the determination of free DNA and free ATP in the extracts. After two extraction steps, up to 3.5 mg polysaccharides/g organic material and 5.5 mg protein/g organic material were extracted, whereas no significant increase in DNA was detected. The role of extracellular polymers in granular stability is discussed. Offsprint requests to: A. J. B. Zehnder     

Sex pheromone perception in male pine sawflies,Neodiprion sertifer (Hymenoptera; Diprionidae)

Summary Electroantennographic and single sensillum recordings were performed on male pine sawfly, Neodiprion sertifer, antennae. Responses to the sex pheromone component (2S, 3S, 7S)- 3,7-dimethyl-2-pentadecenyl (diprionyl) acetate (SSS:OAc), to the behavioral inhibitor (2S, 3R, 7R)-diprionyl acetate (SRR:OAc), to the six other enantiomers of diprionyl acetate, and to the biosynthetic precursor diprionol were recorded. Responses to trans-perillenal, a monoterpene identified in female gland extracts and to (2S, 3S, 7S)-diprionyl propionate (SSS:OPr), a field attractant for N. sertifer and some related sawfly species were also recorded.EAG recordings demonstrated a high antennal sensitivity to SSS:OAc and to SSS:OPr. A somewhat lower response was elicited by SRR:OAc.Single sensillum recordings revealed 8–12 different cells firing in each sensillum, corresponding to the number of cells observed in earlier morphological investigations. Out of these cells all, except one, responded to SSS:OAc and to SSS:OPr. No differences in the response to the two components could be observed. The largest amplitude cell in each sensillum was specifically tuned to the behavioral antagonist, SRR:OAc. The pheromone perception system encountered in male pine sawflies thus differs clearly from that observed in moths.Abbreviation EAG electroantennogram - OAc acetate - OPr propionate     

Interconversion of F430 derivatives of methanogenic bacteria

F₄₃₀ is the prosthetic group of the methylcoenzyme M reductase of methanogenic bacteria. The compound isolated from Methanosarcina barkeri appears to be identical to the one obtained from the only distinctly related Methanobacterium thermoautotrophicum. F₄₃₀ is thermolabile and in the presence of acetonitrile or C10 _in4 ^sup- two epimerization products are obtained upon heating; in the absence of these compounds F₄₃₀ is oxidized to 12, 13-didehydro-F₄₃₀. The latter is stereoselectively reduced under H₂ atmosphere to F₄₃₀ by cell-free extracts of M. barkeri or M. thermoautotrophicum. H₂ may be replaced by the reduced methanogenic electron carrier coenzyme F₄₂₀.Abbreviations CH₃S-CoM methylcoenzyme M, 2-methylthioethanesulfonic acid - HS-CoM coenzyme M, 2-mercaptoethanesulfonic acid - F₄₃₀ Ni(II) tetrahydro-(12, 13)-corphin with a uroporphinoid (III) ligand skeleton - 13-epi-F₄₃₀ and 12,13-di-epi-F₄₃₀ the 12, 13- and 12, 13-derivatives of F₄₃₀ - 12, 13-didehydro-F₄₃₀ F₄₃₀ oxidized at C-12 and C-13 - coenzyme F₄₂₀ 7,8-didemethyl-8-hydroxy-5-deazaflavin derivative - coenzyme F₄₂₀H₂ reduced coenzyme F₄₂₀ - MV⁺ methylviologen semiquinone - HPLC high-performance liquid chromatography     

Book reviews

Book reviewed in this article: Ecological theory and integrated pest management in practice. M. Kogan, editor World crop pests , Vol. 2. Aphids; their biology, natural enemies and control, PART A (Eds. A. K. Minks and P. Harrewijn)     

Dopamine beta-hydroxylase from bovine adrenal medulla contains covalently-bound pyrroloquinoline quinone

Treatment of homogeneous dopamine beta-hydroxylase (DBH) preparations from bovine adrenals with the inhibitor phenylhydrazine (PH) changed the structureless absorption spectrum of DBH into spectra with a maximum at 350 nm. A product with this absorption spectrum could be detached with pronase, enabling its isolation. It appeared to be the C(5) hydrazone of pyrroloquinoline quinone (PQQ) and PH, as judged from its properties and the fact that it could be transformed into PQQ itself. From the yield obtained a ratio of 0.85 PQQ per enzyme subunit was calculated. In contrast to copper-quinoprotein amine oxidases (EC 1.4.3.6), hydrazone formation in DBH did not require saturation of the mixture with O2. DBH is the first copper-quinoprotein hydroxylase found so far. The implications of this finding for the current views on mechanism of action and inhibition by hydrazines are discussed. The success of the recently developed 'hydrazine method' [(1987) FEBS Lett. 221, 299-304] for all different types of amine oxidoreductases, suggest that the method could also be applied to other enzymes for which hydrazines are inhibitors and where the identity of the cofactors has not been established or the presence of PQQ is suspected.     

L-tyrosine is the precursor of PQQ biosynthesis in Hyphomicrobium X

A method was developed to study amino acids as possible precursors of PQQ biosynthesis. Cultures of Hyphomicrobium X, growing on [13C]methanol, were supplemented with unlabelled amino acids. Uptake and participation in metabolism were determined via gas chromatography/mass spectrometry of derivatized amino acids, obtained from hydrolyzed cellular protein, by measuring their 12C content. Several amino acids appeared to be incorporated into the protein to a significant extent, without degradation or conversion. Among these were the aromatic amino acids, L-tyrosine and L-phenylalanine. Using the same replacement approach, their incorporation into PQQ was determined by 1H- and 13C-NMR spectroscopy of purified PQQ obtained from the culture medium. It appeared that the complete carbon skeleton of tyrosine was present, forming the o-quinone and pyrrole-2-carboxylic acid moieties in PQQ, while phenylalanine was not incorporated at all. Starting with L-tyrosine, possible biosynthetic routes to PQQ are discussed.     

Activation by reduction of the resting form of cytochrome c oxidase: tests of different models and evidence for the involvement of CuB

(1) The reaction of the resting form of oxidised cytochrome c oxidase from ox heart with dithionite has been studied in the presence and absence of cyanide. In both cases, cytochrome a reduction in 0.1 M phosphate (pH 7) occurs at a rate of 8.2.10(4) M-1.s-1. In the absence of cyanide, ferrocytochrome a3 appears at a rate (kobs) of 0.016 s-1. Ferricytochrome a3 maintains its 418 nm Soret maximum until reduced. The rate of a3 reduction is independent of dithionite concentration over a range 0.9 mM-131 mM. In the presence or cyanide, visible and EPR spectral changes indicate the formation of a ferric a3/cyanide complex occurs at the same rate as a3 reduction in the absence of cyanide. A g = 3.6 signal appears at the same time as the decay of a g = 6 signal. No EPR signals which could be attributed to copper in any significant amounts could be detected after dithionite addition, either in the presence or absence of cyanide. (2) Addition of dithionite to cytochrome oxidase at various times following induction of turnover with ascorbate/TMPD, results in a biphasic reduction of cytochrome a3 with an increasing proportion of the fast phase of reduction occurring after longer turnover times. At the same time, the predominant steady state species of ferri-cytochrome a3 shifts from high to low spin and the steady-state level of reduction of cytochrome a drops indicating a shift in population of the enzyme molecules to a species with fast turnover. In the final activated form, oxygen is not required for fast internal electron transfer to cytochrome a3. In addition, oxygen does not induce further electron uptake in samples of resting cytochrome oxidase reduced under anaerobic conditions in the presence of cyanide. Both findings are contrary to predictions of certain O-loop types of mechanism for proton translocation. (3) A measurement of electron entry into the resting form of cytochrome oxidase in the presence of cyanide, using TMPD or cytochrome c under anaerobic conditions, shows that three electrons per oxidase enter below a redox potential of around +200 mV. An initial fast entry of two electrons is followed by a slow (kobs approximately 0.02 s) entry of a third electron.(ABSTRACT TRUNCATED AT 400 WORDS)     

The pho-controlled outer membrane porin PhoE does not contain specific binding sites for phosphate or polyphosphates

Purified PhoE-porins were reconstituted into black lipid bilayer membranes, and the selectivity and size of the reconstituted pores were determined. Addition of polyphosphates influenced the internal charge situation of the pore resulting in a shift from anion to cation selectivity. However, the pore size as judged from single channel conductances was not influenced by the addition of polyphosphates. A strong inhibition of the pore conductance only occurred when Mg2+ was also present in the aqueous phase. The inhibition of the pore function is presumably caused by the formation of a chelate between the divalent cation and the polyphosphate. Nevertheless, neither this inhibition nor the selectivity shift are specific to phosphate, because both effects can be mimicked by other polyvalent anions such as citrate. Inhibition of the PhoE pore function by polyphosphate in in vivo experiments confirmed the results of in vitro experiments that polyphosphate is only able to affect the permeability of the outer membrane toward beta-lactam antibiotics if Mg2+ is present. The outcome of the in vivo and the in vitro experiments are consistent with the assumption that the PhoE-porins do not contain a specific binding site for phosphate or polyphosphates but are anion selective because of an excess of positively charged amino acids inside or at the surface of the pore.