Immunohistochemical localization of β-glucosidases in lignin and isoflavone metabolism in Cicer arietinum L. seedlings

Coniferin specific- and isoflavone 7-glucoside specific -glucosidases have been localized in stem and root sections of chick pea (Cicer arietinum L.) seedlings by the indirect immunofluorometrical method. The coniferin specific -glucosidase has been found in the cell walls of the tracheary elements and of the endo-, epi-, and exodermis. All these tissues are known to contain either lignin or polymers, like suberin and cutin, which consist partially of phenylpropanoid elements. The localization of this -glucosidase is therefore in agreement with its postulated relationship to the phenylpropanoid metabolism. The isoflavone 7-glucoside specific -glucosidase, on the other hand, is predominantly located in the parenchymatic cortex cells, and obviously in the cytoplasm. These cells are known to contain the isoflavone formononetin, which has been shown to undergo turnover in chick pea seedlings. We therefore have good reason to assume that this -glucosidase is involved in the metabolism of the 7-glucoside of this isoflavone.Abbreviations SDS sodium dodecylsulfate - PBS physiological phosphate saline The results are part of the thesis of Gerd Burmeister, 1980, University of Münster     

Microbial metabolism of aromatic nitriles. Enzymology of C–N cleavage by Nocardia sp. (Rhodochrous group) N.C.I.B. 11216

1. An organism utilizing benzonitrile as sole carbon and nitrogen source was isolated by the enrichment-culture technique and identified as a Nocardia sp. of the rhodochrous group. 2. Respiration studies indicate that nitrile degradation proceeds through benzoic acid and catechol. 3. Cell-free extracts of benzonitrile-grown cells contain an enzyme that catalyses the conversion of benzonitrile directly into benzoic acid without intermediate formation of benzamide. 4. This nitrilase enzyme was purified by DEAE-cellulose chromatography and gel filtration on Sephadex G-100 in the presence and absence of substrate. The purity of the enzyme was confirmed by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis and isoelectric focusing on polyacrylamide gel. 5. The enzyme shows a time-dependent substrate-activation process in which the substrate catalyses the association of inactive subunits of mol.wt. 45000 to form the polymeric 12-unit active enzyme of mol.wt. 560000. The time required for complete association is highly dependent on the concentration of the enzyme, temperature and pH. 6. The associated enzyme has a pH optimum of 8.0 and K(m) with benzonitrile as substrate of 4mm. The activation energy of the reaction as deduced from the Arrhenius plot is 51.8kJ/mol. 7. Enzyme activity is inhibited by thiol-specific reagents and several metal ions. 8. Studies with different substrates indicate that the nitrilase is specific for nitrile groups directly attached to the benzene ring. Various substituents in the ring are compatible with activity, though ortho-substitution, except by fluorine, renders the nitrile invulnerable to attack. 9. The environmental implications of these findings and the possible significance of the enzyme in the regulation of metabolism are discussed.     

Substrate specificity of an α-amino acid ester hydrolase produced by Acetobacter turbidans A.T.C.C. 9325

A partially purified preparation of an alpha-amino acid ester hydrolase was obtained from Acetobacter turbidans A.T.C.C. 9325, which catalyses synthesis of 7-(d-alpha-amino-alpha-phenylacetamido)-3-cephem-3-methyl-4- carboxylic acid (cephalexin) from methyl d-alpha-aminophenylacetate and 7-amino-3-deacetoxycephalosporanic acid. The enzyme preparation catalysed both cephalosprin synthesis from 7-amino-3-deacetoxycephalosporanic acid and suitable amino acid esters (e.g. methyl d-alpha-aminophenylacetate, l-cysteine methyl ester, glycine ethyl ester, d-alanine methyl ester, methyl dl-alpha-aminoiso-butyrate, l-serine methyl ester, d-leucine methyl ester, l-methionine methyl ester) and the hydrolysis of such esters. The substrate specificity of the enzyme preparation for the hydrolysis closely paralleled the acyl-donor specificity for cephalosporin synthesis, even to the reaction rates. Only alpha-amino acid derivatives could act as acyl donors. The hydrogen atom on the alpha-carbon atom was not always required by acyl donors. The hydrolysis rate was markedly diminished by adding 7-amino-3-deacetoxycephalosporanic acid to reaction mixtures, but no effect on the total reaction rate (the hydrolysis rate plus synthesis rate) was observed with various concentrations of 7-amino-3-deacetoxycephalosporanic acid. Both the hydrolytic and the synthetic activities of the enzyme preparation were inhibited by high concentrations of some acyl donors (e.g. methyl d-alpha-aminophenylacetate, ethyl d-alpha-aminophenylacetate). The enzyme preparation hydrolysed alpha-amino acid esters much more easily than alpha-amino acid derivatives with an acid-amide bond.     

Unexpected wide substrate specificity of C. perfringens α-toxin phospholipase C

Clostridium perfringens phospholipase C (CpPLC), also called α-toxin, is the main virulence factor for gas gangrene in humans. The lipase activity serves the bacterium to generate lipid signals in the host eukaryotic cell, and ultimately to degrade the host cell membranes. Several previous reports indicated that CpPLC was specific for phosphatidylcholine and sphingomyelin. Molecular docking studies described in this paper predict favorable interactions of the CpPLC active site with other phospholipids, e.g. phosphatidylethanolamine, phosphatidylinositol and, to a lesser extent, phosphatidylglycerol. On the basis of these predictions, we have performed experimental studies showing α-toxin to degrade all the phospholipids mentioned above. The molecular docking data also provide an explanation for the observed lower activity of CpPCL on sphingomyelin as compared to the glycerophospholipids.     

Differential Sensitivity of Macrocarpa and Microcarpa Types of Chickpea （Cicer arietinum L.） to Water Stress： Association of Contrasting Stress Response with Oxidative Injury

Chickpea (Cicer arietinum L.) is particularly sensitive to water stress at its reproductive phase and, under conditions of water stress, will abort flowers and pods, thus reducing yield potential. There are two types of chickpea: (i) Macrocarpa (“Kabuli”), which has large, rams head‐shaped, light brown seeds; and (ii) Microcarpa (“Desi”), which has small, angular and dark‐brown seeds. Relatively speaking, “Kabuli” has been reported to be more sensitive to water stress than “Desi”. The underlying mechanisms associated with contrasting sensitivity to water stress at the metabolic level are not well understood. We hypothesized that one of the reasons for contrasting water stress sensitivity in the two types of chickpea may be a variation in oxidative injury. In the present study, plants of both types were water stressed at the reproductive stage for 14 d. As a result of the stress, the “Kabuli” type exhibited an 80% reduction in seed yield over control compared with a 64% reduction observed for the “Desi” type. The decrease in leaf water potential (Ψ_w) was faster in the “Kabuli” compared with the “Desi” type. At the end of the water stress period, Ψ_w was reduced to ?2.9 and ?3.1 MPa in the “Desi” and “Kabuli” types, respectively, without any significant difference between them. On the last day of stress, “Kabuli” experienced 20% more membrane injury than “Desi”. The chlorophyll content and photosynthetic rate were significantly greater in “Desi” compared with “Kabuli”. The malondialdehyde and H₂O₂ content were markedly higher at the end of the water stress in “Kabuli” compared with “Desi”, indicating greater oxidative stress in the former. Levels of anti‐oxidants, such as ascorbic acid and glutathione, were significantly higher in “Desi” than “Kabuli”. Superoxide dismutase and catalase activity did not differ significantly between the two types of chickpea, whereas on the 10th day, the activities of ascorbate peroxidase, dehydroascorbate reductase, and glutathione reductase were higher in “Desi”. These findings indicate that the greater stress tolerance in the “Desi” type may be ascribed to its superior ability to maintain better water status, which results in less oxidative damage. In addition, laboratory studies conducted by subjecting both types of chickpea to similar levels of polyethylene glycol‐induced water stress and to 10 μ.mol/L abscisic acid indicated a greater capacity of the “Desi” type to deal with oxidative stress than the “Kabuli” type. (Managing editor: Ping He)     

Phylogenenetic approach of the section Bulbocodii D.C. of Narcissus based on cpDNA. A case of taxonomic inflation?

In this paper, we analyzed the phylogeny of the section Bulbocodii (genus Narcissus; Amarillydaceae) using the matK and trnL-F fragments of cpDNA in order to review the validity of the recognized taxa. Our results indicate that Narcissus obesus should be considered a valid species, and that N. blancoi is a distinct taxon. In addition, seven previously recognized species, N. juressianus, N. subnivalis, N. graellsii, N. conspicuus, N. citrinus, N. nivalis, and N. quintanilhae, should be assigned to an infraspecific rank under N. bulbocodium, as they are not valid species. In addition, we analyzed the distribution of the three morphological characters widely used in the systematics of this section and found that their variation does not agree with the phylogenetic results, rendering these characters limited taxonomical utility. This result suggests that the section Bulbocodii shows high morphological lability, which can explain the proliferation of nominal species.     

Genetic dissection of drought tolerance in chickpea (Cicer arietinum L.)

Key message

Analysis of phenotypic data for 20 drought tolerance traits in 1–7 seasons at 1–5 locations together with genetic mapping data for two mapping populations provided 9 QTL clusters of which one present on CaLG04 has a high potential to enhance drought tolerance in chickpea improvement.

Abstract

Chickpea (Cicer arietinum L.) is the second most important grain legume cultivated by resource poor farmers in the arid and semi-arid regions of the world. Drought is one of the major constraints leading up to 50 % production losses in chickpea. In order to dissect the complex nature of drought tolerance and to use genomics tools for enhancing yield of chickpea under drought conditions, two mapping populations—ICCRIL03 (ICC 4958 × ICC 1882) and ICCRIL04 (ICC 283 × ICC 8261) segregating for drought tolerance-related root traits were phenotyped for a total of 20 drought component traits in 1–7 seasons at 1–5 locations in India. Individual genetic maps comprising 241 loci and 168 loci for ICCRIL03 and ICCRIL04, respectively, and a consensus genetic map comprising 352 loci were constructed (http://cmap.icrisat.ac.in/cmap/sm/cp/varshney/). Analysis of extensive genotypic and precise phenotypic data revealed 45 robust main-effect QTLs (M-QTLs) explaining up to 58.20 % phenotypic variation and 973 epistatic QTLs (E-QTLs) explaining up to 92.19 % phenotypic variation for several target traits. Nine QTL clusters containing QTLs for several drought tolerance traits have been identified that can be targeted for molecular breeding. Among these clusters, one cluster harboring 48 % robust M-QTLs for 12 traits and explaining about 58.20 % phenotypic variation present on CaLG04 has been referred as “QTL-hotspot”. This genomic region contains seven SSR markers (ICCM0249, NCPGR127, TAA170, NCPGR21, TR11, GA24 and STMS11). Introgression of this region into elite cultivars is expected to enhance drought tolerance in chickpea.     

Interspecific Transition Among Caragana microphylla,C. davazamcii and C. korshinskii Along Geographic Gradient. Ⅱ. Characteristics of Photosynthesis and Water Metabolism

The characteristics of photosynthesis and water metabolism of Caragana microphylla Lam., C. davazamcii Sancz. and C. korshinskii Kom. populations in different sites (117.6o-105.7o E, 44.6o-38.8o N) were studied. (1) From the east to the west, the responses of the three species to photosynthetically available radiation (PAR) in net photosynthesis rate increased, the relative humidity of the air which corresponded to the occurrence of maximum photosynthesis rate decreased, and the corresponding air temperature increased. Along the same gradient, the before-noon superiority of the photosynthesis became evident, and the photosynthesis rate and the light use efficiency (LUE ) increased, while the transpiration rate decreased, thus the water use efficiency (WUE ) increased notably, and the leaf water content decreased gradually. From the east to the west, the plants took a water-saving strategy step by step with higher photosynthesis rate and lower transpiration rate. These physiological changes in the plants were adaptable to the conditions of light, temperature and humidity in the habitat of the plants, and might be the biological foundation for the geographical transition among C. microphylla , C. davazamcii and C. korshinskii. (2) The adaptation of photosynthetic system of C. microphylla , C. davazamcii and C. korshinskii to PAR, air humidity and temperature exhibited the interspecific continuity, which was consistent with the environmental gradient. In different species and different sites, the diurnal changes of net photosynthesis rate, the daily cumulative value of net photosynthesis, the diurnal changes of transpiration rate, the daily cumulative value of transpiration, the water use efficiency and the diurnal changes of leaf water content varied with longitudinal descent (from the east to the west). The characteristics of photosynthesis and water metabolism indicated that the geographical transition among C. microphylla , C. davazamcii and C. korshinskii was in gradual change, and these three species formed a geographical cline.     

The amino acid sequence of cytochrome c′ from Alcaligenes sp. N.C.I.B. 11015

The amino acid sequence of the cytochrome c' from Alcaligenes sp. N.C.I.B. 11015 (Iwasaki's ;Pseudomonas denitrificans') has been determined. This organism is the only non-photosynthetic bacterium in which the protein has been found. The protein consists of a single polypeptide chain of 127 residues, with a single haem covalently attached to two cysteines. Unlike normal cytochromes c, the haem attachment site is very close to the C-terminus. The amino acid sequence around the haem attachment site is very similar to that of Chromatium vinosum D cytochrome c'. Detailed evidence for the amino acid sequence of the protein has been deposited as Supplementary Publication SUP 50022 at the British Library (Lending Division), (formerly the National Lending Library for Science and Technology), Boston Spa, Yorks. LS23 7BQ, U.K., from whom copies may be obtained on the terms given in Biochem. J. (1973) 131, 5.     

Ecological Partitioning of Cercopithecus campbelli, C. petaurista, and C. diana in the Taï Forest

I determined the degree of ecological partitioning among 3 species of guenons (Cercopithecus campbelli, C. petaurista, and C. diana) in the Taï Forest, Côte d’Ivoire and used the partitioning data to understand competitive relationships among them. Over a 13-mo period, I measured ecological partitioning in terms of food and canopy stratum use for 2 habituated groups of each guenon species and also collected data on food availability. I found that the study species diverged primarily in food items consumed and vertical strata occupied. Cercopithecus petaurista ate much more foliage than the other species did and used mostly the middle strata (5–20 m). Cercopithecus diana ate primarily fruit and used mostly the upper strata (>20 m). Cercopithecus campbelli ate mostly fruit together with large amounts of animal matter and primarily occupied the ground and low strata (<5 m). Of the specific pairs, the diets of Cercopithecus campbelli/C. diana overlapped the most overall and decreased during the season of low fruit availability. Cercopithecus campbelli and C. diana age/sex classes also overlapped more than the age/sex classes of other species pairs. The results suggest that the potential for competition was more intense for Cercopithecus campbelli/C.diana relations than it is for other species pairs. I compare my results from Taï with those from other primate and guenon communities and demonstrate that dietary overlaps and seasonal dietary divergence are lower in Taï than in most other guenon communities.