Band 32, Heft 11-12

An NMR 13 C study has shown that proton rearrangement in pyrazole is decreased by adding small amounts of water to the sample. Single water molecules as coordination centres pertrub the mechanism of cooperative proton transfer occurring in pyrazole selfassociates. The decrease of the rearrangement effect due to water is similar to the action of transition metal ions studied by the authors earlier.

NMR methods have been applied to evaluation of prototropic tautomerism, N(1)H ⇌ N (3) H, in several selected pyrimidines, viz. the neutral forms of Isocytosine and 2-alkylthiopyrimidone-4, and the monoanionic forms of uracil, 5-fluorouracil and 4-thiouracil. The predominant tautomeric species of the neutral forms could be estimated only qualitatively from 1 H chemical shifts. For the monoanionic forms this procedure was not applicable, for reasons which are discussed in detail. For the monoanionic form of uracil, 13 C chemical shifts of C(5) provided a suitable criterion for quantitative estimation of the populations of the two known tautomeric species. However, the potential scope of this procedure appears somewhat limited. By contrast, the values of the vicinal proton-proton coupling constants, J (5,6), provided both necessary and adequate criteria for quantitative evaluation of the tautomer populations for all the neutral and monoanionic forms. The results were in satisfactory agreement with those obtained by optical spectroscopic methods. In some instances the results obtained in this way may be more reliable than those derived from optical methods. The range of applicability, and utility, of NMR methods to studies on protropic tautomerism in pyrimidines are critically assessed.

The intermolecular association in aqueous solution of the N 1 -oxide nucleotide analog of adenosine-5′-monophosphate has been investigated by 1 H-NMR spectroscopy over the concentration range 0.0006-0.6 ᴍ. The concentration profile of individual chemical shifts provides evidence for an unusual stacking pattern compared to that of the natural nucleotide AMP, involving only the imidazole moiety of the adenine base. The different stacking pattern of this nucleotide analog suggests that the exact stacking geometry of natural nucleotides interacting with nucleotide-depending enzymes, is achieved by adjusting to the geometry of the individual receptor molecule.

An improved method is described for the separation of 22 compounds normally related to purine and 6-thiopurine metabolism in biological materials using high-pressure liquid cation-exchange chromatography on strongly acidic exchange resin. The column (0.18 × 100 cm) is eluted with 0.4 ᴍ ammonium formate, pH 4.6, at a linear flow velocity of 5.2 cm · min-1 at 50 °C. The elution volumes of sulphate anions, allopurinol, 6-thioxanthine, adenine, adenosine, and guanosine are demonstrated additionally to further 16 purine and 6-thiopurine compounds.

The influence of five different methods of homogenisation (1. The method according to Potter and Elvehjem, 2. A modification of this method called Potter S, 3. The method of Dounce, 4. Homogenisation by hypersonic waves and 5. Coarce-grained homogenisation with the “Mikro-fleischwolf”) on the absolute value and stability of oxygen uptake of guinea pig liver homogenates has been investigated in simultaneous measurements. All homogenates showed a characteristic fall of oxygen uptake during measuring time (3 hours). The modified method according to Potter and Elvehjem called Potter S showed reproducible results without any influence by homogenisation intensity.

The vertical distribution of eight heavy metals (Hg, Cu, Zn, Cr, Co, Ni, Pb, Cd) in dated sediments of four sedimentary profiles from the central part of Lake Constance was examined. From 1900 to about 1960 - 1970, a general increase of all heavy metals was observed. After this time, the concentrations of most of the metals decreased. As compared with pre-1900 “natural” metal concentrations, Cd and Pb have been enriched the strongest (enrichment factors 4.0 and 3.8 resp.), whereas Ni and Co exhibit least civilizational influences (enrichment factors 1.5 and 1.9 resp.). Cu, Cr, Zn and Hg were enriched by a factor between 2 - 3. Elements which were not influenced by anthropogenic processes - such as K and Li - remained unaffected over the whole vertical sequence. Polycyclic aromatic hydrocarbons (PAH) which were also studied in the same sediment material by Grimmer and Böhnke show a similar pattern. Within a sedimentary profile the concentration ratio benzo (a) pyrene: cadmium remained about constant. Combustion of coal is believed to be the common source of both heavy metals and PAH and seems to be responsible for the general distribution pattern of both groups of pollutants. In addition, local sources - both domestic and industrial - are assumed to have delivered additional heavy metal pollution.

The vertical distribution of nitrogen and phosphorus in dated sediments of four sedimentary profiles from the central part of Lake Constance was examined. From about 1900 until now a steady increase of N and P is to be observed. Nitrogen is enriched by a factor of about 5, phosphorus by a factor of about only 2. The N/P-ratio shows a continuous increase since 1900. The explanation for this development is the steady increase of phosphates delivered into the lake since about 1900. Thus, p.ex. the total amount of phosphorus increased between 1930 and 1974 from 274 to 1929 tons per year. Most of the sedimentary nitrogen is incorporated into the organic material, the production of which is clearly bound to the availability of phosphorus in the lake water. Phosphorus is mostly found in an adsorbed position on clay minerals. Between N and P on one hand and certain heavy metals (Cd, Zn) and polycyclic aromatic hydrocarbons on the other hand a positive correlation only can be found as long as the curves for phosphate delivery (responsible for N-and P-concentrations) and coal consumption (responsible for certain heavy metals and polycyclic aromatic hydrocarbons) have a parallel development. The decrease in coal consumption in connection with a very strong increase of phosphate delivery during the past 15 years has ceased the parallel evolution of the different groups of pollutants in the sediments, and also demonstrates, that a causal relationship N and P - heavy metals and polycyclic aromatic hydrocarbons does not exist.

The synthesis of a potassium and rubidium specific ionophore by Streptococcus faecalis has been proved, using bilayer measurements. The characteristics of this substance agree with the ones, published for free mobile carriers, such as valinomycin and the macrotetrolides.

The smallest active form of aminopeptidase I (EC 3.4.11.1) from yeast has a molecular weight of 6.4 × 10 5 . At neutral pH the active enzyme is in equilibrium with two inactive subfragments (M r = 3.2 × 10 5 and 1.1 × 10 5 ) as well as with higher aggregates (Mr ≧ 1.2 × 10 6 ). All of these species may be dissociated to give a single type of subunits with a molecular weight of 5.3 × 10 4 . It is concluded that the active enzyme is a dodecamer whereas the subfragments correspond to dimeric and hexameric forms.

Aminopeptidase, Yeast, Electron Microscopy, Quaternary Structure Electron micrographs of native aminopeptidase I and of isolated subfragments were taken after negative staining with uranyl formate. From these studies and from the chemical evidence sum­ marized in the preceding paper it is concluded that the active enzyme is a dodecamer possessing pseudo-D 3 symmetry with the dimer as the smallest symmetric unit.

The inhibition of the proliferation of hyperdiploid Ehrlich ascites tumor cells in suspension cultures by amobarbital is coupled to an increased glycolytic activity as shown by lactic acid pro­ duction and glucose consumption; higher concentrations of amobarbital than 1 mᴍ enhance the ATP/ADP ratio of the total cell. The actual activity of pyruvate dehydrogenase of intact cells is completely inhibited in the presence of 2 mᴍ amobarbital as was shown by the 14 CO 2 evolution from [1- 14 C] pyruvate or the incorporation of 14 CO 2 into the total lipid fraction of the cells from [U- 14 C] pyruvate or from [U- 14 C] lactate. The pyruvate dehydrogenase complex from Ehrlich ascites tumor cells is completely inhibited by 1 mᴍ amobarbital in vitro. The activity of α-oxoglutarate dehydrogenase is inhibited by amobarbital, too, as was shown by measuring the 14 CO 2 evolution from [1- 14 C] glutamate with intact cells. It is suggested that the inhibition of pyruvate dehydrogenase in the presence of amobarbital is the result of a direct action on the enzyme as well as the consequence of a change in the cellular redox state or its energy charge.

A homogenate of cells of Streptomyces hydrogenans which have been cultivated in the presence of estradiol-17β, was separated by gel filtration on Sephadex G-200. In the presence of NADH, a few distinct fractions converted about 10% of [ 3 H] testosterone to 5α-dihydrotestosterone (5α-DHT), 5α-androstanedione, 4-androstenedione, and dehydroepiandrosterone (DHEA). The metabolites were separated after two consecutive runs on silica gel and propanediol-1,2-impregnated cellulose plates by thin layer chromatography. Identification was achieved by comparison with known steroid samples, specific staining procedures and by crystallization to constant specific radioactivity. The 5α-reductase activity responsible for the formation of 5α-DHT and 5α-androstanedione in vitro required NADH as co-substrate and could only be found after induction of the cells with estradiol-17β in vivo. Within the same chromatographic fractions, a Δ 4 -Δ 5 -ketosteroid isomerase activity can be detected which catalyzes the reverse reaction from the 4-androstene testosterone to the 5-anthrostene DHEA in vitro.

The assimilatory nitrate reductase was purified 60-fold from a newly isolated, nitrate assimilating strain of the photosynthetic bacterium Rhodopseudomonas capsulata. The enzyme had a molecular weight of about 180 000 dalton and was typically prokaryotic in that it was not active with reduced pyridine nucleotides but rather with reduced flavins.

The molar extinction coefficient for phycoerythrobilin (1a) was calculated by two independent methods. It is different from that of the cleaved chromophore, phycobiliviolin (2). By unfolding with urea or tryptic digestion, the chromophore absorption of C-phycoerythrin (PE) was determined free of any protein influence. The chromophore content of PE from various Cyanobacteria was determined with these data to be either 5 or 6, depending on the organism. This corresponds to a chromophore distribution over phycoerythrin subunits α:β of either 2:3 or 2:4. The phylogenetic significance of varying chromophore content is discussed.

Electrochromic Absorption Changes, Photosynthetic Unit A correlation between the amplitude of the electrochromic absorption changes measured at 480 nm and 520 nm and the size of the photosynthetic unit is found. Three tobacco mutants which differ with respect to their photosynthetic unit size exhibit decreased amplitudes of the electrochromic absorption changes which parallel the size of the photosynthetic units. Under these conditions the ATP-yield per flash is unaffected by the photosynthetic unit size. From this it is deduced that the correlation between the amplitude of the electrochromic absorption change and the photosynthetic unit size is due to the fact that the number of pigments undergoing an electro­ chromic bandshift is proportional to the total pigment content. As the influence on the electro­ chromic effect at 520 nm is too pronounced as to be simply explainable by the variation of the carotenoid content in the different mutants it is inferred that either structural effects leading to a modification of the carotenoid orientation in the mutants or an indirect influence of chlorophyll on the electrochromic bandshift which depend on the number of chlorophyll molecules are responsible.

Photoreduction of the artificial electron acceptor, cytochrome c, by isolated spinach chloroplasts in the absence of added catalyst has been found to be resistant to inhibition by 10 mᴍ salicyl­ aldoxime. This contrasts with the pronounced inhibition of the photoreduction of both 2,6-dichloro-phenolindophenol and ferricyanide over a range of electron flow rates, and indicates an inhibition on the acceptor side rather than on the donor side of photosystem 2 (PS2). All photoreductions were susceptible to inhibition by 3-(3′,4′-dichlorophenyl)-1, 1-dimethyl urea (DCMU). Salicyl­ aldoxime (1 -10 mᴍ) inhibited electron flow from PS2 to P700, and also altered the chlorophyll fluorescence induction of dark-adapted chloroplasts. Salicylaldoxime concentrations up to 5 mM did not change the initial fluorescence level, F 0 , or the initial slope, (dF/dt) 0 , but lowered the final steady state fluorescence level, F m , and the value of p 0 [p 0 = (F m - F 0 )/F m ] ; an effect similar to that seen on 1 mᴍ ferricyanide addition, thus indicating an induced oxidation of the fluorescence quencher, Q. Both DCMU and orthophenanthroline, which block electron transport directly after Q, caused an increase in (dF/dt) 0 , F 0 and F m but left p 0 unchanged. This contrasted with the effect of salicylaldoxime. It is proposed that salicylaldoxime inhibits electron transport from PS2 to photosystem 1, at a site which can probably be identified with the plastoquinone pool, by inducing a cyclic flow of electrons around PS2. Cytochrome c, under the conditions used, appears to be photoreduced at a site close to the secondary electron acceptor, R, of PS2.

Protein synthesis after UV-and X-irradiation was investigated in diploid yeast. The incorporation of radioactively labelled lysine and phenylalanine was measured 2.5 and 4 hours after exposure. By varying the specific activity the pool sizes could be estimated. At 2.5 hours there is some increase in pool sizes and a dose-dependent enhancement of protein synthesis. At 4 hours pools are again normal, but the increase of synthetic activity prevails.

The effect of introducing alkyl substituents (methyl to pentyl) to the chain segment n in long-chain alkenyl acetates, CH 3 (CH 2 ) n CH = CH(CH 2 ) m -O-COCH 3 (I), was investigated in pheromone receptors of Noctuidae, Tortricidae, and Cochylidae species. The receptor types studied were maximally responsive to (Z)-7-or (Z)-9-dodecen-1-yl acetate, (Z)-7-, (Z)-9-, or (Z)-11-tetradecen-1-yl acetate, or (Z)-11-hexadecen-1-yl acetate, representing key compounds with n = 1, n = 3, or n = 5 methylene groups. In terms of the relative stimulus amounts required to elicit equivalent EAG amplitudes, the efficacy of the alkyl-branched derivatives was between 1/1000 to 300 times that of the unbranched chain of the same length and double bond position. The effects of branching were specific to the type of receptor, the length and double bond position of the parent chain, and the kind and position of the alkyl substituent. The most pronounced increase observed occurred with receptors for « = 1 type pheromones, (Z)-9-dodecen-1-yl acetate and (Z) -11-tetradecen-1-yl acetate when an α-methyl group was introduced to elongated derivatives; whereas the greatest decreasing effects were obtained on receptors for the n = 5 type compound, (Z)-7-tetradecen-1-yl acetate. The results show basic differences in structure-response relationships between the Noctuidae vs Tortricidae receptors studied. Various possible effects of the substituent groups during sensory transduction are considered. The data should contribute to further elucidation of interaction mechanisms of unbranched alkenyl acetate pheromones (I) with insect olfactory receptors.

Pyrophosphate Synthesis, Sarcoplasmic Reticulum, Calcium Accumulation The vesicles of the sarcoplasmic membranes synthesize pyrophosphate from inorganic phosphate. Pyrophosphate synthesis proceeds as long as a calcium gradient is maintained across the vesicular membranes. Pyrophosphate synthesis is inhibited by low concentrations of nucleoside triphosphates.

Calcification, Collagen Membrane, Ca/P Ratio Dependence Spontaneous calcification of a membrane made of native collagen has been investigated. The method permits independent variation of calcium and phosphate concentrations. With increasing phosphate concentration the precipitation of calcium-phosphate on the collogen occurs at a conspicuously lower calcium concentration as with a number of other membranes.

Whether proteins or nucleic acids were responsible for the emergence of life has been debated for a long time. Taking the observation that families of proteins display a remarkable invariance of their amino acid sequence around critical regions, such as active/binding sites, even though these proteins may represent considerable evolutionary diversity, as the naturally provided evidence of evolutionary selection of a working system, the idea is developed that: 1. Proteins had to have been first informational macromolecules that were necessary and sufficient to lead to the emergence of life; 2. it is impossible for a nucleic acid molecules to have formed, by chance, whose base sequence could yield a biologically active protein. A model is proposed to account for the emergence of the first sucessful cell according to this approach.

The fresh-water medusa Craspedacusta sowerbii (Hydrozoa: Limnomedusae) contains 99.26% water, 0.67% organic matter, and 0.07% inorganic materials. Its cation concentrations are compared to those of Aurelia aurita, sea-water, and habitat water.

Gold-back ferns and silver-back ferns of the genus Pityrogramma are known to produce chalcones and dihydrochalcones, deposited as a farina on the under surface of their fronds. In Cheilanthes and Notholaen a mostly methy­lated flavones and flavonols are excreted, but some species are shown here to accumulate chalcones and dihydrochalcones, too. Their distribution within the genera and within the species is reported and discussed.

Ten flavonol glycosides were isolated from leaves of Cedrus atlantica. Among the six aglycones the rarely occurring syringetin and laricitrin were found in addition to kaempferol, quercetin, isorhamnetin and myricetin. 3-Glyco-sylation with glucose and/or rutinose was the main pattern, whereas acylation with p-coumaric acid occurred for kaempferol-and isorhamnetin-3-glucoside.

Seven isoflavonoid phytoalexins have been isolated from the fungus inoculated hypocotyls of Lablab niger and identified as the isoflavone 2′-hydroxygenistein, the isoflavanones dalbergioidin and kievitone, the pterocarpan phaseollidin and the isofiavans demethyivestitol, isovestitol and iaxifloran. Small quantities of the 2-arylbenzofuran, vignafuran, were also detected.

Erythritol-1-d 1 has been isolated from γ-irradiated aqueous oxygenated solution of DNA after reduction with NaBD 4 , alkali and phosphatase treatment. It is concluded that this product stems from a ᴅ-erythrose 2,4-diphosphate unit in the DNA which is formed via a sequence of reactions following H-abstraction at C-2′ by OH radicals..

A high correlation coefficient r = -0.832 (P r≠0 <0.0001) was estimated in man for the inverse relationship between the diagnostic 59 Fe 2+ -absorption and the serum ferritin concentration which is very close to the correlation r = - 0.88 as described for the relationship between the diagnostic 59 Fe 2+ -absorption and the diffuse cytoplasmic storage iron in the bone marrow macrophages. The increase of the diagnostic 59 Fe 2+ -absorption seems to be an earlier and more sensitive indicator of depleted iron stores whereas the serum ferritin decreases somewhat later during the development of iron deficiency.

During persistent infection of mice with the lymphocytic choriomeningitis (LCM) virus approximately 3% of leukocytes were found to contain viral antigen and to produce infectious virus. Morphologically, infected cells were shown not to be lymphoblasts and their numbers were not reduced by removal of mononuclear phagocytes. We conclude that in LCM virus carrier mice true lymphocytes participate in the infectious process.

A simplified method is described for recording electrical potentials without injury from epidermal insect mechanoreceptors. For this purpose a stationary capillary-electrode is used with an anterior part made of polyethylene and filled with a hygroscopic electrolyte; its wide orifice allows the hair-shaft to keep contact when it is bent under the stimulus.