Comparative effectiveness of lymphotoxin anticarcinogenic and tumor cell growth-inhibitory activities

Prevention of ultraviolet radiation- or chemical carcinogen-induced morphologic transformation and inhibition of tumor-producing transformed cell growth by lymphotoxin and by normal spleen leukocytes were quantitatively compared to define the antineoplastic activity spectra of these natural immune mediators. When Syrian golden hamster embryo cells seeded for colony formation in culture dishes were treated simultaneously with carcinogen and lymphotoxin, the number of morphologically transformed cell colonies was irreversibly reduced by 50% in the presence of 6 units of lymphotoxin/ml. Lymphotoxin inhibition of tumor cell growth, however, was reversible and 50% reduction in tumor cell growth in three transformed lines required 124, 330, and 477 units/ml. Thus, the anticarcinogenic activity of lymphotoxin can be 20-fold or more greater than its tumor growth-inhibitory activity. Similarly spleen leukocytes also were more effective as an anticarcinogen than as an inhibitor of tumor cell growth, consistent with previous observations that naturally occurring spleen leukocyte antineoplastic activity may result from lymphotoxin secretion.     

Lymphokine and phorbol (PMA) regulation of complement (C2) synthesis using U937

Human monocytes synthesize large amounts of the second complement component (C2) after incubation with a T-lymphocyte product called monocyte complement stimulator (MCS). The human monocyte-like cell line, U937, also synthesizes C2 and can be stimulated to increase this synthesis by lymphokine-rich culture supernates. Additionally, phorbol myristate acetate (PMA), an agent which induces maturational changes in other macrophage-like cell lines, also stimulates C2 synthesis by U937 cells. Lymphokine and PMA stimulation of C2 secretion by U937 are both reversibly inhibitable by cycloheximide. At optimal concentrations for stimulation of C2 synthesis, PMA inhibits [³H]thymidine incorporation by U937 indicating that increased C2 is not due to increased numbers of U937 cells.     

Glycerol catabolite regulation of d-cycloserine production in Streptomyces garyphalus

A fluorescent pigment was isolated from the culture fluid of Methanobacterium thermoautotrophicum strain H. This pigment was shown to be 7,8-didemethyl-8-hydroxy-5-deazariboflavin by various spectroscopic and chromatographic techniques. This compound was previously described as the FO acid hydrolysis fragment of coenzyme F₄₂₀. On the basis of the time of appearance of the pigment in the course of fermentation, it is suggested that this substance may be an over-produced biosynthetic precursor of F₄₂₀.     

Die Bedeutung der “feedback”-Hemmung der Threonin-Desaminase für die Synthese von Valin und Leucin

Zusammenfassung In Arthrobacter Stamm 23 führte die durch Mutation verursachte allosterische Unempfindlichkeit der Threonin-Desaminase zur dereprimierten Bildung der Enzyme im Isoleucin-Valin-Leucin-Biosyntheseweg. Derepression erfolgte auch, wenn Wildtypzellen in Gegenwart von -Ketobuttersäure inkubiert wurden. In beiden Fällen wurde Isoleucin überproduziert und ins Kulturmedium ausgeschieden. Wie aus Wachstumsexperimenten hervorging, verursachte der Überschuß an -Ketobuttersäure im Medium primär einen Valin- und Leucin-Mangel, der zu einer vorübergehenden Wachstumshemmung führte. Durch die dereprimierte Bildung der Enzyme im Isoleucin-Valin-Biosyntheseweg konnte die Wachstumshemmung überwunden werden.Der vorübergehende Hemmeffekt der -Ketobuttersäure ließ sich auf eine Konkurrenz der Substrate am ersten gemeinsamen Enzym im Isoleucin-Valin-Biosyntheseweg, der Acetohydroxysäure-Synthase, zurückführen. Wegen des niedrigen K _m-Wertes für -Ketobuttersäure wird dieses Substrat vom Enzym bevorzugt umgesetzt. Durch gaschromatographische Bestimmungen der Acetoin- und Acetyläthylcarbinol-Bildung in Enzymtests mit variierten Substrat-Konzentrationen konnte nachgewiesen werden, daß relativ geringe Konzentrationen an -Ketobuttersäure genügen, um die -Acetolacetat-Bildung vollständig zu unterdrücken. Diese Ergebnisse erklären die durch -Ketobuttersäure verursachte vorübergehende Wachstumshemmung bei Bakterien.

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The effect of the feedback inhibition of threonine deaminase on valine-leucine biosynthesis

In Arthrobacter strain 23 the allosteric insensitivity of threonin deaminase caused by mutation resulted in derepressed formation of the enzymes of the isoleucine-valine-leucine pathway. Derepression was also observed, when wild type cells were incubated in the presence of -oxobutyrate. In both cases isoleucine was overproduced and excreted. As growth experiments indicated the excess of -oxobutyrate in the medium caused endogenous valine and leucine deficiency and a transient inhibition of growth. Derepressed formation of the isoleucinevaline biosynthetic enzymes resulted in relief of growth inhibition.The transient inhibitory effect of -oxobutyrate has been traced back to substrate competition at the first enzyme common to the isoleucine and valine pathway, acetohydroxy acid synthase. Due to the low K _m of the enzyme for -oxobutyrate this substrate is preferentially converted. As proven by gaschromatographical measurements of acetoin and acetylethyl carbinol produced in enzyme (acetohydroxy acid synthase) assays with varied substrate concentrations, relatively low concentrations of -oxobutyrate are able to suppress the formation of -acetolactate completely. These results explain the transient inhibitory effect of -oxobutyrate on the growth of bacteria.

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Abkürzungen -KBS -Ketobuttersäure - FAD Flavin-adenin-dinucleotid - AHS Acetohydroxysäure - IPM Isopropylmalat - TPP Thiaminpyrophosphat     

Isolation of a crustaceann-acetyl-d-glucosamine-1-phosphate transferase and its activation by phospholipids

Summary Then-acetyl-d-glucosamine-1-phosphate: dolichol phosphate transferase fromArtemia has been partially purified and characterized. The enzyme is solubilized from crude microsomes using Triton X-100, and after detergent removal appears to be associated with phospholipids. Using dolichol phosphate and UDP-n-acetyl-d-glucosamine as substrates, the enzyme catalyzes the formation of dolichol-pyrophosphate-n-acetyl-d-glucosamine. the product identity has been verified by TLC and paper chromatography following mild acid hydrolysis. Under the incubation conditions used only one product is made, i.e., Dol-p-p-GlcNAc. The formation of product is linear with increasing amounts of added protein and with time of incubation. The enzyme requires magnesium ions for activity. Activity of the enzyme is stimulated 6-fold by exogenous dolichol phosphate and is also stimulated by added phospholipids, with optimal activity being obtained in the presence of mixtures of phosphatidylcholine and phosphatidylglycerol. Enzymatic activity is not increased upon addition of GDP-mannose or dolichol phosphate mannose. The enzyme is rapidly inactivated by exposure to several detergents, including Triton X-100 and deoxycholate. The activity is inhibited by tunicamycin and by the purified B₂ homologue of this antibiotic. Other antibiotic inhibitors such as diumycin and polyoxin D have little effect on the enzyme. Both the microsomal and solubilized enzyme preparations are inactivated by 70% upon treatment with phospholipase A₂; activity may be restored by addition of phospholipids. Following hydrophobic interaction chromatography on Phenyl Sepharose, gel filtration chromatography on Sepharose CL-4B indicated that the enzyme, purified 81-fold, contained phophatidylcholine and phosphatidyl-ethanolamine.Abbreviations SDS sodium dodecyl sulfate - PMSF phenyl methanesulfonylfluoride - HEPES 4-(2-hydroxyethyl)-1-piperazineethane sulfonic acid - GlcNAc N-acetyl-d-glucosamine - Dol-PP-GlcNAc dolichol pyrophosphate N-acetyl-d-glucosamine - Dol-P-man dolichol-phosphate-mannose - Dol-PP- (GlcNAc)₂ dolichol-pyrophosphate-di-N- acetylchitobiose - DMSO dimethylsulfoxide - C:M (2:1) chloroform:methanol (2:1) - C:M:W (10:10:3) chloroform:methanol:water (10:10:3) - GlcNAc-1-P N-acetyl-d-glucosamine-1-phosphate - Dol-P dolichol phosphate - EGTA ethylene glycol bis (b-aminoethyl ether)-NNNN tetraacetic acid     

The biosynthesis of the lantibiotics epidermin,gallidermin, Pep5 and epilancin K7

Lantibiotics are antibiotic peptides that contain the rare thioether amino acids lanthionine and/or methyllanthionine. Epidermin, Pep5 and epilancin K7 are produced by Staphylococcus epidermidis whereas gallidermin (6L-epidermin) was isolated from the closely related species Staphylococcus gallinarum. The biosynthesis of all four lantibiotics proceeds from structural genes which code for prepeptides that are enzymatically modified to give the mature peptides. The genes involved in biosynthesis, processing, export etc. are found in gene clusters adjacent to the structural genes and code for transporters, immunity functions, regulatory proteins and the modification enzymes LanB, LanC and LanD, which catalyze the biosynthesis of the rare amino acids. LanB and LanC are responsible for the dehydration of the serine and threonine residues to give dehydroalanine and dehydrobutyrine and subsequent addition of cysteine SH-groups to the dehydro amino acids which results in the thioether rings. EpiD, the only LanD enzyme known so far, catalyzes the oxidative decarboxylation of the C-terminal cysteine of epidermin which gives the C-terminal S-aminovinylcysteine after addition of a dehydroalanine residue.Abbreviations Dha 2,3-didehydroalanine - Dhb 2,3-didehydrobutyrine - Lan lanthionine - Melan methyllanthionine     

Biosynthesis of a trypanocide by Chromobacterium violaceum

Radio-isotope studies indicated not only that l-tryptophan can serve as carbon source for synthesis of the trypanocide, violacein by Chromobacterium violaceum (BB-78 strain) but also that isatin and indole 3-acetic acid are both important metabolic intermediates. Using 3-indolyl [2-¹⁴C] and [1-¹⁴C] acetic acid, it was found that the carboxylic carbon was not eliminated and that indole-3-acetic acid was incorporated intact into the pigment structure. N-Ethyl(5-hydroxy-indol-3-yl)-2-indolylethylamide is also an important metabolic intermediate in the violacein biosynthesis. This is the first report of a metabolic scheme for violacein synthesis which includes an intermediate other than l-tryptophan.     

Flora Palinologica Italiana,Sezione Aeropalinologica — Specie esotiche — S 231:Sequoia sempervirens (Lamb.) Endl. (Taxodiaceae)

Riassunto La scheda morfopalinologica diSequoia sempervirens (Lamb.) Endl. continua lo studio sulle Pinohyta ornamentali più frequenti in Torino. È esaminato polline fresco ed acetolizzato proveniente da due stazioni e la provenienza non determina variazioni significative dei parametri morfometrici.

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Summary The morphopalynological card ofSequoia sempervirens (Lamb.) Endl. is a new contribution to the comprehensive study on pollen of ornamental Pinophyta in Turin. Fresh and acetolysed grains from two stations are examined, and for all parameters no significative differences were observed in relation to the collection site.

Parole chiave: Sequoia sempervirens(Lamb.) Endl., Flora Palinologica Italina, Sezione Aeropalinologica.     

林带中阻力分布的理论与实验研究

推导了风向垂直于林带走向时林带内的阻力分布的解析式,比较了３种所面形状林带的阻力分布特点,并用风洞实验资料进行了验证,简析了在实际生产中的应用．     

Structure and biosynthesis of teichoic acids in the cell walls of Staphylococcus xylosus DSM 20266

The simultaneous occurrence of a N-acetylglucosaminyl poly(ribitolphosphate) (-GlcNAc) and a N-acetylglucosaminyl poly(glycerolphosphate) (-GlcNAc) in the cell walls of Staphylococcus xylosus DSM 20266 was demonstrated by different experimental lines:(1) Fractionation of extracted cell wall teichoic acid on DEAE-cellulose, (2) investigation of the composition of cell walls in the growth cycle, (3) in vitro biosynthesis using crude membranes as the source of enzyme.The polymerization of these polymers starts from CDP-ribitol and CDP-glycerol, respectively. In the presence of UDP-N-acetylglucosamine both polymers are substituted with N-acetylglucosamine at a level and with the identical anomeric configuration found in the native cell wall teichoic acids. The in vitro biosynthesis of poly(glycerolphosphate) was unique in that it was highly stimulated by UDP-N-acetylglucosamine and to a lower extent by other UDP-activated sugars. Kinetic studies have provided evidence that this stimulation is due to an increase of V _max while K _m is unchanged. Competition experiments have indicated that poly(ribitolphosphate) and poly(glycerolphosphate) were synthesized in the in vitro system in a close spatial relationship.Abbreviations ADP adenosine 5-diphospho - CDP cytidine 5-diphospho - GDP guanosine 5-diphospho - GalNAc N-acetyl-galactosamine - Glc glucose, glucosyl - GlcNAc N-acetyl-glucosamine - N acetylglucosaminyl - GlcUA glucuronic acid - Gro glycerol - Man mannose, mannosyl - Rit ribitol - SDS sodium dodecyl sulfate - UDP uridine 5-diphospho