Effect of benzimidazole on nicotinamide adenine dinucleotide phosphate phosphomonosterase activity in wheat leaves

Summary Nicotinamide adenine dinucleotide phosphate phosphomonoesterase was isolated and partially purified from wheat (Triticum aestivum L. var. Selkirk) leaves. The enzyme hadK _NADP value of 1.4×10^–4 M and a pH optimum of 5.9.In vitro activity of this enzyme was unaffected by precursors of NAD (nicotinamide and nicotinic acid) or cytokinis (kinetin and benzimidazole). However, when detached wheat leaves were treated with solutions of these compounds, the precursors lowered the specific activity while the cytokinins enhanced the activity. It is suggested that spatial separation and compartmentation of the enzyme and its substrate NADP account for the similar effect of benzimidazole on both.This work was supported by a grant No. A2698 from the National Research Council, Canada.     

Relationship of prodigiosin condensing enzyme activity to the biosynthesis of prodigiosin and its precursors in Serratia marcescens

Prodigiosin condensing enzyme (PCE) activities were present in Serratia marcescens wild type 08, mutants OF, WF and 9-3-3. Their specific activities exhibited different maxima and at different times during the late log phase or the early stationary phase of cell growth. The levels of prodigiosin and its precursors also showed a significant increase at this period. The results support that prodigiosin and/or its precursors are secondary metabolites. The ubiquity of the PCE activity in mutants deficient in prodigiosin biosynthesis suggest that this particular enzyme may also be present in non-pigmented clinical isolates.     

Relationship of prodigiosin condensing enzyme activity to the biosynthesis of prodigiosin and its precursors inSerratia marcescens

Summary Prodigiosin condensing enzyme (PCE) activities were present inSerratia marcescens wild type 08, mutants OF, WF and 9-3-3. Their specific activities exhibited different maxima and at different times during the late log phase or the early stationary phase of cell growth. The levels of prodigiosin and its precursors also showed a significant increase at this period. The results support that prodigiosin and/or its precursors are secondary metabolites. The ubiquity of the PCE activity in mutants deficient in prodigiosin biosynthesis suggest that this particular enzyme may also be present in non-pigmented clinical isolates.     

Presence of a protein disulfide isomerase (EC 5.3.4.1) in wheat germ]

The presence of a protein disulfide isomerase (rearrangease) in wheat embryo has been demonstrated by its ability in reactivating randomly cross-linked ribonuclease. This activity requires a dialysable cofactor; after dialysis, the activity is recovered by addition of reduced glutathione. The enzyme can be precipitated by 70% saturation ammonium sulfate.     

Comparative study of oxalate oxidase in three genotypes ofSorghum vulgare

Summary The presence of an oxalate oxidase (EC 1.2.3.4) has been demonstrated in 15,000×g supernatants prepared from 10-day-old seedlings of three genotypes ofSorghum vulgare: grain sorghum hybrid (CSH-5), grain-cum-forage sorghum (PC-6) and forage sorghum (PC-1). The specific activity of the enzyme in the different tissues of seedlings was found to be present in the order leaves > stems > roots in PC-6 and PC-1, but this order was reversed in CSH-5. A comparison of the different properties of the leaf enzyme of these three genotypes of sorghum revealed that the enzyme has maximum activity in the acidic pH range from 4.0 to 5.0 and in the temperature range from 37°C to 40°C. The enzyme was stimulated by Cu²⁺ and Fe²⁺. The rate of H₂O₂ formation in the enzyme reaction was linear up to 5 min and was stoichiometrically related to oxalate consumption. The enzyme is unaffected by Na⁺ at physiological concentration (0.15 M). The superiority of this enzyme over moss and other plant enzymes for enzymic determination of urinary oxalate is discussed.     

Inactivation of N-assimilating enzymes and proteolytic activities in wheat leaf extracts: Effect of pyridine nucleotides and of adenylates

Summary Nitrate reductase was protected from inactivation in wheat leaf extracts by NADH, while NADPH was less effective. NAD, NADP or adenylates did not affect nitrate reductase inactivation in vitro. Glutamine synthetase was more stable than nitrate reductase and was protected from inactivation by ATP. ADP, AMP or pyridine nucleotides had no or only a minor effect on the decrease of glutamine synthetase activity in extracts. The caseolytic activity extracted from senescing leaves was slightly decreased by NADH and NADPH but this effect was not sufficient to explain the stabilization of nitrate reductase by NADH. Oxidized pyridine nucleotides and adenylates had no major effect on the caseolytic activity under the conditions used.This work was supported by grant 3.067-0.81 from the Swiss National Science Foundation. Author for correspondence: U.F.     

Chitin synthetase activity and inhibition in different insect microsomal preparations

Summary To facilitate massive screening and for structure-activity relationship studies of chitin synthesis inhibitors, methods to obtain the chitin synthetase (CS) containing microsomal fraction from the postmitochondrial supernatant were examined. Compared with fractionation by differential centrifugation, the CaCl₂ precipitate yielded the most active CS preparation. Acidification (pH 5.6) and polyethylene glycol 8000 (5%) treatments resulted in relatively low CS activity. Inhibitory effects were detected with polyoxin-D and 1-geranyl-2-methyl benzimidazole, a novel CS inhibitor, but not with benzoylphenyl ureas.     

Influence of sex,developmental time and food onβ-N-acetylglucosaminidase activity in the rice weevilSitophilus oryzae L.

Summary The effects of some biological parameters on -N-acetylglucosaminidase activity have been investigated inS. oryzae. There is no significant influence of sex and developmental time on the enzyme activity level, which appears in contrast to be greatly influenced by food (wheat or sorghum). Sorghum contains competitive inhibitors which are almost completely removed after dialysis. Fasting relieves this inhibition very quickly, suggesting that inhibitors act directly at the gut level.     

Comparative kinetics of greening in etiolated intact and excized wheat leaves

Summary Differential characteristics of lag phase of chlorophyll accumulation during greening of etiolated intact and excized wheat leaves are discussed. Relatively, a prolonged lag phase in case of excized leaves is attributed to the lack of proper hormonal level in the leaves after excision.Acknowledgment. We thank Prof. M. C. Dash for encouragement and facilities. The assistance of Mr A. N. Misra is acknowledged.     

Effects of 2-β-D-glucosyl-4-hydroxy-7-methoxy-1,4-benzoxazin-3-one onSchizaphis graminum (rondani) (insecta,aphididae) feeding on artificial diets

Summary 2--Glucosyl-4-hydroxy-7-methoxy-1,4-benzoxazin-3-one (DIMBOA-Glc), the main hydroxamic acid from maize and wheat, and its aglucone, decreased survival ofSchizaphis graminum reared on artificial diets. Both compounds were toxic for aphids and acted as feeding deterrents, at concentrations as low as 1 mM. The natural concentrations of glucosides of hydroxamic acids in wheat leaves reach up to 6 mmoles/kg fresh weight, thus falling within the range in which DIMBOA-Glc causes delecterious effects to diet-fed aphids.To whom correspondence should be addressed. THis work was supported by grants from International Foundation for Science (484), Universidad de Chile (N-1654) and the Agency for International Development.     

Nachweis pflanzlicher Ribonukleasen in Polyacrylamid-Gelen nach Disk-Elektrophorese

Summary Detection of ribonucleases in polyacrylamide gels after disc-electrophoresis is possible by incubation of the gels in a solution of low molecular RNA followed by staining with methylene blue. Application of this method to protein extracts of wheat leaves, sugar beet leaves and roots of different bean varieties as well as callus cultures and roots of the same bean variety shows differences in number and position of RNase zones.     

Post-proteasomal and proteasome-independent generation of MHC class I ligands

Peptide ligands presented by MHC class I molecules are produced by intracellular proteolysis, which often involves multiple steps. Initial antigen degradation seems to rely almost invariably on the proteasome, although tripeptidyl peptidase II (TPP II) and insulin-degrading enzyme (IDE) may be able to substitute for the proteasome in rare cases. Recent evidence suggests that the net effect of cytosolic aminopeptidases is destruction of potential class I ligands, although a positive role in selected cases has been documented. This may apply particularly to the trimming of long precursors by TPP II. In contrast, trimming of ligand precursors in the endoplasmic reticulum is essential for the generation of suitable peptides and has a substantial impact on the repertoire of ligands presented. Trimming by the ER aminopeptidase (ERAP) enzymes most likely acts on free precursors and is adapted to the needs of class I molecules by way of a molecular ruler mechanism. Trimming by ERAP enzymes also occurs for cross-presented ligands, which can alternatively be processed in a special endosomal compartment by insulin-regulated aminopeptidase.     

H2 receptor antagonists and human granulopoiesis

The effect of 2 H2 receptor antagonists (ranitidine and cimetidine) on the in vitro growth of human granulomonopoietic precursors (CFU-GM) was studied. Ranitidine, although having an anti H2 receptor activity much greater than that of cimetidine, displays the same toxicity for CFU-GM.     

Secretion of chitinase by Aphanocladium album, a hyperparasite of wheat rust

Aphanocladium album, a hyperparasite of wheat rust, Puccinia graminis var. tritici, secretes chitinase when grown on a chitin-containing medium. This enzyme was purified and shown to attack the mycelium of the parasitized rust fungus.     

ADAMTS-2 functions as anti-angiogenic and anti-tumoral molecule independently of its catalytic activity

ADAMTS-2 is a metalloproteinase that plays a key role in the processing of fibrillar procollagen precursors into mature collagen molecules by excising the amino-propeptide. We demonstrate that recombinant ADAMTS-2 is also able to reduce proliferation of endothelial cells, and to induce their retraction and detachment from the substrate resulting in apoptosis. Dephosphorylation of Erk1/2 and MLC largely precedes the ADAMTS-2 induced morphological alterations. In 3-D culture models, ADAMTS-2 strongly reduced branching of capillary-like structures formed by endothelial cells and their long-term maintenance and inhibited vessels formation in embryoid bodies (EB). Growth and vascularization of tumors formed in nude mice by HEK 293-EBNA cells expressing ADAMTS-2 were drastically reduced. A similar anti-tumoral activity was observed when using cells expressing recombinant deleted forms of ADAMTS-2, including catalytically inactive enzyme. Nucleolin, a nuclear protein also found to be associated with the cell membrane, was identified as a potential receptor mediating the antiangiogenic properties of ADAMTS-2.     

Administration of D-alanine did not cause increase of D-amino acid oxidase activity in mice.

D-amino acid oxidase (DAAO) activity was not altered in the liver and kidney by oral administration of D-alanine to adult mice. The enzyme was apparently not induced by the enteric microflora either, since the enzyme activity in the liver and kidney of germ-free mice was not different from that of specific-pathogen-free mice. The times of appearance of DAAO activity and of free D-amino acids in the kidney were elucidated using suckling mice. DAAO activity started to increase 7 days after birth, and reached almost the adult level by 28 days. The content of free neutral D-amino acids also increased with age, in a similar fashion. A possible conclusion is that the enzyme activity normally increases during this period, to eliminate the free D-amino acids which have increased with age in the suckling mice. Consequently, the administration of D-alanine had no further effect in increasing enzyme activity.     

Administration of D-alanine did not cause increase of D-amino acid oxidase activity in mice

D-amino acid oxidase (DAAO) activity was not altered in the liver and kidney by oral administration of D-alanine to adult mice. The enzyme was apparently not induced by the enteric microflora either, since the enzyme activity in the liver and kidney of germ-free mice was not different from that of specific-pathogen-free mice. The times of appearance of DAAO activity and of free D-amino acids in the kidney were elucidated using suckling mice. DAAO activity started to increase 7 days after birth, and reached almost the adult level by 28 days. The content of free neutral D-amino acids also increased with age, in a similar fashion. A possible conclusion is that the enzyme activity normally increases during this period, to eliminate the free D-amino acids which have increased with age in the suckling mice. Consequently, the administration of D-alanine had no further effect in increasing enzyme activity.     

A histochemical study of leucine amino peptidase activity in the testes of immature and mature guinea-pigs

Summary The distribution of the enzyme leucine amino peptidase (LAP) has been studied histochemically in immature and mature guinea-pig testes. Immature guinea-pig testis showed a very feeble LAP activity, while in mature ones strong LAP activity was noted. The present communication indicates a direct relationship between the activity of the enzyme LAP and sexual maturation.     

H2 receptor antagonists and human granulopoiesis

Summary The effect of 2 H₂ receptor antagonists (ranitidine and cimetidine) on the in vitro growth of human granulomonopoietic precursors (CFU-GM) was studied. Ranitidine, although having an anti H₂ receptor activity much greater than that of cimetidine, displays the same toxicity for CFU-GM.Acknowledgments. This work was supported by CNR, Rome, PFCCN and AIRC, Milan.     

Debriding ability of a novel multi-enzyme preparation isolated from Antarctic krill (Euphausia superba)

Summary The wound-debriding activity of various types of proteolytic enzymes and proteases from Antarctic krill (multi-enzyme system consisting of both endo- and exopeptidases) was evaluated. The results, based on the enzymatically acieved weight reduction of a necrotic animal material (excised rat skin) in vitro, clearly showed that the multi-enzyme system (krill) had a higher degrading activity than the single enzyme preparation, or that with only a few enzymes. The debriding effect of the krill enzymes was markedly related to the enzyme concentration, resulting in 70–100% substrate degradation after 24 h. The digesting capacity of trypsin reached about 50%, but an increase in concentration of this enzyme did not substantially influence its overall activity. The effect of streptokinase-streptodornase, collagenase and plasmin-desoxyribonuclease was weak (10–20% digested).