Chemical studies on the lipopolysaccharide of Rhizobium meliloti 10406 and its lipid A region

The structure of the lipopolysaccharide from Rhizobium meliloti 10406, a derivative of the wild-type strain MVII-1, was examined. The compositional analysis of its polysaccharide moiety demonstrated lack of heptose(s), but high contents in glucose, galacturonic acid and 2-keto-3-deoxy-octonate (dOclA) as characteristic features. The lipid A moiety consisted of a -1,6 linked glucosamine disaccharide carrying ester (at C-4) and glycosidically (at C-1) linked phosphate residues, both present exclusively as monoester phosphates but not as phosphodiesters. Ester- and amidelinked 3-hydroxy fatty acids were mostly present as non-3-O-acylated residues. Laser desorption mass spectrometry (LD-MS) revealed heterogeneity in the fatty acid substitution, as was also indicated by the non-stoichiometric ratios obtained by quantitative fatty acid analysis. The predominating lipid A structure contained at the reducing glucosamine residue ester-linked 3-hydroxy-tetradecanoic acid (3-OH-14:0) and amide-linked 3-OH-18:0, or 3-OH-18:1, respectively. The distal (non-reducing) glucosamine carried ester-bound the recently discovered 27-hydroxyoctacosanoic acid and 3-OH-14:0 and, as amide-linked fatty acid, mostly 3-hydroxy-stearic acid (3-OH-18:0).The isolated lipopolysaccharide exhibited a high extent of lethal toxicity in galactosamine-treated mice, comparable to that of enterobacterial lipopolysaccharide. The structural relationship of LPS and lipid A of Rhizobium meliloti to other rhizobial lipopolysaccharides and lipid A's with respect to questions of taxonomy and of phylogenetic relationships will be discussed.Abbreviations LPS lipopolysaccharide - dOclA 3-deoxy-D-mannooctulosonic acid (KDO) - GalA galacturonic acid - DOC sodium deoxycholate - PAGE polyacrylamide gel electrophoresis - LD-MS laser desorption-mass spectrometry     

Nontoxic and toxic oligopeptides with D-amino acids and unusual residues in Microcystis aeruginosa PCC 7806

Toxic and nontoxic peptides were isolated from the cyanobacterium Microcystis aeruginosa PCC 7806 by a procedure including extraction of cells with water-saturated 1-butanol, chromatography of the extract on silica gel plates and high performance liquid chromatography (HPLC) on Partisil-5. The toxin was shown to be only a minor constituent, being negatively charged and thus separable by electrophoresis, within the HPLC-purified fraction. It contained erythro-β-methyl-D-Asp, D-Glu, D-Ala, L-Leu, and L-Arg known to be part of the Microcystis peptide-toxin with M_r 994. The major part of the HPLC-purified fraction was assigned, however, to a nontoxic peptide with a M_r of 956. Partial hydrolysis studies of the nontoxic peptide(s) revealed amino acid sequences composed of D-Glu, N-methyl-Phe, and 3,4-dehydro-Pro, aside from the common L-amino acids. Cyclic linkage in the nontoxic peptide(s) appears likely.     

Functional role of newly formed pore complexes in postmitotic nuclear reorganization

Many nuclear proteins are released into the cytoplasm at prometaphase and are transported back into the daughter nuclei at the end of mitosis. To determine the role of this reentry in nuclear remodelling during early interphase, we experimentally manipulated nuclear protein uptake in dividing cells. Recently we and others have shown that signal-dependent, pore complex-mediated uptake of nuclear protein is blocked in living cells on microinjection of the lectin wheat germ agglutinin (WGA), or of antibodies such as PI1 that are directed against WGA-binding pore complex glycoproteins. In the present study, we microinjected mitotic PtK₂ cells with WGA or antibody PI1 and followed nuclear reorganization of the daughter cells by immunofluorescence and electron microscopy. The inhibitory effect on nuclear protein uptake was monitored by co-injection of the karyophilic protein nucleoplasmin. When injected by itself early in mitosis, nucleoplasmin became sequestered into the daughter nuclei as they entered telophase. In contrast, nucleoplasmin was excluded from the daughter nuclei in the presence of WGA or antibody PI1. Although PtK₂ cells with blocked nuclear protein uptake completed cytokinesis, their nuclei showed a telophaselike organization characterized by highly condensed chromatin surrounded by a nuclear envelope containing a few pore complexes. These findings suggest that pore complexes become functional as early as telophase, in close coincidence with nuclear envelope reformation. They further indicate that the extensive structural rearrangement of the nucleus during the telophase-G1 transition is dependent on the influx of karyophilic proteins from the cytoplasm through the pore complexes, and is not due solely to chromosome-associated components.Abbreviations WGA wheat germ agglutinin - GlcNAc N-acetylglucosamine     

S-adenosyl-L-methionine:trans-caffeoyl-coenzyme A 3-O-methyltransferase from elicitor-treated parsley cell suspension cultures

An S-adenosyl-L-methionine:caffeoyl-CoA 3-O-methyltransferase was purified 82-fold from elicitor-induced parsley cell suspension cultures by ammonium sulfate fractionation, anionic exchange and hydrophobic interaction chromatographies, and chromatofocusing. The enzyme has an apparent pI of 5.7 and a molecular weight of approx 48,000 determined by gel filtration chromatography. Maximal activity was observed at pH 7.5 in 50 mM phosphate or Tris-HCl buffers and the additional presence of 0.5 M NaCl. The methyltransferase activity was dependent on Mg2+, whereas EDTA, Mn2+, and Ca2+ inhibited the reaction. The partially purified enzyme efficiently catalyzed the methylation of caffeoyl-CoA, but also accepted with low affinity various other caffeic esters as substrates. Dark-grown parsley cells contained considerable methyltransferase activity which was nevertheless increased approx threefold within 12 h following the addition of a crude fungal elicitor to the cell suspensions. We propose that the O-methyltransferase activity is an important component in the rapid resistance response of the cells, which depends on the formation of cell wall-bound ferulic polymers.     

Shifts in Methanogenic Subpopulations Measured with Antibody Probes in a Fixed-Bed Loop Anaerobic Bioreactor Treating Sulfite Evaporator Condensate

A fixed-bed loop, high-rate anaerobic bioreactor treating sulfite evaporator condensate was sampled when it reached steady state and afterwards following perturbations during a 14-month period. By using immunotechnology, it was observed that shifts in methanogenic subpopulations occurred in association with perturbations, such as restarting and relocating the biomass into a different tank. Methanogens related to Methanobacterium bryantii MoHG and Methanobrevibacter smithii ALI were numerous throughout the observation period, while Methanosarcina mazei S6 and Methanosarcina thermophila TM1 were found in the early and late samples, respectively. Also, Methanobacterium formicicum was more numerous at the top portion of the bioreactor, while Methanobrevibacter arboriphilus AZ and DC were at the bottom. Sample formalinization required for prolonged storage proved suitable for antigen preservation.     

Seasonal variation of calcium,magnesium, potassium,and manganese contents in xylem sap of beech(Fagus sylvatica L.) in a 35-year-old limestone beech forest stand

Summary In a 35-year-old calcareous beech forest stand five beech trees (Fagus sylvatica L.) were felled every 2 weeks, and xylem sap was obtained by means of water displacement from the lowest trunk sections, each 100 cm in length. From mid-October 1988 to mid-October 1989 a total of 130 trees were investigated. The seasonal variations of the Ca, Mg, K and Mn contents, as well as those of pH, show four characteristic phases. Additionally, distribution of the mineral contents along the trunk was studied in four trees. The seasonal increase and decrease of xylem sap mineral contents along the trunk is shown for the characteristic phenophases. The Ca, Mg, K, and Mn contents of xylem saps were determined by means of atomicabsorptionspectrophotometry.     

A new recombinant procoagulant protein derived from the cDNA encoding human factor VIII

We have constructed new B domain deletion derivatives of human factor VIII (FVIII) by manipulating the cDNA using recombinant DNA techniques. One of these new derivatives, FVIII delta II, in which amino acids 771(pro)-1666(asp) have been deleted, no longer contains the protease cleavage site at amino acid position 1648(arg)-1649(glu) known to be involved in the initial step of FVIII processing. We have expressed this molecule in both baby hamster kidney (BHK) 21 cells using the vaccinia virus (VV) expression system and have established Chinese hamster ovary (CHO) derived permanent cell lines expressing either recombinant (r)FVIII or FVIII delta II. The characteristics of FVIII delta II have been compared to those of rFVIII and/or plasma derived (pd) FVIII. FVIII delta II has the following properties: (i) it exhibits FVIII procoagulant activity; (ii) it is expressed at 5-fold higher levels than is the complete molecule in comparable systems; (iii) it migrates for the most part as a single major band on SDS-PAGE, in contrast to the complete molecule; (iv) it is activated to a greater extent by thrombin than is either rFVIII or pdFVIII; and (v) it retains the ability to bind von Willebrand factor (vWf).     

The stimulating effect of a cold, dark pretreatment on the etioplast/chloroplast transformation of angiosperms. I. The stimulating effect of cold predarkening on different stages of greening under white light

Schönbohm, E., Stute, U., Thienhaus, P. and Werner, U. 1988. The stimulating effect of a cold, dark pretreatment on the etioplast/chloroplast transformation of angiosperms I. The stimulating effect of cold predarkening on different stages of greening under white light. - Physiol. Plant. 72: 541–546.
The etioplast/chloroplast transformation in angiosperms is controlled by light; most of the processes are mediated by phytochrome. We have shown that in the primary leaves of etiolated seedlings of wheat ( Triticum aestivum L. cv. Kolibri), fire-bean ( Phaseolus multiflorus L. cv. Preisgewinner) and in the cotyledons of etiolated sun flower seedlings ( Helianthus annuus L. cv. macrocarpa) the chlorophyll accumulation in the phase after the end of the lag phase can be greatly stimulated by a cold predarkening period. This effect is not necessarily coupled with a red preirradiation. Furthermore the lag phase can be dramatically shortened by the cold, dark pretreatment, whereas the amount of photoconvertible protochlorophyll(ide) in the darkness remains unaffected by the cold, dark pretreatment. The stimulating effect of a cold, predarkening period on greening is fully reversible by a warm, dark phase that is placed between the cold period and the onset of the continuous white light phase. These findings cannot be generalized: We could demonstrate that in the tropical plant Momordica charantia greening under white light was not affected by different temperature pretreatments during predarkening. The stimulating effect of a cold, predarkening period on greening is assumed to have ecological relevance.     

Twelve loci form a continuous linkage map for human chromosome 18

We have constructed a primary genetic map of human chromosome 18 consisting of 11 DNA markers and one serological marker (JK). Two of these loci define highly polymorphic VNTR systems. The markers define a continuous genetic linkage map of 97 cM in males and 205 cM in females; female genetic distances in a panel of 59 three-generation families were consistently about twice those observed in males. The high odds in support of the linear order of the markers on this recombination map, and the extent of coverage of chromosome 18, indicate that this map will permit efficient linkage studies of human genetic diseases that may be segregating on chromosome 18 and will provide anchor points for development of high-resolution maps for this chromosome.