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Basic β-1,3-Glucanase from Drosera binata Exhibits Antifungal Potential in Transgenic Tobacco Plants.

ABSTRACT: The basic β-1,3-glucanase of the carnivorous plant Drosera binata was tested as a purified protein, as well as under the control of a double CaMV35S promoter in transgenic tobacco for its capability to inhibit the growth of Trichoderma viride, Rhizoctonia solani, Alternaria solani, and Fusarium poae in an in-vitro assay. The purified protein inhibited tested phytopathogens but not the saprophytic fungus T. viride. Out of the analysed transgenic plants, lines 13, 16, 19, and 22 exhibited high DbGluc1 transcript abundance normalised to the actin transcript. Because of DbGluc1 transgene expression, lines 13 and 16 showed a 1.7-fold increase and lines 19 and 22 showed more than a 2-fold increase in total β-1,3-glucanase activity compared to the non-transgenic control. In accordance with the purified β-1,3-glucanase in-vitro antifungal assay, crude protein extracts of lines 19 and 22 significantly inhibited the growth of phytopathogens (14-34%). Further analyses revealed that the complementary action of transgenic β-1,3-glucanase and 20% higher activity of endogenous chitinase(s) in these lines were crucial for maximising the antifungal efficiency of crude protein extracts.

SUBMITTER: Rajninec M

PROVIDER: S-EPMC8401921 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Red rot resistant transgenic sugarcane developed through expression of β-1,3-glucanase gene.

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| S-EPMC5489175 | biostudies-literature

Improved barley broiler feed with transgenic malt containing heat-stable (1,3-1,4)-beta-glucanase.

Project description:The low nutritional value of barley for poultry is because of the absence of an intestinal enzyme for efficient depolymerization of (1, 3-1,4)-beta-glucan, the major polysaccharide of the endosperm cell walls. This leads to high viscosity in the intestine, limited nutrient uptake, decreased growth rate, and unhygienic sticky droppings adhering to chickens and floors of the production cages. Consequently, the 7.5 billion broiler chickens produced annually in the United States are primarily raised on corn-soybean diets. Here we show that addition to normal barley of 6.2% transgenic malt containing a thermotolerant (1,3-1,4)-beta-glucanase (4.28 microg.g(-1) soluble protein) provides a weight gain equivalent to corn diets. The number of birds with adhering sticky droppings is drastically reduced. Intestines and excrements of chickens fed the barley control diet contained large amounts of soluble (1,3-1,4)-beta-glucan, which was reduced by 75 and 50%, respectively, by adding transgenic malt to the diet. The amount of active recombinant enzyme in the small intestine corresponded to that present in the feed, whereas an 11-fold concentration of the enzyme was observed in the ceca, and a 7.5-fold concentration occurred in the excrement. Glycosylation of the beta-glucanase isolated from the ceca testified to its origin from the transgenic barley. Analysis of the data from this trial demonstrates the possibility of introducing individual recombinant enzymes into various parts of the gastrointestinal tract of chickens with transgenic malt and thereby the possibility of evaluating their effect on the metabolism of a given ingredient targeted by the enzyme.

| S-EPMC17606 | biostudies-literature

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| S-EPMC4532416 | biostudies-literature

An antifungal exo-alpha-1,3-glucanase (AGN13.1) from the biocontrol fungus Trichoderma harzianum.

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| S-EPMC93379 | biostudies-literature

Overexpression of an acidic endo-β-1,3-1,4-glucanase in transgenic maize seed for direct utilization in animal feed.

Project description:BackgroundIncorporation of exogenous glucanase into animal feed is common practice to remove glucan, one of the anti-nutritional factors, for efficient nutrition absorption. The acidic endo-β-1,3-1,4-glucanase (Bgl7A) from Bispora sp. MEY-1 has excellent properties and represents a potential enzyme supplement to animal feed.Methodology/principal findingsHere we successfully developed a transgenic maize producing a high level of Bgl7AM (codon modified Bgl7A) by constructing a recombinant vector driven by the embryo-specific promoter ZM-leg1A. Southern and Western blot analysis indicated the stable integration and specific expression of the transgene in maize seeds over four generations. The β-glucanase activity of the transgenic maize seeds reached up to 779,800 U/kg, about 236-fold higher than that of non-transgenic maize. The β-glucanase derived from the transgenic maize seeds had an optimal pH of 4.0 and was stable at pH 1.0-8.0, which is in agreement with the normal environment of digestive tract.Conclusion/significanceOur study offers a transgenic maize line that could be directly used in animal feed without any glucanase production, purification and supplementation, consequently simplifying the feed enzyme processing procedure.

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Analysis of gene families encoding acidic and basic beta-1,3-glucanases of tobacco.

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| S-EPMC55038 | biostudies-other

Action of an endo-β-1,3(4)-glucanase on cellobiosyl unit structure in barley β-1,3:1,4-glucan.

Project description:β-1,3:1,4-Glucan is a major cell wall component accumulating in endosperm and young tissues in grasses. The mixed linkage glucan is a linear polysaccharide mainly consisting of cellotriosyl and cellotetraosyl units linked through single β-1,3-glucosidic linkages, but it also contains minor structures such as cellobiosyl units. In this study, we examined the action of an endo-β-1,3(4)-glucanase from Trichoderma sp. on a minor structure in barley β-1,3:1,4-glucan. To find the minor structure on which the endo-β-1,3(4)-glucanase acts, we prepared oligosaccharides from barley β-1,3:1,4-glucan by endo-β-1,4-glucanase digestion followed by purification by gel permeation and paper chromatography. The endo-β-1,3(4)-glucanase appeared to hydrolyze an oligosaccharide with degree of polymerization 5, designated C5-b. Based on matrix-assisted laser desorption/ionization (MALDI) time-of-flight (ToF)/ToF-mass spectrometry (MS)/MS analysis, C5-b was identified as β-Glc-1,3-β-Glc-1,4-β-Glc-1,3-β-Glc-1,4-Glc including a cellobiosyl unit. The results indicate that a type of endo-β-1,3(4)-glucanase acts on the cellobiosyl units of barley β-1,3:1,4-glucan in an endo-manner.

| S-EPMC4673573 | biostudies-literature

Amyloid-like protein inclusions in tobacco transgenic plants.

Project description:The formation of insoluble protein deposits in human tissues is linked to the onset of more than 40 different disorders, ranging from dementia to diabetes. In these diseases, the proteins usually self-assemble into ordered ?-sheet enriched aggregates known as amyloid fibrils. Here we study the structure of the inclusions formed by maize transglutaminase (TGZ) in the chloroplasts of tobacco transplastomic plants and demonstrate that they have an amyloid-like nature. Together with the evidence of amyloid structures in bacteria and fungi our data argue that amyloid formation is likely a ubiquitous process occurring across the different kingdoms of life. The discovery of amyloid conformations inside inclusions of genetically modified plants might have implications regarding their use for human applications.

| S-EPMC2964307 | biostudies-literature

Beta-1,4-glucanase-like protein from the cyanobacterium Synechocystis PCC6803 is a beta-1,3-1,4-glucanase and functions in salt stress tolerance.

Project description:In the present study, we characterized the gene (Cyanobase accession number slr0897) designated Ssglc encoding a beta-1,4-glucanase-like protein (SsGlc) from Synechocystis PCC6803. The deduced amino acid sequence for Ssglc showed a high degree of similarity to sequences of GH (glycoside hydrolase) family 9 beta-1,4-glucanases (cellulases) from various sources. Surprisingly, the recombinant protein obtained from the Escherichia coli expression system was able to hydrolyse barley beta-glucan and lichenan (beta-1,3-1,4-glucan), but not cellulose (beta-1,4-glucan), curdlan (beta-1,3-glucan), or laminarin (beta-1,3-1,6-glucan). A 1H-NMR analysis of the enzymatic products revealed that the enzyme hydrolyses the beta-1,4-glycosidic linkage of barley beta-glucan through an inverting mechanism. The data indicated that SsGlc was a novel type of GH9 glucanase which could specifically hydrolyse the beta-1,3-1,4-linkage of glucan. The growth of mutant Synechocystis cells in which the Ssglc gene was disrupted by a kanamycin-resistance cartridge gene was almost the same as that of the wild-type cells under continuous light (40 micromol of photons/m2 per s), a 12 h light (40 micromol of photons/m2 per s)/12 h dark cycle, cold stress (4 degrees C), and high light stress (200 micromol of photons/m2 per s). However, under salt stress (300-450 mM NaCl), growth of the Ssglc-disrupted mutant cells was significantly inhibited as compared with that of the wild-type cells. The Ssglc-disrupted mutant cells showed a decreased rate of O2 consumption and NaHCO3-dependent O2 evolution as compared with the wild-type cells under salt stress. Under osmotic stress (100-400 mM sorbitol), there was no difference in growth between the wild-type and the Ssglc-disrupted mutant cells. These results suggest that SsGlc functions in salt stress tolerance in Synechocystis PCC6803.

| S-EPMC1925248 | biostudies-literature

Suppression of GhGLU19 encoding β-1,3-glucanase promotes seed germination in cotton.

Project description:BackgroundIn eudicots, germination begins with water uptake by the quiescent dry seed and is greatly related to the permeability of micropyle enriched callose layers. Once imbibition starts, seeds undergo a cascade of physiological, biochemical, and molecular events to initiate cellular activities. However, the effects of callose on water uptake and following seed metabolic events during germination are largely unknown. Cotton (Gossypium hirsutum) is a eudicot plant with natural fiber and edible oil production for humans. Here, we addressed this question by examining the role of GhGLU19, a gene encoding β-1,3-glucanase, in cotton seed germination.ResultsGhGLU19 belongs to subfamily B and was expressed predominately in imbibed seeds and early seedlings. Compared to wild type, GhGLU19-suppressing and GhGLU19-overexpressing transgenic cotton lines showed the higher and lower seed germination percentage, respectively. Callose was enriched more at inner integument (ii) than that in embryo and seed coat in cotton seeds. In GhGLU19-suppressing lines, callose at ii of cotton seeds was greatly increased and brought about a prolonged water uptake process during imbibition. Both proteomic and transcriptomic analysis revealed that contrary to GhGLU19-overexpressing lines, the glycolysis and pyruvate metabolism was decreased, and abscisic acid (ABA) biosynthesis related genes were downregulated in imbibed seeds of GhGLU19-suppressing lines. Also, endogenous ABA was significantly decreased in GhGLU19-suppressing line while increased in GhGLU19-overexpressing line.ConclusionsOur results demonstrate that suppression of GhGLU19 improves cotton seed germination via accumulating callose of inner integument, modulating glycolysis and pyruvate metabolism, and decreasing ABA biosynthesis. This study provides a potential way for improving germination percentage in cotton seed production, and other eudicot crops.

| S-EPMC9308338 | biostudies-literature

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