11 S seed storage proteins fromHelianthus species (Compositae): Biochemical,size and charge heterogeneity

The seeds of 19 sunflower species were compared on the basis of their protein contents and the relative proportions of their protein fractions. The globulin content varied from 50% to about 70% and the albumin content from 18% to 35% according to the species. The level of intermediateMr polypeptides showed a great variability (9.6 to 24.3%). Comparative studies onMr polymorphism were carried out by means of sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) of non reduced and/or reduced samples using both mono- and bidimensional procedures. Polypeptide constituents of helianthinin were compared including both number and molecular size (cultivatedH. annuus was used as a standard). Studies focused on differences observed between the major two (Mr 38 000), (Mr 32 000) and (Mr 25 500), (21 000) polypeptides families constituting the main A, B, and C subunits. and polypeptides analyses permit to discriminate easilyH. petiolaris from the other species. Charge polymorphism was studied using isoelectric focusing (IEF) and IEF-PAGE in mono and bidimensional procedures in the presence or absence of 2-mercaptoethanol (2-ME). Only a specific ₄ polypeptide enables an easy discrimination betweenH. petiolaris and all the other species. Detailed nomenclature of the , and , polypeptides constituting the different helianthinin globulin subunits is given via the results of pI andMr analyses. Monodimensional IEF patterns of the more basic albumins (pI > 8.0) appear to provide a more valuable approach to identifying specific protein markers.     

Phylogenetic relationship of zeins and coixins as determined by immunological cross-reactivity and Southern blot analysis

Zeins from Zea mays L cv. Maya and coixins from Coix lacryma-jobi L. cv. Adlay were fractionated to obtain -, -, and -zein and -, -, and -coixin. The -coixins were composed of 4 polypeptide classes of 27 kDa (C1), 25 kDa (C2), 17 kDa (C4) and 15 kDa (C5) with solubility properties very similar to those of the 22 kDa and 19 kDa -zeins. Like the -zeins, the C1 and C2 -coixins corresponded to 80% of total Coix prolamins. The fraction corresponding to -coixin contained only one protein band of 22 kDa (C3). This coixin fraction has solubility properties similar to those of -zein and represents 15% of the total coixin. The -zein fraction was composed of a major 17 kDa protein band, while the -coixin fraction consisted of a mixture of - and -coixins.Polyclonal antibodies raised against C1 recognized C1 and C2 and cross-reacted strongly with the 22 kDa -zein, as did C4 and C5 antisera. The antiserum against -coixin showed strong cross-reaction with -zein. The homology between coixins and zeins was further investigated by using Southern hybridization analyses. The genomic DNA of maize and Coix were digested with several restriction enzymes and probed with cDNA clones representing 19 and 22 kDa -zeins as well as the 28 and 16 kDa -zeins. The Coix genome showed complex cross-hybridization sequences with the 22 kDa -zein cDNA, while no cross-hybridization was observed with the 19 kDa cDNA clone. The cDNA clone representing the 28 kDa -zein cross-hybridized with only one band of Coix genomic DNA, in contrast to the three bands observed in maize. This same Coix sequence also cross-hybridized with the cDNA clone representing the 16 kDa -zein. The relevance of these findings are discussed in the context of the origin of zein and coixin genes.     

Analysis of the effect of rye chromosomes 4R, 6R and telomeric heterochromatin on 7R on homologous chromosome pairing in pentaploid hybrids (AABBR,AABBD)

Summary Meiotic pairing in Triticum turgidum cv. Ma (4x) with a mean chiasmata frequency of 27.16 per cell was compared with chiasmata frequencies in its hybrids with several triticale strains, Chinese Spring wheat and its addition lines for Imperial rye chromosomes 4R and 6R. In hybrids between Ma and x Triticosecale cv. Rosner the chiasmata frequency was marginally reduced by an average of 1.25%, by 8.8% in hybrids with x Triticosecale cv. DRIRA HH and by 6.7% with DRIRA EE (lacking 90% telomeric heterochromatin from chromosome arm 7RL). In pentaploid hybrids between Ma and T. aestivum cv. Chinese Spring the reduction was an average of 10.30%, while addition lines with rye chromosome 6R reduced chiasmata frequencies by an average of 7.4% and rye addition line for 4R showed the greatest depression in chiasmata frequency in hybrids by a 25.04% reduction. An interchange difference involving long chromosome segments was observed between Ma and Rosner.Contribution No. 819 Ottawa Research Station     

Conformational changes of α-lactalbumin and its fragment, Phe31-Ile59, induced by sodium dodecyl sulfate

Conformational changes of bovine -lactalbumin in sodium dodecyl sulfate (SDS) solution were studied with the circular dichroism (CD) method using a dilute phosphate buffer ofpH 7.0 and ionic strength 0.014. The proportions of -helix and -structure in -lactalbumin were 34% and 12%, respectively, in the absence of SDS. In the SDS solution, the helicity increased to 44%, while the -structure disappeared. In order to verify the structural change from -structure to -helix, the moiety, assuming the -structure in the -lactalbumin, was isolated by a chymotryptic digestion. The structure of this -lactalbumin fragment, Phe³¹-Ile⁵⁹, was almost disordered. However, the fragment adopted a considerable amount of -helical structure in the SDS solution. On the other hand, the tertiary structure of -lactalbumin, detected by changes of CD in the near-ultraviolet region, began to be disrupted before the secondary structural change in the surfactant solution. Dodecyl sulfate ions of 80 mol were cooperatively bound to -lactalbumin. Although the removal of the bound dodecyl sulfate ions was tried by the dialysis against the phosphate buffer for 5 days, 4 mol dodecyl sulfates remained per mole of the protein. The remaining amount agreed with the number of stoichiometric binding site, determined by the Scatchard plot, indicating that the stoichiometric binding was so tight.     

Rapid plant regeneration of pea using thidiazuron

A simple and rapid pea regeneration procedure was developed. An average of up to 20 shoots formed from hypocotyl explants of cvs. Sugar Ann and Patriot cultured on Murashige and Skoog basal medium supplemented with 0.5 or 1.0 M thidiazuron (N-phenyl-N-1,2,3-thiadiazol-5-ylurea). Hypocotyls of Puget and Sugar Daddy did not respond. Regenerated shoots rooted rapidly when cultured on Murashige and Skoog basal medium containing either 2.0 M -naphthaleneacetic acid or 1.0–2.0 M indole-3-butyric acid. Seeds were harvested from regenerated plants after only 9–11 weeks.Abbreviations BAP 6-benzyladenine - 2,4-d 2,4-dichlorophenoxy acetic acid - GA₃ gibberellic acid - IBA indole-3-butyric acid - MS Murashige and Skoog (1962) medium - NAA -naphthaleneacetic acid - TDZ thidiazuron (N-phenyl-N-1,2,3-thiadiazol-5-ylurea)     

1H and13C-NMR analysis of four pentasaccharides from urine of blood-group A and B,Leb adults. Confirmation of the structure of two new oligosaccharides: Fuc(α1–2)[GalNAc(α1–3)]Gal(β1–3)[Fuc(α1–4)]Glc and Fuc(α1–2)[Gal(α1–3)]Gal(β1–3)[Fuc(α1–4)]Glc

The major pentasaccharides Fuc(1-2)[GalNAc(1-3)]Gal(1-4)[Fuc(1-3)]Glc and Fuc(1-2) [Gal(1-3)]Gal(1-4)[Fuc(1-3)]Glc, which are normally present in the urine of bloodgroup A Le^b and B Le^b healthy subjects, were each found to be contaminated by a minor component when analysed by¹H-NMR. The determination of these structures, Fuc(1-2) [GalNAc(1-3)]Gal(1-3)[Fuc(1-4)]Glc and Fuc(1-2) [Gal(1-3)]Gal(1-3)[Fuc(1-4)]Glc, was based on the results of methylation analysis and¹H/¹³C-NMR spectroscopy.Abbreviations HPLC high performance liquid chromatography - GLC gas liquid chromatography - NMR nuclear magnetic resonance - COSY correlation spectroscopy - Gal d-galactopyranose - GalNAc 2-acetamido-2-deoxy-d-galactopyranose - Glc d-glucopyranose - Fuc l-fucopyranose - LNDFH I lacto-N-difucohexaose I (Le^b determinant     

α-and β-Glycosidases in maize roots

Four glycosidases were analyzed in 10 mm apical segments prepared from growing roots (15 mm) of Zea mays L. The pH optima were found to be 5.8 for -glucosidase, 4.4 for -galactosidase, 6.4 for -glucosidase and 6.0 for -galactosidase. The -glucosidase showed 4-fold higher activity than the -galactosidase. The distribution of the -glucosidase activity was signifcantly different from that of the -galactosidase, -glucosidase and -galactosidase.Abbreviations -Glu -glucosidase - -Gal -galactosidase - -Glu -glucosidase - -Gal -galactosidase     

Sterols of male and female compound inflorescences of Zea mays L.

A comparison has been made between the sterols of male and female inflorescences and of pollen from Zea mays. The female inflorescence was shown to contain cholesterol, 24-methylcholesterol, 24-ethyl-5,22-cholestadien-3-ol, 24-ethylcholesterol and (28Z)-24-ethylidenecholesterol. Themale inflorescence contained the same five compounds together with 24-methylenecholesterol. Pollen contained 24-methylenecholesterol as its main sterol together with lesser amounts of cholesterol, 24-ethylcholesterol, (28Z)-24-ethylidenecholesterol, 24-methylene-5-cholest-7-en-3-ol and 4-methyl-24-methylene-5-cholest-7-en-3-ol.     

Tissue culture ofKarwinskia humboldtiana—a plant producing toxins with antitumoural effects

Isolated embryos ofKarwinskia humboldtiana were cultured in vitro. The growth of embryos and development to plantlets on woody plant medium supplemented with indole-3-acetic acid 6.10^-2 mol l^–1, gibberellic acid (GA₃) 3.10^-2 mol l^–1, and 6-benzylaminopurine (BA) 2 mol l^–1 was obtained. Multiplication of shoots and rooting of excised shoots has been achieved. Callus formation on modified Murashige-Skoog medium supplemented with 1-naphthaleneacetic acid 10 mol l^–1, GA₃ 14 mol l^–1, and kinetin 5 mol l^–1 on hypocotyls, or on root cultures on medium supplemented with 2.4-dichlorophenoxyacetic acid 10 mol l^–1 and BA 10 mol l^–1 was induced.Abbreviations BA 6-benzylaminopurine - 2,4-d 2,4-dichlorophenoxyacetic acid - GA₃ gibberellic acid - IAA indole-3-acetic acid - NAA 1-naphthaleneacetic acid - TEM transmission electron microscopy     

Evaluation of structural and physiological plant characteristics in relation to the distribution of cadmium in maize inbred lines

To establish the structural and physiological characteristics related to the genotypic variation in Cd distribution between maize inbred lines (shoot Cd excluders and non-shoot Cd excluders), shoot and root morphological parameters were studied on plants grown in nutrient solution. Furthermore, the xylem sap composition and the desorbability of Cd from roots of these inbreds have been compared. No relationship between the morphological characteristics of either shoot (specific leaf area and leaf area ratio) or root (specific root length, specific surface area, and average diameter) and Cd distribution could be assessed. Cadmium concentrations in the xylem exudates from non-shoot Cd excluders were higher than those from shoot Cd excluders, but not related to citrate and malate concentrations. The absolute and relative amounts of Cd desorbed from roots of shoot Cd excluders were about twice as high compared to those of the non-shoot Cd excluders, especially at the lowest Cd concentration in solution. The absence of a relationship between shoot or root morphological parameters and Cd partitioning and the differences between both groups in the amounts of Cd desorbed, even at similar root Cd concentrations, indicate that the differential Cd distribution between shoot Cd excluders and non-shoot Cd excluders may be related to the observed differences in root Cd concentration, desorption characteristics and binding capacity of Cd inside and/or outside the root and its distribution within the roots.     

Abscisic acid metabolism —vacuolar/extravacuolar distribution of metabolites

The biotransformation of abscisic acid (ABA) was studied in cell suspension cultures of Lycopersicon esculentum. The ABA was converted by the cells to phaseic acid, nigellic acid, dihydrophaseic acid, abscisic acid--D-glucopyranosyl ester (ABA-Glc) and other ABA and phaseic acid conjugates. Investigation of their cellular distribution showed that the conjugated forms were located only in the vacuoles whereas ABA and its acidic metabolites were found mainly in the extravacuolar fractions. Our results, together with a number of studies on the increase of ABA-Glc as a response to stress, allow us to propose that ABA-Glc is irreversibly compartmented in the vacuoles of plant cells.Abbreviations ABA abscisic acid - ABA-Glc -D-glucopyranosyl ester of ABA - DPA 4-dihydrophaseic acid; nigellic acid=3-methyl-5-(1-hydroxy-2-hydroxymethyl-6-dimethyl-4-oxo-cyclohex-2-enyl)-penta-2Z, 4E-dienoic acid - PA phaseic acid     

Cell multiplication and ultrastructure ofMicrasterias denticulata (Desmidiaceae) grown under salt stress

Cells ofMicrasterias denticulata Bréb. were kept in nutrient solution of high osmolality (salt stress) for four weeks. In a special cell multiplication test it was established that cell division is gradually inhibited at increasing salt concentrations and totally arrested at the highest concentration (26 mosm/kg). Recovery studies proved that even cells from the highest concentration range start dividing immediately after being placed in aqua bidest. thus indicating the full reversibility of the inhibiting effect. — Cells of the highest concentration range show marked ultrastructural changes. Besides an enormous accumulation of starch and oil bodies and a condensed appearance of the ground plasma, a reduction of mitochondria, ER and the Golgi-system is found. The most striking effect occurs on the vacuolar system which appears extremely reduced and condensed. The cell wall is thickened by the formation of an additional cell wall layer with a spongy electron microscopical appearance. Through the cell wall many droplets of a probably fat-like substance are excreted. — In summary, salt stress induces growth-inhibited akinete cells in the sense ofFritsch; these can be reactivated by decreasing the salt concentration. The salt-induced akinete state seems to be an ecological adaption to unfavourable conditions rather than a degeneration of the cells.Dedicated to Prof. DrLothar Geitler on the occasion of his 90th birthday.23. 12. 1988     

The catalysis of nucleotide polymerization by compounds of divalent lead

Summary Soluble lead salts and a number of lead-containing minerals catalyze the formation of oligonucleotides from nucleoside 5-phosphorimidazolides. The effectiveness of lead compounds correlates strongly with their solubility. Under optimal conditions we were able to obtain 18% of pentamer and higher oligomers from ImpA. Reactions involving ImpU gave smaller yields.Abbreviations A adenosine - U uridine - Im imidazole - MeIm 1-methyl-imidazole - EDTA ethylenediaminetetraacetic acid - pA adenosine 5-phosphate - pU uridine 5-phosphate - Ap adenosine cyclic 2:3-phosphate - ATP adenosine 5-triphosphate - AppA P₁,P₂-diadenosine 5-diphosphate - pNp (N = A,U) nucleotide 2(3), 5-diphosphate - ImpA adenosine 5-phosphoreimidazolide - ImpU uridine 5-phosphorimidazolide - A ²pA adenylyl-[25]-adenosine - A ³pA adenylyl-[35]-adenosine - pA ²pA 5-phospho-adenylyl-[25]-adenosine - pA ³pA 5-phospho-adenylyl-[35]-adenosine - pUpU 5-phospho-uridylyl-uridine - pApU 5-phospho-adenylyl-uridine - pUpA 5-phospho-uridylyladenine - (pA)n (n, 2,3,4,) oligoadenylates with 5 terminal phosphate - ImpApA 5-phosphorimidazolide of adenylyl adenosine - (pA) ₅₊ pentamer and higher oligoadenylates with 5 terminal phosphate - (Ap)nA (n = 2,3,4) oligoadenylates without terminal phosphates In the following we do not specify the nature of the internucleotide linkageIn the following we do not specify the nature of the internucleotide linkage     

A general strategy for the isolation of carbohydrate chains fromN-,O-glycoproteins and its application to human chorionic gonadotrophin

For the structural analysis of the carbohydrate chains ofN-,O-glycoproteins a straightforward strategy was developed based on the cleavage of theN-linked chains with immobilized peptide-N ⁴-(N-acetyl--glucosaminyl) asparagine amidase-F (PN-Gase-F) fromFlavobacterium meningosepticum, followed by alkaline borohydride treatment of the remainingO-glycoprotein material. This methodology was applied to the isolation of the Asn- and Ser-linked carbohydrate chains of human chorionic gonadotrophin. The structures of the isolated oligosaccharides were verified by 500-MHz¹H-NMR spectroscopy. The Asn-linked sugar chains were shown to be: NeuAc2-3Gal1-4GlcNAc1-2Man1-6[NeuAc2-3Gal1-4GlcNAc1-2Man1-3]Man 1-4GlcNAc1-4[Fuc1-6]_0-1GlcNAc and Man1-6[NeuAc2-3Gal1-4GlcNAc1-2Man 1-3]Man1-4GlcNAc1-4GlcNAc. Also some minor constituents occurred. The structures of the Ser-linked oligosaccharides were established in the form of their oligosaccharide-alditols as: NeuAc2-3Gal1-3[NeuAc2-6]GalNAc, NeuAc2-3Gal 1-3GalNAc and NeuAc2-3Gal1-3[NeuAc2-3Gal1-4GlcNAc1-6]GalNAc.Abbreviations hCG human chorionic gonadotrophin - hCG- -subunit - hCG- -subunit - ElA enzyme immunoassay - PNGase-F peptide-N ⁴-(N-acetyl--glucosaminyl)asparagine amidase-F (EC 3.5.1.52) - SDS sodium dodecyl sulphate - GalNAc N-acetylgalactosamine - GlcNAc N-acetylglucosamine - NeuAc N-acetylneuraminic acid - Man mannose - Gal galactose - Fuc fucose     

Fromβ-lactams toα- andβ-amino acid derived peptides

Summary The potential of-lactams as intermediates for the access to- and-amino acid-derived peptides is shortly reviewed, with major focus on the technologies developed in our group. The two general strategies lie, on one side, in the oxidative ring expansion of 3-hydroxy-lactams toN-carboxy-amino acid anhydrides or Leuch's anhydrides and subsequent coupling with-amino acid esters and, on the other side, in the nucleophilic ring opening ofN-Boc--lactams. Both approaches have been successfully applied to the synthesis of,-diamino acid,-amino--hydroxy acid, polyhydroxylated-amino acid,,-disubstituted-amino acid,-amino acid,-amino--hydroxy acid and,-disubstituted-amino acid derived peptides. Because of the mild reaction conditions needed for the above transformations and the highly stereoselective procedures employed for the construction of the starting-lactam ring, the whole process allows the production of optically pure final products.     

A model of associative memory in the brain

A model of associative memory for time varying spatial patterns is proposed and simulated on a digital computer. This is a network composed of many neuron-like elements, and shows an ability for associative memory similar to that of the brain.Suppose a number of sequences of spatial patterns are presented to this network, for example, 12345, ABC, and so on. Then, these patterns are memorized in the network. After that, if any part of one of these sequences, say 23, is presented to the circuit, the rest of the sequence, 45, is recalled following to it. It resembles to such a situation — if we hear a part of a melody which we have memorized in the past, the rest of the melody is recalled even after it is stopped half-way. Although the recalled patterns are not always 100% correct, they are not completely destroyed even if the presented patterns are imperfect.     

(Z)-9-tetradecenal: a potent inhibitor of pheromone-mediated communication in the oriental tobacco budworm moth,Helicoverpa assulta

The behavioural significance of (Z)-9-tetradecenal to male H. assulta was tested by comparing the number of moths attracted to lures containing a standard synthetic female sex pheromone with lures in which (Z)-9-tetradecenal was also added. The standard pheromone mixture used contained 1000 g (Z)-9-hexadecenal, 50 g (Z)-11-hexadecenal, 300 g (Z)-9-hexadecenyl acetate and 15 g (Z)-11-hexadecenyl acetate impregnated on rubber septa. Addition of (Z)-9-tetradecenal to the standard pheromone was shown to significantly reduce the caught of male H. assulta when added in amounts greater than 10 g or 1% of the major pheromone component in both field and net-house experiments. The reduction in catch was found to be dependent on the quantity of (Z)-9-tetradecenal added to the standard pheromone. The implications of these results on conspecific and inter-specific pheromone-mediated communication in H. assulta and related sympatric heliothine species is discussed.Abbreviations Z9-16:AL (Z)-9-hexadecenal - Z11-16:AL (Z)-11-hexadecenal - Z9-16:AC (Z)-9-hexadecenyl acetate - Z11-16:AC (Z)-11-hexadecenyl acetate - Z9-14:AL (Z)-9-tetradecenal - Z9-16:OH (Z)-9-hexadecen-1-ol - Z11-16:OH (Z)-11-hexadecen-1-ol - RH relative humidity     

Structure of a new nonasaccharide isolated from human milk: VI2Fuc,V4Fuc,III3Fuc-p-lacto-N-hexaose

The structure of a new nonasaccharide isolated from human milk has been investigated. By using methylation analysis, FAB-MS and¹H-and¹³C-NMR spectroscopy as basic methods of structural investigation, this oligosaccharide was identified as VI²--Fuc,V⁴-Fuc,III³--Fuc-p-lacto-n-hexaose: Fuc1-2Gal1-3[Fuc1-4]GlcNAc1-3Gal1-4[Fuc1-3]GlcNAc1-3Gal1-4Glc.Abbreviations COSY correlation spectroscope - DP degree of polymerisation - FAB-MS fast atom bombardment-mass spectrometry - HPLC high performance liquid chromatography - NMR nuclear magnetic resonance - GLC gas-liquid chromatography     

Nucellar callus of ‘Femminello’ lemon,selected for tolerance toPhoma tracheiphila toxin,shows enhanced release of chitinase and glucanase into the culture medium

Phoma tracheiphila is the causative agent of the disease mal secco. Citrus cultivars differ substantially in respect to their sensitivity to the pathogenP. tracheiphila and its toxin. Some cultivars (e.g., Femminello lemon) are inherently sensitive while others (e.g., Tarocco orange) are tolerant. Cell lines derived from nucellar tissue of Femminello, Tarocco and a cell line selected for tolerance to the fungal toxin (Femminello-S) were used to study host-pathogen interaction. Our results showed that calli or conditioned media of Tarocco and Femminello-S inhibited the size of co-cultivated fungal colonies when compared to Femminello. In addition, conditioned medium of Tarocco as well as FemminelloS, but not Femminello, promoted bursting of hyphal tips. A ten-fold increase in chitinase and glucanase enzymatic activity, as evaluated by radiometric assay and laminarin hydrolysis respectively, was detected in Femminello-S extracellular extracts as compared to Femminello. An increase in chitinase was also shown by immunoblot analysis. Our findings suggest a positive correlation between the presence of chitinase and glucanase in the conditioned media of the cultured cells and the tolerance of those cells toP. tracheiphila toxin.     

Circadian changes in protein-synthesis rate and protein phosphorylation in cell-free extracts of Gonyaulax polyedra

The polysomal pattern of the dinoflagellate Gonyaulax polyedra, cultured under constant conditions, demonstrates a circadian rhythm. The relative amount of polysomes increases during the phase corresponding to the previous night period (=subjective night phase) when the rate of protein synthesis reaches its maximum (Cornelius et al., 1985, Planta 160, 365–370). Cell-free extracts were isolated at different circadian phase. The rate of protein synthesis in the extracts changed rhythmically in the same manner as the rate of protein synthesis in vivo. Substances in the postribosomal supernatants influenced the protein-synthesis rate of the cell-free system, depending on the phase when they were isolated: night factors stimulated protein synthesis in day extracts whereas day factors inhibited protein synthesis in night extracts. These effects were abolished by heating the postribosomal supernatant. In-vitro phosphorylation in parallel probes showed changes in the pattern of phosphorylated proteins. Phosphorylation of one of the proteins (95 kDa) was decreased after addition of night factor(s) and increased after addition of day factor(s). Cyclic-AMP enhanced the rates of protein synthesis and phosphorylation in the day extracts.Abbreviations cAMP cyclic-AMP - CT circadian time - D (N) subjective day (night)phase