Synthesis of [6″-H]-, (6″-H)- and (2-H)-maltotriose

To prepare labeled precursors for biosynthetic studies, methods for the specific introduction of tritium and deuterium into the reducing and the terminal glucose unit of maltotriose were developed. Thus [6″-³H]- and (6″-²H)-maltotriose (17) and (18) were prepared via selective methoxytritylation, deprotection and subsequent modified Pfitzner-Moffatt oxidation, followed by reduction with sodium borotritiide or sodium borodeuteride, respectively. A simple two step procedure utilizing the Lobry de Bruyn/van Ekenstein transformation gave (2-²H)maltotriose (20).     

Contribution of Monsoon Asia to the carbon budget of the biosphere,past and future

The High Resolution Biosphere Model (HRBM), which has been developed by the group of the author, was used to investigate the carbon balance of the vegetation and the soil in the ecosystems of Monsoon Asia in comparison to the rest of the world. The HRBM is a global grid-based (0.5 degree resolution) model with a monthly time step. It includes modules for natural vegetation, land use, vegetation fires, vegetation composition. A historical carbon budget was calculated for the period 1860–1978 and, on a global scale, validated using atmospheric CO₂ data. Based on the per-country development of the population and their requirements, different reasonable scenarios were used to investigate the potential impacts of land use and deforestation in the period 1990–2050. The HRBM calculates considerable contributions of Monsoon Asia to the global CO₂ emissions due to land use changes in the past. Between 1860 and 1978, about 1/4 of the global releases from land use changes came from South Asian and Southeast Asian biota. The future contributions in the period 1990–2050 depend on the assumed development of the agricultural methods. If the intensity of agriculture and the agricultural productivity will stay the same as in the 1980s, there will be a strong need to increase agricultural areas, and thus deforestation will dominate. If there will be a change over to intensive methods of agricultural production, the presently used areas might be sufficient to provide resources to the growing population.     

Metabolism of Pyrene by the Basidiomycete Crinipellis stipitaria and Identification of Pyrenequinones and Their Hydroxylated Precursors in Strain JK375

The metabolism of pyrene, a polycyclic aromatic hydrocarbon, by submerged cultures of the basidiomycete Crinipellis stipitaria was studied. After incubation for 68 h at 25°C in a 20-liter fermentor with complex medium and 20 mg of pyrene per liter, five metabolites were detected. The compounds were isolated by preparative high-performance liquid chromatography on RP18 and DIOL gels. By UV, infrared, and ¹H nuclear magnetic resonance spectroscopy and mass spectrometry, 1-hydroxypyrene, 1,6-dihydroxypyrene, 1,8-dihydroxypyrene, 1,6-pyrenequinone, and 1,8-pyrenequinone were identified. 1,6- and 1,8-dihydroxypyrene were obtained from fungal cultures for the first time. The formation of these metabolites was confirmed by investigations with [4,5,9,10-¹⁴C]pyrene.     

α2-Adrenoceptor-Mediated Inhibition of Electrically Evoked [3H]Noradrenaline Release from Chick Sympathetic Neurons: Role of Cyclic AMP

Abstract: This study explores the role of cyclic AMP in electrically evoked [³H]noradrenaline release and in the α₂-adrenergic modulation of this release in chick sympathetic neurons. Along with an increase in stimulation-evoked tritium overflow, applications of forskolin enhanced the formation of intracellular cyclic AMP. Both effects of forskolin were potentiated by the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine. The forskolin-induced increase in overflow was abolished by the Rp-diastereomer of cyclic AMP-thioate, an antagonist at cyclic AMP-dependent protein kinases, and 1,9-dideoxy-forskolin, an inactive analogue at adenylyl cyclase, had no effect on the evoked overflow. A 24-h pretreatment with either cholera toxin or forskolin reduced the subsequent forskolin-induced accumulation of cyclic AMP and inhibited the stimulation-evoked release. Basal cyclic AMP production, however, remained unaltered after forskolin treatment and was enhanced after 24 h of cholera toxin exposure. The α₂-adrenergic agonist bromoxidine did not affect the formation of cyclic AMP stimulated by forskolin but reduced electrically evoked release. However, effects of bromoxidine on ³H overflow were attenuated by forskolin as well as by 8-bromo-cyclic AMP. Effects of bromoxidine on [³H]noradrenaline release were paralleled by an inhibition of voltage-activated Ca²⁺ currents, primarily through a delayed time course of current activation. This effect was abolished when either forskolin or 8-bromo-cyclic AMP was included in the pipette solution. Both substances, however, failed to affect Ca²⁺ currents in the absence of bromoxidine. These results suggest that the signaling cascade of the α₂-adrenergic inhibition of noradrenaline release involves voltage-activated Ca²⁺ channels but not cyclic AMP. Elevated levels of cyclic AMP, however, antagonize this α₂-adrenergic reduction, apparently through a disinhibition of Ca²⁺ channels.     

Pool sequencing of natural HLA-DR,DQ, and DP ligands reveals detailed peptide motifs,constraints of processing,and general rules

We have approached the problem of MHC class II ligand motifs by pool sequencing natural peptides eluted from HLA-DR, DQ, and DP molecules. The results indicate surprisingly clear patterns, although not quite as clear as with natural class I ligands. The most striking feature is a highly dominant Proline at position 2. We interpret this to be a consequence of aminopeptidase N-like activity in processing. Another general aspect is the existence of three to four hydrophobic or aromatic anchors, whereby the first and the last are separated by five to eight residues. The peptide motifs for HLA-DR1, DR5, DQ7, and DPw4 are allele-specific and differ by spacing and occupancy of anchors. The anchors tend to be flanked by clusters of charged residues, and small residues, especially Ala, are frequent in the motif centers. These detailed motifs allow one to interpret most previous (DR-) motifs as fitting one or more of the anchors or conserved clusters. The relative motif symmetry suggests the possibility of bidirectional binding of peptides in the class II groove.     

PET1402, a nuclear gene required for proteolytic processing of cytochrome oxidase subunit 2 in yeast

The nuclear mutation pet ts1402 prevents proteolytic processing of the precursor of cytochrome oxidase subunit 2 (cox2) in Saccharomyces cerevisiae. The structural gene PET1402 was isolated by genetic complementation of the temperature-sensitive mutation. DNA sequence analysis identified a 1206-bp open reading frame, which is located 215 by upstream of the PET122 gene. The DNA sequence of PET1402 predicts a hydrophobic, integral membrane protein with four transmembrane segments and a typical mitochondrial targeting sequence. Weak sequence similarity was found to two bacterial proteins of unknown function. Haploid cells containing a null allelle of PET1402 are respiratory deficient.     

Changes in in vivo fluorescence quenching in rye and barley as a function of reduced PSII light harvesting antenna size

The relationship between the size of the light harvesting antenna to photosystem II (LHCII) and quenching of non-photochemical and dark level fluorescence was studied in wild-type rye (Secale cereale L. cv. Musketeer) and barley (Hordeum vulgare L. cv. Gunilla) as well as in the barley chlorophyll b-less chlorina F2 mutant (H. vulgare L. cv. Dornaria, chlorina-F2). Exposure for 10 min to an irradiance of 500 μmol m^?2 s^?1 resulted in a strong (0.71–0.73) non-photochemical (q_s) quenching of the fluorescence yield in wild-type (WT) material, while the barley chlorina F2-mutant was quenched to 75% of this level. Relaxation of q_s in darkness revealed a fast initial decay, related to relaxation of the high-energy-state dependent (q_E) part of q_s. Etiolated seedlings of rye and barley exposed to intermittent light (IML) for 36 cycles of 2 min light and 118 min darkness had suppressed Chl b and LHCII-production in both WT rye and barley, while the barley chlorina F2-mutant became totally devoid of all LHCII-polypeptides. It was found that the levels of q_s and q_s were similar in control grown barley chlorina F2 and IML-grown WT rye and barley, but q_s was reduced by 30 to 35% and q_s by 50 to 65%, respectively, as compared to control-grown. WT plants. No significant q_s could be detected in IML-grown barley chlorina F2. It is clear, from these changes in in vivo fluorescence quenching in rye and barley that a significant level of q_s is detectable even in the absence of LHCII. Only when the proximal antennae are totally absent, does q_E completely disappear. We conclude that the presence of LHCII is not an absolute requirement for q_E-quenching and suggest that distal as well as proximal antenna may contribute to q_E in vivo.     

Streptococcal erythrogenic toxin type C is not a phosphorylated protein. Description of two different purification procedures and investigation of its phosphorylation state

Abstract Erythrogenic toxin type C (ETC) from different streptococcal group A strains was successively purified by absorption on phenylsepharose, acidic dialysis of the eluate at 40% saturated ammonium sulphate solution, CM-Sepharose chromatography, finally by immunoaffinity chromatography on monoclonal antibodies. Second, after growing of bacteria in the presence of [³²P]orthophosphate to phosphorylate ETC, the ETC was purified with phenylsepharose following immunoaffinity chromatography. The occurrence of phosphoamino acids in the purified ETC was investigated by an immunoassay. No phosphoamino acids could be detected in the ETC molecule. Also after radiolabelling with ³²P it was not possible to demonstrate a radioactive signal. The treatment with alkaline phosphatase has no influence on the mitogenicity or position of ETC in isoelectric focusing. The results obtained led to the conclusion that in contrast to the literature, ETC is not a phosphorylated protein.