Abbau von 14C-, 3H- und 36Cl-markiertem γ-Hexachlorcyclohexan durch anaerobe Bodenmikroorganismen

Zusammenfassung Durch eine anaerobe Mischflora aus Ackerboden wurde -Hexachlorcyclohexan (-HCH) in 4–5 Tagen zu 90% abgebaut. Dabei erfolgte eine schnelle Abspaltung des Chlors in Form von Chloridionen und danach eine Freisetzung des C- und H-Anteiles in Form flüchtiger Verbindungen, in denen kein Chlor und auch kein CO₂ nachzuweisen war.Die Verwendung von ¹⁴C/³H- und ³⁶Cl/³H-doppelmarkiertem -HCH zeigte, daß die Cl- und H-Abspaltung nicht im Verhältnis von 1:1 erfolgte, sondern mehr Cl als H abgespalten wurde. Die flüchtigen Verbindungen enthielten andererseits höhere ¹⁴C- als ³H-Anteile. Gaschromatographische Untersuchungen zeigten ebenfalls eine rasche Verminderung des -HCH und die Bildung verschiedener Metabolite. Es wurde jedoch kein -Pentachlorcyclohexen nachgewiesen. Bei steigenden O₂-Gehalten in der Gasphase verminderte sich der -HCH-Abbau. Jedoch fanden auch noch bei 5% O₂ Chlorabspaltung und die Freisetzung flüchtiger Metabolite statt.-HCH wurde ebenfalls, jedoch langsamer, durch die anaerobe Mischflora abgebaut. Auch hier wurde Chlorid abgespalten, und es traten ebenfalls flüchtige Verbindungen auf, die kein Chlor enthielten.

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Degradation of ¹⁴C-, ³H- and ³⁶Cl-labelled -hexachlorocyclohexane by anaerobic soil microorganisms

Up to 90% of the -Hexachlorocyclohexane (-HCH) applied to an anaerobic mixed bacterial flora enriched from an arable soil were degraded within 4–5 days. Degradation resulted in a rapid release of chloride and in formation of chlorine-free volatile metabolites. CO₂ formation from the molecule was not detected.Investigations with ¹⁴C/³H- and ³⁶Cl/³H double-labelled -HCH indicated that the release of Cl and H did not occur in the ratio of 1:1. More Cl than H was split off. The volatile compounds contained more ¹⁴C than ³H. Gas chromatographic studies also showed the rapid decrease of -HCH and the formation of several metabolites. -Pentachlorocyclohexene was not detected. Increasing O₂-contents in the gas phase of cultures resulted in decreases of the compound's degradation. Release of chloride and of volatile metabolites were observed with O₂ contents in the gas phase up to 5%.-HCH was also, but more slowly as with -HCH, degraded by the anaerobic mixed flora. Chloride was released and volatile, chlorine-free metabolites were found.

     

UDP-GlcNAc: Galβ3GalNAc-Mucin: (GlcNAc → GalNAc) β6-N-Acetylglucosaminyltransferase and UDP-GlcNAc: Galβ3(GlcNAcβ6) GalNAc-Mucin (GlcNAc → Gal)β3-N-Acetylglucosaminyltransferase from Swine Trachea Epithelium

Summary Two specific -N-acetylglucosaminyltransferases involved in the branching and elongation of mucin oligosaccharide chains, namely, a 1,6 N-acetylglucosaminylsaminyltransferase that transfers N-acetylglucosamine from UDP-N-acetylglucosamine to Gal3GalNAc-Mucin to yield Gal3(GlcNAc6)GalNAc-Mucin and a 3-N-acetylglucosaminyl transferase that transfers N-acetylglucosamine from UDP-N-acetylglucosamine to Gal3(GlcNAC6)GalNAc-mucin to yield GlcNAc3Gal3 (GlcNAc6)GalNAc-Mucin were purified from the microsomal fraction of swine trachea epithelium. The 1,6-N-acetylglucosaminyltransferase was purified about 21,800-fold by procedures which included affinity chromatography on DEAE columns containing bound asialo Cowper's gland mucin glycoprotein with Gal1,3GalNAc side chains. The apparent molecular weight estimated by gel filtration was found to be about 60 Kd. The purified enzyme showed a high specificity for Gal1,3GalNAc chains and the most active substrates were mucin glycoproteins containing these chains. The apparent Km of the 6-glucosaminyltrans-ferase for Cowper's gland mucin glycoprotein containing Gal1,3GalNAc chains was 0.53 µM; for UDP-N-acetylglucosamine, 12 µM; and for Gal 1,3GalNAc NO₂ø, 4 mM. The activity of the 6-glucosaminyltransferase was dependent on the extent of glycosylation of the Gal3GalNAc chains in Cowper's gland mucin glycoprotein.The best substrate for the partially purified 3-Glucosaminyltransferase was Cowper's gland mucin glycoprotein containing Gal1,3(GlcNAc6)GalNAc side chains. This enzyme showed little or no activity with intact sialylated Cowper's gland mucin glycoprotein or derivatives of this glycoprotein containing GalNAc or Gal1,3GalNAc side chains.The radioactive oligosaccharides formed by these enzymes in large scale reaction mixtures were released from the mucin glycoproteins by treatment with alkaline borohydride, isolated by gel filtration on Bio-Gel P-6 and characterized by methylation analysis and sequential digestion with exoglycosidases. The oligosaccharide products formed by the 6- and 3-glucosaminyltransferases were shown to be Gal3(GlcNAC6) GalNAc and GlcNAc3 Gal3(GlcNAC6)GalNAc respectively.Taken collectively, these results demonstrate that swine trachea epithelium contains two specific N-acetylglucosaminyltransferases which catalyze the initial branching and elongation reactions involved in the synthesis of O-linked oligosaccharide chains in respiratory mucin glycoproteins. The first enzyme a 6-glucosaminyltransferase converts Gal3GalNAc chains in mucin glycoproteins to Gal3(GlcNAc6)GalNAc chains. This product is the substrate for a second 3-glucosaminyltransferase which converts the Gal3(GlcNAc6)GalNAc chains to GlcNAc3Gal(GlcNAc6)GalNAc chains in the glycoprotein. The 3-glucosaminyltransferase did not utilize Gal3GalNAc chains as a substrate and this results in an ordered sequence of addition of N-acetylglucosamine residues to growing oligosaccharide chains in tracheal mucin glycoproteins.Abbreviations NeuNAc N-acetylneuraminic acid - GalNAcol N-acetylgalactosaminitol - CGMG Cowper's gland mucin glycoprotein - GalNAc-CGMG Cowper's gland mucin glycoprotein containing GalNAc side chains O-glycosidically linked to serine or threonine - Gal3GalNAc-CGMC Cowper's gland mucin glycoprotein containing Gal3GalNAc side chains - MES 2-(N-morpholino) Ethane Sulfonic acid - PBS Phosphate Buffered Saline     

Methodological problems in evolutionary biology VII. The species plague

Various philosophers and evolutionary biologists have recently defended the thesis that species are individuals rather than sets. A decade of debates, however, did not suffice to settle the matter. Conceptual analysis shows that many of the key terms involved (individuation, evolutionary species, spatiotemporal restrictedness, individual) are ambiguous. Current disagreements should dissolve once this is recognized. Explication of the concepts involved leads to new programs for philosophical research. It could also help biology by showing how extant controversies concerning evolution may have conceptual rather than factual roots.     

Harzkonservierte fossile Vogelfedern aus der untersten Kreide

Zusammenfassung Harzkonservierte Fossilien ermöglichen bei Anwendung adäquater Methoden die morphologische Analyse der Feinmerkmale bis zur Auflösungsgrenze des Lichtmikroskops, Beobachtung in verschiedenen Ebenen und Richtungen, und somit konkrete Rückschlüsse auf die Wirkung und Bedeutung der Einzelelemente und des Gesamtgefüges.Eine so eingehende funktionsmorphologische Analyse mit Berücksichtigung der Positionsvariation (graduell verschiedene Gestaltung in gesetzmäßiger Abhängigkeit von der Lage innerhalb der Gesamtfeder) der Einzelelemente wie Abzweigungs-, Knick-, Neigungswinkel, Krümmung, Länge, Dicke, Querschnitt, Dichte, Differenzierungsgrad der verschiedenen Abschnitte von Rhachis, Rami, Radii inklusive Häkchen und Cirren wird erstmals für fossile Vogelfedern geliefert (hier als Abriß zu einer dokumentarisch und thematisch ausführlicheren Darstellung in Stuttgarter Beiträge zur Naturkunde).Diese Federn entstammen der untersten Unterkreide und sind damit nur relativ wenig jünger alsArchaeopteryx. Sie weisen extrem differenzierten Aufbau auf, der auf hohe flugtechnische und wärmeisolierende Leistungsfähigkeit schließen läßt.Die hier vorgelegten funktionsmorphologischen Ermittlungen an fossilen Körperkonturfedern mögen auch zu einer intensiveren Analyse der bis jetzt stark vernachlässigten Untersuchung ganz normaler Körperfedern rezenter Vögel anregen. Erst dann, nach umfassender Kenntnis ihrer Ausgestaltung innerhalb der verschiedensten rezenten Vogelgruppen, läßt sich überzeugend begründen, ob und wieweit die hier vorgelegten Federn dieses Unterkreide-Vogels noch ursprüngliche Elemente (Plesiomorphien) oder ihnen eigene Sonderbildungen (Autapomorphien) aufweisen; das gilt sowohl für morphologische wie für funktionelle Elemente der Gesamtstruktur.

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Resin-preserved fossil bird's feathers from the Lowermost Cretaceous

Summary Parts of some feathers, originating from a single bird, were discovered in our collections of Lower Cretaceous amber from the Lebanon mountains — which, in general, contains the oldest terrestrial microfossils preserved with all morphological details.These contour feathers of the trunk, which are nearly as old as Archaeopteryx (Lowermost Cretaceous: Neocomian/Uppermost Jurassic: Kimmeridigian) were studied with magnifications of 500–900 in several levels by a special technique. (In normal fossils, i.e., impressions, the granulation of the sediment and the fossil's bulky carbon remainders cause a blurred image even at a magnification of merely 100).Special emphasis was laid on the study of the individual elements' gradual variation, depending on the respective position within the total feather (position variation). Where appropriate, an analysis of lengths, quantity, degree of differentiation, angle of inclination, break, and branching, cross-sectional view, curvature, etc. of the rhachis, rami, distal and proximal radii, barbicles, hooklets, etc. were undertaken. [Through measurements of the depth of details the effects caused by a sloping position (apparent variation) may be precisely separated from the real variation.]On the basis of such a detailed knowledge of structure and relative position a thorough functional analysis of the single elements as well as the total system is given.Principal features: The production of plain stability in the feather's center, and of flexibility in its apical and lateral rims; dispersion of forces in case of pressure or a pulling load; function of the hooklets (which donot serve as an interlocking mechanism while the feather is in the normal resting position, but function with increasing braking action only when a neighboring ramus diverges to a precisely defined extent from its resting position) including the mechanism of their unhooking; devices for the avoidance of harmful hooking into contacted parts of other feathers; production of maximal stability by minimal air resistance, and of minute chambers (<0,00001 mm³) with still air for optimal heat isolation.Apart from this abstract, further information, accompanied by numerous figures, will be given in a later paper in Stuttgarter Beiträge zur Naturkunde.

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Veränderte Fassung eines am 11. 10. 1971 gehaltenen Vortrages auf der 83. Jahresversammlung der Deutschen Ornithologengesellschaft in Bonn.     

Methodological problems in evolutionary biology VIII. Biology and culture

Biology cannot accommodate all aspects of culture. Aspects of culture that a biological approach can take into account can be covered by the biological categories of phenotype and environment. There is no need to treat culture as a separate category. Attempts to elaborate biological explanations of cultural variation will meet with success only if biologists expand theories of development, and integrate them in evolutionary biology. The alternative — elaborating the idea of so-called cultural inheritance — makes little sense from a biological point of view.     

Cell cycle,morphogenesis, and ultrastructure in the pseudoheliozoan clathrulina elegans

Summary The development of the sessile pseudoheliozoan Clathrulina elegans Cienkowski (Desmothoraca, Rhizopoda, Protozoa) was followed by light and electron microscopy. The cell cycle comprises a heliozoan-like stage sitting in an organic perforated capsule, an amoeboid resp. biflagellate stage ensuring dispersal, and an encysted stage. Special reference is given to the morphogenetic processes during the flagellate-to-heliozoan transformation. The differentiation of the stalk is described in detail and a hypothesis of the morphogenesis of the capsule is presented. Granules in the filopods displaying a jerky movement, are identified as a hitherto underscribed type of haptocyst-like organelles. These kinetocysts are shown to be a useful marker for studying the turnover of the plasma membrane at organelle level. Evidence is presented that in the flagellate stage a surplus of plasma membrane is stored in numerous lacunae thus facilitating the rapid transformation into the heliozoan stage with its comparatively large surface area. The morphogenetic significance of microtubules during the formation of the stalk, the latter being an asymmetric extracellular secretion product, is elucidated. The role of microtubule nucleation sites, the precisely timed assembly and disassembly of microtubules and other factors which probably control development are discussed to some extent.The author thanks Mrs. G. Borges and Mrs. B. Gauss for excellent technical assistance. This investigation was supported by the Deutsche Forschungsgemeinschaft.     

DNA homology and adsorption specificity of Pseudomonas aeruginosa virulent bacteriophages

Summary The DNA homology and adsorption specificity of newly isolated virulent bacteriophages of P. aeruginosa have been studied. On the basis of this analysis all phages were divided into four groups: k, m, mnP78-like and mnF82-like bacteriophages. DNA's of k as well as m phages were shown to possess different restriction patterns although they have an extensive homology. Unlike other groups, k phages were characterized by the presence of T4 DNA ligase-repaired, single-chain breaks.Abbreviations kbp kilobase pairs - EM electron microscopy     

Microbial transformation of ginsenoside Rb1 by Rhizopus stolonifer and Curvularia lunata

Of 49 microbial strains screened for their capabilities to transform ginsenoside Rb₁, Rhizopus stolonifer and Curvularia lunata produced four key metabolites: 3-O-[-d-glucopyranosyl-(1,2)--d-glucopyranosyl]- 20-O-[-d-glucopyranosyl]-3,12, 20(S)-trihydroxydammar-24-ene (1), 3-O-[-d-glucopyranosyl-(1,2)--d- glucopyranosyl]-20-O-[-d-glucopyranosyl]-3,12, 20(S)-trihydroxydammar-24-ol (2), 3-O-[-d-gluco- pyranosyl-(1,2)--d-glucopyranosyl]-3, 12, 20(S)-trihydroxydammar-24-ene (3), and 3-O--d-glucopyranosyl-3, 12, 20(S)-trihydroxydammar-24-ene (4), identified by TOF-MS, ¹H- and ¹³C-NMR spectral data. Metabolites 1, 3 and 4 were from the incubation with R. stolonifer, and 1 and 2 from the incubation with C. lunata. Compound 2 was identified as a new compound.     

Adoptions expérimentales de larves entre des fourmis de genres différents (II):Myrmica laevinodis Nylander etAnergates atratulus Schenck

Résumé Nous avons fait élever des larves d'Anergates atratulus par des ouvrières deMyrmica laevinodis à 22°C. Pour y parvenir, il n'est pas utile de faire hivernerensemble les larves d'Anergates et les ouvrières deMyrmica. La présence de larves autochtones n'empêche pas lesMyrmica d'élever des larves d'Anergates. Dans toutes les expériences lesMyrmica ont été soumises au fridavant de recevoir des larves d'Anergates. Aucune reine deMyrmica n'a été utilisée dans ces expériences.Sur les 64 larves d'Anergates que nous avons utilisées, 38 se sont transformées en imagos. C'est au début de l'adoption et au moment des métamorphoses que périrent la plupart des 26Anergates perdus. Les femelles vécurent en général 2 ou 3 jours et cherchèrent très tôt à quitter le nid natal. Les mâles vécurent 2 à 3 semaines.

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Summary Larvae ofAnergates atratulus were experimentally reared by workers ofMyrmica laevinodis, at 22°C. An overwintering of both larvae ofAnergates and workers ofMyrmica is not necessary for the success of that experiment. The presence of larvae ofMyrmica does not keep theMyrmica from rearing larvae ofAnergates. The workers ofMyrmica have been cooled, in all the experiments, before receiving larvae ofAnergates. No queen ofMyrmica have been used in that experiments.38 of the 64 larvae ofAnergates used became imagos. Most of the 26 lostAnergates died at the beginning of the adoption and during the metamorphosis. The females lived generally 2 or 3 days and tried, very early, to leave their native nest. The males lived 2 or 3 weeks.

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Anergates atratulus Myrmica laevinodis, 22 . bmecme Anergates Myrmica. Myrmica Anergates. Myrmica Anergates. Myrmica . 64 Anergates , 38 . 26 Anergates 2 3 . 2 3 .

     

Plant regeneration from cytoplasmic hybrids of rice (Oryza sativa L.)

Summary We obtained cybrid plants by electrofusing -irradiated protoplasts of a cytoplasmic male-sterile line A-58 CMS (Oryza sativa L.) and iodoacetamide (IOA)-treated protoplasts of the fertile (normal) rice cultivar Fujiminori. The cybridity of the plants was confirmed by mitochondrial (mt) DNA restriction endonuclease, and plasmid-like DNA analyses, and by isozyme, cytological and morphological investigations. The chromosome number of the cybrid plants is 24.     

Action of rat liver Galβ1-4GlcNAc α(2-6)-sialyltransferase on Manβ1-4GlcNAcβ-OMe,GalNAcβ1-4GlcNAcβ-OMe,Glcβ1-4GlcNAcβ-OMe and GlcNAcβ1-4GlcNAcβ-OMe as synthetic substrates

Incubation of synthetic Man\1-4GlcNAc-OMe, GalNAc1-4GlcNAc-OMe, Glc1-4GlcNAc-OMe, and GlcNAc1-4GlcNac-OMe with CMP-Neu5Ac and rat liver Gal1-4GlcNAc (2-6)-sialyltransferase resulted in the formation of Neu5Ac2-6Man1-4GlcNAc-OMe, Neu5Ac2-6GalNAc1-4GlcNAc-OMe, Neu5Ac2-6Glc1-4GlcNAc-OMe and Neu5Ac2-6GlcNAc1-4GlcNAc-OMe, respectively. Under conditions which led to quantitative conversion of Gal1-4GlcNAc-OEt into Neu5Ac2-6Gal1-4GlcNAc-OEt, the aforementioned products were obtained in yields of 4%, 48%, 16% and 8%, respectively. HPLC on Partisil 10 SAX was used to isolate the various sialyltrisaccharides, and identification was carried out using 1- and 2-dimensional 500-MHz¹H-NMR spectroscopy.Abbreviations 2D 2-dimensional - CMP cytidine 5-monophosphate - CMP-Neu5Ac cytidine 5-monophospho--N-acetylneuraminic acid - COSY correlation spectroscopy - DQF double quantum filtered - HOHAHA homonuclear Hartmann-Hahn - MLEV composite pulse devised by M. Levitt - Neu5Ac N-acetylneuraminic acid - Neu5Ac2en 2-deoxy-2,3-didehydro-N-acetylneuraminic acid     

Evaluation of laboratory indexes of absorption of soil phosphorus by plants: II

Summary A comparison was made of several laboratory methods for estimating the yield of phosphorus in plants grown in greenhouse cultures on samples of 22 soils from different parts of the United States. The methods investigated and their rank in order of increasing precision of the estimates of the yield of phosphorus were as follows: extraction with lactic acid, calcium lactate buffer solution extraction with 2 per cent citric acid solution < extraction with 0.03N NH₄F, 0.025N HCl solution < percentage phosphorus saturation (found as follows: 100 × labile phosphorus by isotopic dilution/ phosphorus adsorption capacity according to Langmuir adsorption equation) labile phosphorus by isotopic dilution phosphorus extracted by water.Journal Paper No. J-3747 of the Iowa Agricultural and Home Economics Experiment Station, Ames, Iowa. Project No. 1183. Contribution from the Department of Agronomy.     

A framework for classifying subsistence production types of Nepal

This paper defines a four-tiered, hierarchical system for classifying subsistence production types in Nepal and uses the system to describe the subsistence communities of central Nepal. The system first divides Nepal into a grid of homogenous parts or cells, second, it divides, the cells into ridge-valley slope sequences, third, defines production types for the ridge-valley slope sequences, and, finally, distinguishes local variants of the production types. After describing these tiers, I apply these categories to the northern portions of the central and western development regions where I have done research. I then define how representatives of Nepal as a whole are the production types defined for the central and western development regions. Finally, the paper locates some completed studies within its framework.     

Ganglioside-cholera toxin interactions: a binding and lipid monolayer study

Summary On t.l.c. plates ¹²⁵I-cholera toxin binds to a disialoganglioside tentatively identified as GD_lb with about 10 times less capacity than to ganglioside GM₁. Binding of labeled toxin to both gangliosides was abolished in presence of excess amounts of unlabeled B subunit. Ganglioside extracts from human or pig intestinal mucosa showed toxin binding to gangliosides GM₁ and GD_1b. In ganglioside-containing lipid monolayers the penetration of the toxin was independent of the ganglioside binding capacity.Abbreviations GM₂ Gal-NAc14Gal(3-2NeuAc)14G1c1Cer - GM₁ Gal3Ga1-NAc14Gal(32NeuAc)14G1c11Cer - GD_1a NeuAc23Ga113Gal-NAc14Gal(32NeuAc)14G1c11Cer - GD1b Gall3Gal-NAcl4Gal(32NeuAc82NeuAc)14Glc11Cer - GT_1b NeuAc23Ga113Ga1-NAcal4Gal(3-2NeuAc82NeuAc)14G1c11Cer - dpPC 1,2-hexadecanoyl-sn-glycero-3-phosphocholine - dpPE 1,2-hexadecanoyl-sn-glycero-3-phosphoethanolamine     

The use of BRM-activated killer cells in adoptive immunotherapy: A pilot study with nine advanced cancer patients

Adoptive immunotherapy using MHC-nonrestricted-lymphocytes, peripheral blood T cells and NK cells was devised. Peripheral blood mononuclear cells (3 x 107) were selected by immobilization to anti-CD3 monoclonal antibody for 4 days and cultured for 2 weeks in the presence of IL-2. Thereafter they were reactivated by 500 U/ml of IFN- and 1000 U/ml of IL-2 for 1 hour. Enhancement of NK and LAK activities was confirmed. Peripheral blood T cells proliferated in response to immobilized anti-CD3 antibody (3% to 30%). Approximately 6 x 109 BRM-activated killer (BAK) cells composed of CD56+ T cells and CD56+ NK cells, were dispensed to cancer patients via intravenous drip infusion. Nine patients were treated with BAK cells every 2 weeks or every month on an outpatient basis. During the course of adoptive immunotherapy, the crossed affinity immunoelectrophoresis (CAIE) pattern of serum immunosuppressive acidic protein (IAP) was analysed. Both the production and glycosylation pattern of IAP is changed in response to tumor enlargement and may therefore act as a marker of the disease progression. During the course of BAK therapy, the glycosylation IAP pattern of 6 patients changed from tumor (T) to normal (N). In addition, the performance status of all patients was maintained at 90–100% of the Karnofsky scale and any side effects including fever were not observed during treatments with BAK cells. Moreover, the overall quality of life (QOL) of the patients, scored at the Face scale was favorable. In addition, blood levels of activated T cells producing IFN- were assayed as an indication marker of BAK therapy. The normal range of IFN- producing T cells comprised 6.9 ± 0.9% of peripheral blood mononuclear cells (PBMC), according to a single cell FACScan analyses of PBMCs derived from normal individuals. IFN- producing T cells of Patients No. 8 and 9, who received extensive chemotherapy before initiation of BAK therapy, comprised only 0.2% and 2% of PBMC, respectively. These patients died 3 and 6 months after beginning BAK therapy. Peripheral blood T cells of Patients Nos. 1–7 proliferated in response to immobilized anti-CD3 antibody and the frequency of IFN- producing T cells in PBMC preparation of these patients were over 3% before initiation of BAK therapy. Since our data show a positive correlation between survival time and initial T cell counts, a low frequency of these cells may contraindicate BAK therapy.     

Phosphate repression of cephamycin and clavulanic acid production by Streptomyces clavuligerus

Summary Production of cephamycin and clavulanic acid by Streptomyces clavuligerus is controlled by the phosphate concentration. Phosphate represses the biosynthesis of cephamycin synthetase, expandase and clavulanic acid synthetase. In the presence of 2 mM phosphate, the specific activities of expandase, cephamycin synthetase and clavulanic acid synthetase were higher than in the presence of 75 mM phosphate. The specific activity of cephamycin synthetase is maximal with an initial phosphate concentration of 10 mM, whereas the specific activity of expandase is maximal with 1 mM phosphate. A correlation between cephamycin synthetase specific activity and expandase specific activity was established at phosphate concentrations higher than 10 mM. This shows that the expandase is an important enzyme in the mechanism by which the phosphate concentration affects the biosynthesis of cephamycin.     

Sparse distribution of γ/δ T lymphocytes around human epithelial tumors predominantly infiltrated by primed/memory T cells

Summary Evidence from the mouse system has suggested that T lymphocytes accumulating in non-lymphoid tissue, in particular epithelia, may preferentially express the T cell receptor (TCR) . In this study, we characterize the T cell receptor or phenotype of lymphocytes infiltrating human tumors of epithelial origin using monoclonal antibodies (mAb) for immunohistology and flow cytometry on cells extracted by enzyme digestion. This report shows that the majority of CD3⁺ tumor-infiltrating lymphocytes are TCR ⁺ but a small percentage of TCR can be clearly defined scattered throughout the tumor tissue with apparently no microanatomical selection. So far there has been little evidence for an accumulation of activated T cells in human tumor tissues as defined by mAb against molecules appearing transiently during the acute phase of activation. Now mAb are available that can identify primed or memory T cells such as mAb UCHL-1 recognizing the CD45RO antigen. Here we show that CD3⁺ tumor-infiltrating lymphocytes have a statistically significant accumulation of primed T cells, as compared to the autologous peripheral blood lymphocytes, suggesting their immune stimulation by tumor cells.     

Purification and properties of a novel β-galactosidase or exo-(1 → 4)-β-d-galactanase from the cotyledons of germinated Lupinus angustifolius L. seeds

The main polysaccharide component of the thickened cell walls in the storage parenchyma of Lupinus angustifolius L. cotyledons is a linear (1 4)--linked d-galactan, which is mobilised after germination (L.A. Crawshaw and J.S.G Reid, 1984, Planta 160, 449–454). The isolation from the germinated cotyledons of a -d-galactosidase or exo-(1 4)--d-galactanase with a high specificity for the lupin galactan is described. The enzyme, purified using diethylaminoethyl-cellulose, carboxymethyl-cellulose and affinity chromatography on lactose-agarose, gave two bands (major 60 kDa, minor 45 kDa) on sodium dodecyl sulphate-gel electrophoresis, and two similar bands on isoelectric focusing (major, pI 7.0, minor pI 6.7, both apparently possessing enzyme activity). The minor component cross-reacted with an antiserum raised against, and affinity-purified on, the major band. Both components had a common N-terminal sequence. The minor component was probably a degradation product of the major one. The enzyme had limited -galactosidase action, catalysing the hydrolysis of p-nitrophenyl--d-galactopyranoside and (1 4)- and (1 6)--linked galactobioses. Lactose [-d-galactopyranosyl-(1 4)-d-glucose] was hydrolysed only very slowly and methyl--d-galactopyranoside not at all. Lupin galactan was hydrolysed rapidly and extensively to galactose, whereas other cell-wall polysaccharides (xyloglucan and arabinogalactan) with terminal non-reducing -d-galactopyranosyl residues were not substrates. A linear (1 4)--linked galactopentaose was hydrolysed efficiently to the tetraose plus galactose, but further sequential removals of galactose to give the tetraose and lower homologues occurred more slowly. Galactose, -galactonolactone and Cu⁺² were inhibitory. No endo--d-galactanase activity was detected in lupin cotyledonary extracts, whereas exo-galactanase activity varied pari passu with galactan mobilisation. Exo-galactanase protein was detected, by Western immunoblotting of cotyledon extracts, just before the activity could be assayed and then increased and decreased in step with the enzyme activity. The exo-galactanase is clearly a key enzyme in galactan mobilisation and may be the sole activity involved in depolymerising the dominant (1 4)--galactan component of the cell wall.Abbreviations CM carboxymethyl - DEAE diethylaminoethyl - SDS-PAGE sodium dodecyl sulphate-polyacrylamide gel electrophoresis - TLC thin-layer chromatography We wish to thank CNPq (Conselho Nacional de Desenvolvimento Científico e Tecnológico) for the award of a studentship to M.S. Buckeridge, and the Government of São Paulo State, Brazil for granting him leave of absence. We are grateful to Dr. Amanda Heyller (Unilever Research Laboratory, Colworth House, Bedford, UK) for N-terminal sequence determinations, to Dr. Stuart Wilson (Stirling) for preparing gelatin SDS-gels and to Cristina Fanutti (Stirling) for purifying the xyloglucan oligosaccharide.     

Effects of trichloroethylene,tetrachloroethylene and dichloromethane on enzymatic activities in soil

Summary Enzyme assays for -glucosidase, -acetylglucosaminidase, phosphatase, phosphodiesterase, and proteinase were made in soil samples incubated for two months after contamination with trichloroethylene, tetrachloroethylene, and dichloromethane. These volatile chlorinated hydrocarbons were added at doses of 10, 100, and 1000 g per 100 g dry soil, respectively. Almost no effect was observed in soil sample contaminated with 10 g of the chemicals when compared with control soil. When 100 g of the volatile chlorinated hydrocarbons was added, the activity of -glucosidase, -acetylglucosaminidase and, in part, also of proteinase, was reduced during the first 28 days of incubation but returned to the same or slightly higher level than in the control soil after 2 months. Trichloroethylene, tetrachloroethylene, and dichloromethane at a concentration of 1000 g per 100 g soil primarily inhibited activity of all enzymes under test. However, after two months, the enzymatic activities especially in soil samples contaminated with tetrachloroethylene and dichloromethane were found to be at the same or higher level than in the control soil.     

Regioselective deacetylation of peracetylated monosaccharide derivatives by polyethylene glycol-modified lipase for the oligosaccharide synthesis

Lipase modified with polyethylene glycol became soluble and active in organic solvents, and catalyzed regioselective deacetylation of peracetylated monosaccharide derivatives in 1,1,1-trichloroethane. The deacetylation occurred only at the positions of C-4 and C-6 of the glycopyranoside ring. Especially, peracetylated methyl -D-xylopyranoside and peracetylated L-serine--D-xylopyranoside were hydrolyzed only at the position of C-4. Subsequently, one of the resulting products, that is L-serine-2,3-di-O-acetyl--D-xylopyranoside, was coupled with galactose residue to obtain L-serine-4-O-(-D-galactopyranosyl)--D-xylopyranoside, a model compound of the carbohydrate-protein linkage region of proteoglycans.