A lectin from the liverwortMarchantia polymorpha L

Bryophytes have been screened for lectins. From the liverwortMarchantia polymorpha (Marchiantiales) a lectin could be purified to homogeneity using a combination of ultrafiltration, size exclusion chromatography and ion exchange chromatography. SDS polyacrylamide gel electrophoresis, size exclusion chromatography and electrospray mass spectroscopy showed that the lectin is a monomeric protein with a M_r of 16,134.64 ± 2.93.Marchantia polymorpha lectin agglutinates erythrocytes of different mammalia and exhibits carbohydrate specifity against complex carbohydrate structures. This is the first report of a lectin isolated from liverworts.This article forms Publication No. 71 of the Arbeitskreis Biologie und Chemie der Moose.     

Lectin receptors in snail proteogalactans. II.Archachatina marginata andAchatina achatina (preliminary report)

Summary The lectin receptor sites on the proteogalactans from the albumin glands of West African land snails,Archachatina marginata andAchatina achatina have been studied by precipitin reactions using the agar-gel double diffusion technique with various lectins. The proteogalactans from both snails have predominantly terminal -D-galactose structures; but they show characteristic differences in the topographical features at the surfaces of the carbohydrate structures presumed to be compatible with the combining site for these lectins.     

Effect of actinomycin D onTrypanosoma cruzi

Summary Viable metacyclic forms ofT. cruzi, Y strain, treated with an adequate dose of actinomycin D (50 g Act-D/ml/10⁷ parasites/ml for 72 h at 28° C) showed the following properties: 1) they lost their ability to replicate in culture medium, in blood and in tissues of normal mice and were no longer able to incorporate tritiated thymidine; 2) they could no penetrate into Vero cells and could not replicate inside normal macrophages; 3) they retained their immunogenicity and the ability to protect mice against a virulent infection; 4) they did not induce histological lesions as described in chronic experimental Chagas' disease.     

Partial purification and characterization of acid phosphatase from sporulated oocysts ofEimeria tenella

Summary Acid phosphatase ofEimeria tenella oocysts (Peak II) was purified 77-fold with a recovery of 26% using protamine sulfate precipitation, DEAE-cellulose chromatography and Sephadex G-200 gel filtration. This enzyme occurs in multiple forms as indicated by two peaks which can be separated by DEAE-cellulose chromatography and polyacrylamide gel electrophoresis. The partially purified enzyme has optimal activity at pH 4.5. With p-nitrophenyl phosphate the Km and V_max values for (Peak II) were 25 mM and 1.57 mol/min/mg protein, respectively. The enzyme (Peak II) ist strongly inhibited by Hg⁺⁺, Cu⁺⁺, iodoacetamide, fluoride and molybdate. Tartrate and other divalent metal ions have no effect on enzyme activity. The partially purified Peak II phosphatase is not a glycoprotein as it is not absorbed on concanavalin-A Sepharose and its treatment with bacterial neuraminidase does not alter its elution profile through DEAE cellulose.     

Oxidative stress in the moderately halophilic bacteriumDeleya halophila: Effect of NaCl concentration

The sensitivity ofDeleya halophila to oxidative stress caused by hydrogen peroxide (H₂O₂) was found to vary, depending on the NaCl concentration of the growth medium. Pretreatment of the bacteria at a low concentration of H₂O₂ (50 M) protected the cells against the lethal effects of higher levels (1–2 mM) of H₂O₂. Exposure ofD. halophila cells to 50 M H₂O₂ resulted in the induction of several proteins (hydrogen peroxide-inducible proteins, hips). However, the kinetics of induction, the extent of induction and the number of hips appear to be influenced by the salt concentration of the growth medium. Five of the hips exhibited apparent molecular masses identical to those of five heat shock proteins (hsps).     

Determination of diel periodicity of sex pheromone release in three species of Lepidoptera by ‘closed-loop-stripping’

Summary By means of closed-loop-stripping and subsequent GC analyses the diel periodicity of release of (Z)-11-hexadecenyl acetate, (E)-8-dodecenyl acetate, and (Z)-9-tetradecenyl acetate, the main constituents of the respective sex pheromone blends ofMamestra brassicae, Cryptophlebia leucotreta andSpodoptera sunia females, was determined.Pheromones, 64. For the 63rd contribution we have taken from: Szöcs, G., Toth, M., Bestmann, H. J., Vostrowsky, O., Heath, R. R., and Tumlinson, J. H., Z. Naturforsch.42c (1987) 165; Pheromones, 62: Bestmann et al.¹³.     

Detection of toxic metabolites in the hemolymph ofBeauveria bassiana infectedSpodoptera exigua larvae

Highly active metabolites have been detected in the hemolymph of the lepidopteranSpodoptera exigua infected with the mycopathogen,Beauveria bassiana. A combination of phenyl sepharose and CM ion exchange chromatography was utilized to extract the active metabolites from infected hemolymph samples. The active in vivo metabolites, having a molecular mass greater than 10 KDa, were thermolabile and were inactivated by proteinase K. These metabolites were characterized by their ability to disrupt metamorphosis, killing treated larvae at the wandering or pupal stage. Additionally, injection ofS. exigua larvae with active samples caused a reduction in the number of filopodial-producing hemocytes. The biological activities and biochemical properties suggest that novel compounds are produced duringB. bassiana mycosis.     

Isolation and partial characterization of a phytotoxic glycoprotein from culture filtrates ofRhynchosporium secalis

Summary Rhynchosporium secalis (Oud.) Davis produces a phytotoxic compound with a mol. wt of 275×10³ which is able to induce chlorotic symptoms in both susceptible and resistant barley leaves. Collectively, the data suggest that the toxin is a glycoprotein. Mild base treatment, by elimination, indicates that threonine and serine are involved in o-glycosidic linkages with the carbohydrate moiety. Sugar residues occur in the molecule in the ratio of mannose, rhammnose, galactose, glucosamine 13.6111.Acknowledgments. We acknowledge the assistance of Mr M. McNeil and P. Albersheim in doing the sugar analyses shown in this report. Financial support for this project was provided by a BARD grant 1-31-79, the French government, and the Montana Ag. Expt. Station.     

Role of <Emphasis Type="Italic">N</Emphasis>-linked polymannose oligosaccharides in targeting glycoproteins for endoplasmic reticulum-associated degradation

Misfolded or incompletely assembled multisubunit glycoproteins undergo endoplasmic reticulum-associated degradation (ERAD) regulated in large measure by their N-linked polymannose oligosaccharides. In this quality control system lectin interaction with Glc₃Man₉GlcNAc₂ glycans after trimming with endoplasmic reticulum (ER) -glucosidases and -mannosidases sorts out persistently unfolded glycoproteins for N-deglycosylation and proteolytic degradation. Monoglucosylated (Glc₁Man₉GlcNAc₂) glycoproteins take part in the calnexin/calreticulin glucosylation-deglucosylation cycle, while the Man₈GlcNAc₂ isomer B product of ER mannosidase I interacts with EDEM. Proteasomal degradation requires retrotranslocation into the cytosol through a Sec61 channel and deglycosylation by peptide: N-glycosidase (PNGase); in alternate models both PNGase and proteasomes may be either free in the cytosol or ER membrane-imbedded/attached. Numerous proteins appear to undergo nonproteasomal degradation in which deglycosylation and proteolysis take place in the ER lumen. The released free oligosaccharides (OS) are transported to the cytosol as OS-GlcNAc₂ along with similar components produced by the hydrolytic action of the oligosaccharyltransferase, where they together with OS from the proteasomal pathway are trimmed to Man₅GlcNAc₁ by the action of cytosolic endo--N-acetylglucosaminidase and -mannosidase before entering the lysosomes. Some misfolded glycoproteins can recycle between the ER, intermediate and Golgi compartments, where they are further processed before ERAD. Moreover, properly folded glycoproteins with mannose-trimmed glycans can be deglucosylated in the Golgi by endomannosidase, thereby releasing calreticulin and permitting formation of complex OS. A number of regulatory controls have been described, including the glucosidase-glucosyltransferase shuttle, which controls the level of Glc₃Man₉GlcNAc₂-P-P-Dol, and the unfolded protein response, which enhances synthesis of components of the quality control system.Received 26 January 2004; accepted 25 February 2004     

The in vivo role of α-mannosidase IIx and its role in processing of <Emphasis Type="Italic">N</Emphasis>-glycans in spermatogenesis

The surfaces of mammalian cells are covered by a variety of carbohydrates linked to proteins and lipids. N-glycans are commonly found carbohydrates in plasma membrane proteins. The structure and biosynthetic pathway of N-glycans have been analyzed extensively. However, functional analysis of cell surface N-glycans is just under way with recent studies of targeted disruption of genes involved in N-glycan synthesis. This review briefly introduces the potential role of processing -mannosidases in N-glycan biosynthesis and recent findings derived from the -mannosidase IIx (MX) gene knockout mouse, which shows male infertility. Thus, the MX gene knockout experiment unveiled a novel function of specific N-glycan, which is N-acetylglucosamine-terminated and fucosylated triantennary structure, in the adhesion between germ cells and Sertoli cells. Analysis of the MX gene knockout mouse is a good example of a multidisciplinary approach leading to a novel discovery in the emerging field of glycobiology.Received 29 November 2002; received after revision 30 December 2002; accepted 20 January 2003     

Molecular basis of homocysteine toxicity in humans

Because of its similarity to the protein amino acid methionine, homocysteine (Hcy) can enter the protein biosynthetic apparatus. However, Hcy cannot complete the protein biosynthetic pathway and is edited by the conversion to Hcy-thiolactone, a reaction catalyzed by methionyl-transfer RNA synthetase in all organisms investigated, including human. Nitrosylation converts Hcy into a methionine analogue, S-nitroso-Hcy, which can substitute for methionine in protein synthesis in biological systems, including cultured human endothelial cells. In humans, Hcy-thiolactone modifies proteins posttranslationally by forming adducts in which Hcy is linked by amide bonds to -amino group of protein lysine residues (Hcy-N-Lys-protein). Levels of Hcy bound by amide or peptide linkages (Hcy-N-protein) in human plasma proteins are directly related to plasma total Hcy levels. Hcy-N-hemoglobin and Hcy-N-albumin constitute a major pool of Hcy in human blood, larger than total Hcy pool. Hcy-thiolactone and Hcy-thiolactone-hydrolyzing enzyme, a product of the PON1 gene, are present in human plasma. Modification with Hcy-thiolactone leads to protein damage and induces immune response. Autoantibodies that specifically recognize the Hcy-N-Lys-epitope on Hcy-thiolactone-modified proteins occur in humans. The ability of Hcy to interfere with protein biosynthesis, which causes protein damage, induces cell death and elicits immune response, is likely to contribute to the pathology of human disease.Received 30 May 2003; received after revision 21 July 2003; accepted 15 August 2003     

Single amino acid substitutions insn-glycerol-3-phosphate dehydrogenase allozymes fromDrosophila virilis

Summary The amino acid sequence was compared among the three allelic variants (allozymes) ofsn-glycerol-3-phosphate dehydrogenase inD. virilis, which are detected by one-dimensional electrophoresis. The GPDH^f variant was different from the GPDH^m by only one substitution of 68-lysine for asparagine; GPDH^s differed from GPDH^m by substitution of 127-glycine for arginine. No electrophoretically silent substitutions were found in a total of 352 amino acid residues.     

Total synthesis of orellinine,a minor toxic component of the fungusCortinarius orellanus fries

Summary The 3,3,4,4-tetrahydroxy-2,2-bipyridyl-N-oxide has been synthesized by dealkylation of the corresponding tetramethyl derivative. The chemical properties of this compound are identical to those reported for the minor fungal toxin ofCortinarius orellanus, orellinine.This work is part of the Tesi di Laurea of Miss Cinzia Pierucci.     

Volatiles from clover headHypera meles (Fab.) and alfalfaH. postica (Gyllenhal) weevils: search for pheromones

Summary Volatile fractions of the clover head,Hypera meles (Fab.), and alfalfa,Hypera postica (Gyllenhal), weevils contained three of four boll weevil,Anthonomus grandis (Boh.), pheromone components, (Z)-3,3-dimethylcyclohexane ^1,-ethanol and (Z)- and (E)-3,3-dimethylcyclohexane-^1,-acetaldehyde. Also found were eight oxygenated monoterpenes, previously identified as precursors and intermediates of the boll weevil pheromones.     

Selective protection of cells against X-irradiation. Isoproterenol protects only those cells that possesβ-adrenoreceptors

Summary In mixed culture of Chinese hamster fibroblatst, clone 431, and transformed murine L fibroblasts, clone B-82, isoproterenol was found to protect only 431 cells against ionizing radiation. It was shown that 431 cells, in contrast to B-82 cells, possess-adrenoreceptors, and the radioprotective effect of isoproterenol can be realized only if this agent interacts with-adrenoreceptors coupled with the cAMP system. Since malignization often causes the disappearance of-adrenergic and other hormone receptors, the combined culturing and irradiation of the cells studied can be regarded as a model of the growth of malignant cells (B-82) among normal tissue cells (431 cells) under conditions of radiation therapy. A possibility of selective protection against radiation damage of normal tissue cells, with retention of the former radiosensitivity of tumor cells, is discussed.     

Adipokinetic hormones: cell and molecular biology

Adipokinetic hormones AKH I (pGlu-Leu-Asn-Phe-Thr-Pro-Asn-Trp-Gly-Thr-NH₂) and AKH II (pGlu-Leu-Asn-Phe-Ser-Trp-Gly-Thr-NH₂) are synthesized by neurosecretory cells (NSC) of the corpora cardiaca (CC) in the locust,Schistocerca gregaria. These NSC constitute a homogeneous peptide factory as each cell synthesizes both AKH I and AKH II. This homogeneity makes the CC an excellent system in which to study aspects of neuropeptide biosynthesis. This report summarizes recent findings on AKH inactivation and metabolism, as well as on AKH prohormone processing and biosynthesis.     

Identification of mucondialdehyde as a novel stress metabolite

In a survey of antifungal stress compounds induced by cupric chloride we found that leaves ofChenopodium album exuded a highly fungitoxic metabolite mucondialdehyde (trans-2,trans-4-hexadienedial), which was associated with 13-oxo-9,11-tridecadienoic acids (cis-9,trans-11 andtrans-9,trans-11 isomers) presumably resulting from -scission of 13-hydroperoxy-octadecadi(tri)enoic acid. The biogenesis and role as a general defensive agent in plants are briefly discussed.     

Inhibition of respiratory oxidation of reduced sulfur compounds by intact cells ofThiobacillus denitrificans (strain RT) grown on thiosulfate

Thiobacillus denitrificans strain RT, an obligate sulfur-oxidizing chemolithoautotroph, was grown under microaerophilic conditions with thiosulfate as the only energy source. The rates of tetrathionate, thiosulfate, elemental sulfur (S^o) and sulfite oxidation were measured respirometrically with an oxygen electrode, using actively growing cells. Cells oxidized thiosulfate, elemental sulfur (S^o) and sulfite, but not tetrathionate. The thiosulfateoxidizing activity and elemental sulfur-oxidizing activity (SOA) were almost totally inhibited by 50 M myxothiazol (>80%), an inhibitor of the quinone-cytochrome b region, and by 10 M of the uncoupler carbonyl cyanide m-chlorophenylhydrazone (CCCP) (>82%). Sulfite-oxidizing activity was also significantly inhibited (>60%) by 50 M myxothiazol and 10 M CCCP. 1 mM KCN totally inhibited (>90%) all respiratory activities. This study confirms that a sulfur-oxidizing activity appears during microaerophilic growth ofThiobacillus denitrificans strain RT on thiosulfate. The SOA is linked to the respiratory chain, probably releasing electrons in the quinone-cytochrome b region.To whom correspondence should be addressed. Submitted by R. Bachofen.     

Okadaxanthin,a novel C50-carotenoid from a bacterium,Pseudomonas sp. KK10206C associated with marine sponge,Halichondria okadai

To study the origins of biologically active substances in marine sponges, a carotenoid produced by a marine bacterium,Pseudomonas sp. strain number KK10206C, which was associated with a marine sponge,Halichondra okadai, was investigated. A visible absorption spectrum-guided isolation procedure led to the isolation of a novel C₅₀-carotenoid, okadaxanthin. Its structure, 2,2-bis(4-hydroxy-2-methyl-2-butenyl)-,-carotene, was elucidated mainly by spectroscopic methods. Okadaxanthin turned out to be a potent singlet oxygen quencher, approximately 10 times as strong as -tocopherol.     

(E)-3-tridecen-2-one,an antibiotic from the interdigital glands of black-tailed deerOdocoileus hemionus columbianus

(E)-3-tridecen-2-one, the major volatile component of interdigital gland extracts from the black-tailed deer,Odocoileus hemionus columbianus, inhibited the growth of gram-positive bacteria and fungi. The bacteria,Propionibacterium acnes, and the fungi,Trichophyton mentagrophytes had a minimum inhibitory concentration (MIC) of 12.5 g/mL and 25 g/mL, respectively.