Hexaheme nitrite reductase from Desulfovibrio desulfuricans. M?ssbauer and EPR characterization of the heme groups

M?ssbauer and EPR spectroscopy were used to characterize the heme prosthetic groups of the nitrite reductase isolated from Desulfovibrio desulfuricans (ATCC 27774), which is a membrane-bound multiheme cytochrome capable of catalyzing the 6-electron reduction of nitrite to ammonia. At pH 7.6, the as-isolated enzyme exhibited a complex EPR spectrum consisting of a low-spin ferric heme signal at g = 2.96, 2.28, and 1.50 plus several broad resonances indicative of spin-spin interactions among the heme groups. EPR redox titration studies revealed yet another low-spin ferric heme signal at g = 3.2 and 2.14 (the third g value was undetected) and the presence of a high-spin ferric heme. M?ssbauer measurements demonstrated further that this enzyme contained six distinct heme groups: one high-spin (S = 5/2) and five low-spin (S = 1/2) ferric hemes. Characteristic hyperfine parameters for all six hemes were obtained through a detailed analysis of the M?ssbauer spectra. D. desulfuricans nitrite reductase can be reduced by chemical reductants, such as dithionite or reduced methyl viologen, or by hydrogenase under hydrogen atmosphere. Addition of nitrite to the fully reduced enzyme reoxidized all five low-spin hemes to their ferric states. The high-spin heme, however, was found to complex NO, suggesting that the high-spin heme could be the substrate binding site and that NO could be an intermediate present in an enzyme-bound form.     

E. coli membranes become permeable to ions following T7-virus-infection

Summary Infection of E. coli with the viruses T7 or T3 leads to a dramatic efflux of potassium ions. This ion efflux is caused by the virus particle since no concomitant protein synthesis is required. T7 mutants carrying deletions in the M-gene (Schweiger et al., 1975), however, yield virus particles disturbed in the ion release.     

Multiplication of Coxsackie B1 Virus in Synchronized HeLa Cells

A higher yield of Coxsackie B(1) virus was obtained when HeLa cells were infected late during S phase as compared to the amount produced by random cultures.     

Chelating derivatives of chitosan obtained by reaction with ascorbic acid

Ascorbic acid immediately dissolves Euphausia superba chitosan upon mixing and forms chitosan ascorbate; during the 6-h period after dissolution in water at pH 5–7, ascorbate is oxidized to dehydroascorbate which undergoes Schiff reaction with the amino groups of chitosan, thus yielding a viscous solution of a polymeric ketimine. The latter is characterized by infrared spectrometry, circular dichroism spectropolarimetry, viscometry and alkalimetry. When brought into contact with transition metal ions, the chitosan ascorbate ketimine yields insoluble metal chelates. Upon reduction with sodium cyanoborohydride, the water-insoluble N-[2-(1,2-dihydroxyethyl)tetrahydrofuryl] chitosan (NDTC) is obtained, which shows enhanced capacity for uranium, up to 800 mg U/g from solutions at pH 4·5.     

Phenyl-substituted aminomethylene-bisphosphonates inhibit human P5C reductase and show antiproliferative activity against proline-hyperproducing tumour cells

In certain cancers, such as breast, prostate and some lung and skin cancers, the gene for the enzyme catalysing the second and last step in proline synthesis, δ¹-pyrroline-5-carboxylate (P5C) reductase, has been found upregulated. This leads to a higher proline content that exacerbates the effects of the so-called proline-P5C cycle, with tumour cells effectively using this method to increase cell survival. If a method of reducing or inhibiting P5C reductase could be discovered, it would provide new means of treating cancer. To address this point, the effect of some phenyl-substituted derivatives of aminomethylene-bisphosphonic acid, previously found to interfere with the catalytic activity of plant and bacterial P5C reductases, was evaluated in vitro on the human isoform 1 (PYCR1), expressed in E. coli and affinity purified. The 3.5-dibromophenyl- and 3.5-dichlorophenyl-derivatives showed a remarkable effectiveness, with IC₅₀ values lower than 1 µM and a mechanism of competitive type against both P5C and NADPH. The actual occurrence in vivo of enzyme inhibition was assessed on myelogenous erythroleukemic K562 and epithelial breast cancer MDA-MB-231 cell lines, whose growth was progressively impaired by concentrations of the dibromo derivative ranging from 10⁻⁶ to 10⁻⁴M. Interestingly, growth inhibition was not relieved by the exogenous supply of proline, suggesting that the effect relies on the interference with the proline-P5C cycle, and not on proline starvation.     

Validity of the reduced version of the Morningness-Eveningness Questionnaire

Sleep and Biological Rhythms - The aim of the present work was to validate the reduced version of the Morningness-Eveningness Questionnaire (MEQr) using circadian motor activity as external...     

Functional Anatomy of the Ovule in Broad Bean (Vicia faba L.): Ultrastructural Seed Development and Nutrient Pathways

Ovules of broad bean (Vicia faba L.) were studied to discloseultrastructural features, which can facilitate nutrient transportto the embryo sac from 10 d after pollination (DAP) to the matureseed. Fertilization occurs during the first 24 h after pollination.The endosperm is a coenocyte, which is eventually consumed bythe embryo. By 10 DAP the inner integument is degraded and theouter integument adjoins the embryo sac boundary. The heart-shapedembryo approaches the embryo sac boundary at two sites, whichhere are named contact zones. Small integument cells in theneighbourhood of the first formed contact zones become separatedby prominent intercellular spaces. A heterogenous scatteringmaterial, probably representing secretion products accumulatesin these spaces. By 14-16 DAP the integument exudate disappears,and the suspensor degenerates. As the contact zones increasein size, wall ingrowths form a bridging network in the narrowspace between the embryo sac boundary and the extra-embryonicpart of the endosperm wall. The epidermal cells of the embryoseparate adjacent to these zones, and develop conspicuous wallingrowths. At 20 DAP vacuoles showing various stages in formationof protein bodies appear in the cells of the embryo.Copyright1994, 1999 Academic Press Vicia faba, broad beans, ovule, seed, nutrient transport