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Synechococcus (Anacystis nidulans, strain L 1402-1) were grown at + 37 °C in an atmosphere of 0.04 vol.% CO2 using different light conditions. Changing the culture conditions caused alterations in pigment ratios and ultrastructure of Synechococcus. In comparison to the low white and red light grown cells under strong white light the number of thylakoids decreased and an accumulation of storage carbohydrates could be observed. The number of the polyhedral bodies also varied with culture conditions. The results are discussed with reference to the pigment composition and the function of the polyhedral bodies.
14C-and 15N-Assimilation, 15N-Labelled Amino Acids, M arine D iatom s The marine diatoms Bellerochea yucatanensis and Skeletonema costatum were grown at +20 °C in 0.03 vol.% CO2 with nitrate or ammonia. The 15N -am m onia and 15N -nitrate assim ila tion and 15N -incorporation into various amino acids were studied of both diatom s during exponential growth phase in dependence of different nitrogen conditions. In all experiments the 15N -am m onia uptake was lower than the 15N -nitrate assim ilation rate up to 20-40 min photo synthesis. N itrate lim itation -cells grown in nitrate followed by growth in nitrogen-free m edium for 24 h — caused a strong 15N-label into aspartate after adding 15NH 4C1 (1 m M). In cells grown in nitrate highest enrichment of 15N was found in glutamine. Results were discussed with reference to the operating of the GS/GOGAT system and glutam ic acid dehydrogenase pathway. Photosynthetic 14CO2 fixation experiments showed a very high labelling of aspartate which was interpreted with a phosphoenolpyruvate carboxylation catalysed by phosphoenolpyruvate carb-oxykinase.
Chromatin, RNA Polymerase, Potato Tuber Tissue, Aging Phenomenon The synthesis of RNA by chromatin-bound RNA polymerase (E.C. 2.7.7.6.) from white potato tubers proceeds at a low rate, which is enhanced after slicing the tissue, however. Concomitantly DNA template availability as measured with saturating amounts of Escherichia coli polymerase is diminished drastically. Nearest neighbor frequency analysis proved that the RNA synthesized on chromatin of intact tubers is different from that synthesized on chromatin of sliced tissue.
The RNA polymerase of white potato tubers is dependent on all four ribonucleoside triphos phates and a divalent metal ion such as Mg2+ or Mn2+ and totally inhibited by the presence of pyrophosphate. Actinomycin D blocks the formation of the RNA product, which could be shown to be a heteropolymer by nearest neighbour frequency technique. The Km of the chromatin-bound enzyme with regard to ATP, GTP, CTP and UTP was 5.1 X10-5 M, 1.6X10-5 M, 0.9X10-5 M and 0.45 X 10-5M/1 respectively, α-amanitin inhibits the overall activity to about 50%, which indicates the presence of equal amounts of polymerase I and polymerase If.
A screening procedure is presented which allows the isolation of yeast mutants (typ tlr) with highly efficient utilization of exogenous deoxythymidine-5′-monophosphate (5′-dTMP) (>50% ). Data are given concerning the phenomenon of 5′-dTMP utilization in general: (i) The ability of S. cerevisiae to incorporate exogenous 5′-dTMP was found to already be a wild type feature of this yeast, i. e. apparently not to be due to any mutation such as typ , tup, tmp per or tum. Consequently these mutations are interpreted as amplifiers of a pre-given wild type potency. So far eight stages of 5′-dTMP utilization were detected as classified by the optimal 5′-dTMP requirement, with 5′-dTMP biosynthesis blocked, of the corresponding mutant strains isolated. All of them fit well into a mathematical series of the type “2n × 1.5” (n = 0, 1, 2, … , 11), where the product term for n = 11 represents the 5′-dTMP requirement (μg/ml) of the best 5′-dTMP utilizing wild type strain found, (ii) Amplification of the 5′-dTMP utilizing potency obviously is due to any genetically determined alteration of the yeast 5′-dTMP uptaking principle itself or of physiological processes accompanying the monophosphate’s uptake, (iii) The functioning of 5′-dTMP uptake requires acidic (≦ pH 6) conditions in the yeast cell’s outer environment, (iv) Some yeast typ and typ tlr mutants were found to exhibit a more or less pronounced sensitivity towards exogenously offered 5′dTM P. The response of a sensitive strain towards inhibitory concentrations of the nucleotide apparently is co-conditioned by the presence or absence of thymidylate biosynthesis. With 5′-dTMP biosynthesis blocked the 5′-dTMP mediated inhibition is a permanent one and finally leads to the death of a cell. With a functioning thymidylate biosynthesis, in contrast, the inhibition is only temporary, (v) Yeast typ or typ tlr strains were observed to dephosphorylate exogenous 5′-dTMP to thymidine due to a phosphatase activity which cannot be eliminated at pH 7 + 70 mм inorganic phosphate conditions in the growth medium. This 5′-dTMP cleavage obviously occurs outside the cell and does not seem to be correlated both to the monophosphate’s uptake and to the phenomenon of 5′-dTMP sensitivity. The destruction of 5′-dTMP does not disturb (5′-dTMP) DNA-specific labelling.
In haploid and diploid S. cerevisiae the dimer yield ratio TT̂/CT̂ is found to be 1.2/1 and 1.3/1, resp., at the UV (254 nm) unit dose 1 erg/mm2, the share of TT̂ and CT̂ in a UV (254 nm) lethal hit being 0.7 TT̂ and 0.6 CT̂. A general formulation of the UV lethal hit is given and discussed. The TT̂ + CT̂ yields obtained for S. cerevisiae are compared to those reported for other organisms. It is found that there obviously exists a directly proportional linear correlation between genome size and TT̂ + CT̂ yield for the UV dose range well below the stationary levels of the TT̂ and CT̂ formation kinetics.
Testosterone, Androst-4-en-3,17-dione, Enzyme Induction, S trep to m yces hydrogenans After cultivation of S trep to m yces hydrogenan s in the presence of 3H-labelled testosterone, radio active steroids were extracted separately from the cytosolic, ribosomal and cell wall-membrane fraction of the cells and from the culture medium, respectively.. The separation of the steroids was performed by one-and two-dimensional thin layer chromatography (TLC). The identification of the main metabolites was achieved by crystallization to constant specific radioactivity, specific staining procedures and acetylation. The oxidation of testosterone to androst-4-en-3,17-dione is by far the predominating reaction, which is almost finished after 3 h cultivation. Androst-4-en-3,17-dione is mainly transferred into the culture medium and partly accumulated within the cell wall-membrane fraction. High polar steroid metabolites and androstane derivatives are present in very small amounts only.
Resting potato tuber tissue possesses only faint activity of the two dehydrogenases of the oxidative pentose phosphate cycle, glucose-6-phosphate- and 6-phosphogluconate dehydrogenase. Slicing of the tissue, however, greatly enhances the action of both enzymes. The slicing-induced increase in activity is a consequence of intensified action of at least 5 glucose-6-phosphate dehydrogenase isozymes and a more differentiated activation/inactivation of seven 6-phosphogluconate dehydrogenase isozymes.
Using density labelling and isopycnic equilibrium centrifugation it could be demonstrated, that the bulk of both enzymes appearing after slicing the tissue is the result of de novo synthesis rather than activation of pre-existing proenzymes.
An improved method for isolation of yeast m utants auxotrophic for 5′-dTM P is presented. The procedure employs the two folic acid antagonists am inopterin and sulfanilam ide (SAA). Selectiveness of the procedure depends on concentration of SAA and time of incubation.
44 mutants auxotrophic and 3 conditionally auxotrophic for 5′-dTMP were isolated. All belong to one complementation group. The corresponding gene was designated TMP1. Tetrad dissection revealed its chromosomal nature. TMP1 is not closely linked to the genes ADE2,, LEU1, ARG 4, ILV2, HIS5, LYS1 and the mating type locus. With the centromere-linked genes ARG4 and LEU1 I gene TMP1 exhibited second division segregation frequencies of 0.42 and 0.53 respectively, indicative of centromere-linkage.
Strains auxotrophic and conditionally auxotrophic for 5′-dTM P were all respiratory deficient (petite). Genetical analysis indicates that the petite phenotype is due to loss of the rho factor in cells harbouring either tmp1 or tmp1ts alleles.
One of the earliest consequences of slicing plant storage organs such as potato tubers into thin disks is the formation of polysomes, which in potato slices is complete after 9 hours and is dependent on transcription. Fresh disks do not incorporate 32P, 3H-uridine or 14C-leucine into their ribosomes, whereas ribosomes and polysomes of aged disks use these precursors effectively. This development can be completely blocked by actinomycin D. Among the different RNAs synthesized during aging is 28S- and 16S—rRNA, 5S—RNA, tRNA, and a component sedimenting around 15—18S with a base-composition different from 16S—rRNA, 5S- and 4S—RNA and which supports peptide formation in an in vitro incorporation system.
It is suggested that this compound represents mRNA, which is not available immediately after slicing the tissue. These findings are consistent with the view of a derepression phenomenon in sliced storage tissue.