Development of β-1, 3‐glucanase activity in germinated tomato seeds.
Morohashi, Y. & Matsushima, H. (2000). Journal of Experimental Botany, 51(349), 1381-1387.
Laminarin‐hydrolysing activity developed in the endosperm of tomato (Lycopersicon esculentum) seeds following germination. The enzyme was basic (pI>10) and the apparent molecular mass was estimated to be 35 kDa by SDS‐PAGE. It was specific for linear β-1,3‐glucan substrates. Laminarin was hydrolysed by the enzyme to yield a mixture of oligoglucosides, indicating that the enzyme had an endo‐action pattern. Thus, the enzyme was identified as β-1,3‐ endoglucanase (EC 22.214.171.124). The activity of the enzyme developed in the endosperm after radicle protrusion (germination) had occurred and the enzyme activity was localized exclusively in the micropylar region of the endosperm where the radicle had penetrated. When the lateral endosperm region, where no induction of the enzyme occurred, was wounded (cut or punctured), there was a marked enhancement of β-1,3‐glucanase activity. Thus the post‐germinative β-1,3‐glucanase activity in the micropylar endosperm portion might be brought about by wounding resulting from endosperm rupture by radicle penetration.
Purification, characterization and structural analysis of an abundant β-1, 3‐glucanase from banana fruit.
Peumans, W. J., Barre, A., Derycke, V., Rougé, P., Zhang, W., May, G. D., Delcour, J. A., Van Leuven, F. & Van Damme, E. J. (2000). European Journal of Biochemistry, 267(4), 1188-1195.
An abundant, catalytically active β-1,3-endoglucanase (EC 126.96.36.199) has been isolated from the pulp of ripe bananas. Biochemical analysis of the purified protein, molecular modelling, and molecular cloning of the corresponding gene indicate that this banana enzyme closely resembles previously characterized plant β-glucanases with respect to its amino-acid sequence, structure and biological activity. The results described in this paper demonstrate both the occurrence of an abundant active β-1,3-endoglucanases in fruits and also readdress the question of the possible involvement of these enzymes in the ripening and/or softening process.
Lentinula edodes tlg1 encodes a thaumatin-like protein that is involved in lentinan degradation and fruiting body senescence.
Sakamoto, Y., Watanabe, H., Nagai, M., Nakade, K., Takahashi, M. & Sato, T. (2006). Plant Physiology, 141(2), 793-801.
Lentinan is an antitumor product that is purified from fresh Lentinula edodes fruiting bodies. It is a cell wall component, comprising β-1,3-glucan with β-1,6-linked branches, which becomes degraded during postharvest preservation as a result of increased glucanase activity. In this study, we used N-terminal amino acid sequence to isolate tlg1, a gene encoding a thaumatin-like (TL) protein in L. edodes. The cDNA clone was approximately 1.0 kb whereas the genomic sequence was 2.1 kb, and comparison of the two indicated that tlg1 contains 12 introns. The tlg1 gene product (TLG1) was predicted to comprise 240 amino acids, with a molecular mass of 25 kD and isoelectric point value of 3.5. The putative amino acid sequence exhibits approximately 40% identity with plant TL proteins, and a fungal genome database search revealed that these TL proteins are conserved in many fungi including the basidiomycota and ascomycota. Transcription of tlg1 was not detected in vegetative mycelium or young and fresh mushrooms. However, transcription increased following harvest. Western-blot analysis demonstrated a rise in TLG1 levels following harvest and spore diffusion. TLG1 expressed in Escherichia coli and Aspergillus oryzae exhibited β-1,3-glucanase activity and, when purified from the L. edodes fruiting body, demonstrated lentinan degrading activity. Thus, we suggest that TLG1 is involved in lentinan and cell wall degradation during senescence following harvest and spore diffusion.
Effects of benzothiadiazole and acetylsalicylic acid on β-1, 3‐glucanase activity and disease resistance in potato.
Bokshi, A. I., Morris, S. C. & Deverall, B. J. (2003). Plant Pathology, 52(1), 22-27.
Benzothiadiazole (BTH), as Bion WG50, and acetylsalicylic acid (ASA) treatments of potato foliage of field- and glasshouse-grown potato plants were compared for control of two foliar diseases, early blight (Alternaria solani) and powdery mildew (Erysiphe cichoracearum). The effect of these treatments on harvested tubers wound-inoculated with the dry rot fungus (Fusarium semitectum) was also evaluated. BTH (50 mg a.i. L-1) gave almost complete control of both foliar pathogens on inoculated glasshouse-grown plants and reduced the severity of leaf spotting diseases (mainly early blight) in the field. BTH (100 mg a.i. L-1) and ASA (400 mg a.i. L-1) reduced the severity of dry rot in field-grown tubers in some post-harvest wound-inoculated treatments but not others and a similar reduction occurred with tubers inoculated post-harvest from BTH-treated plants grown under glasshouse conditions. BTH treatment increased β-1,3-glucanase activity in leaves > stem > tubers > stolons but not in roots. Increased enzyme activity was recorded for up to 45 days post-treatment.
Endo-β-1, 3-glucanase GLU1, from the fruiting body of Lentinula edodes, belongs to a new glycoside hydrolase family.
Sakamoto, Y., Nakade, K. & Konno, N. (2011). Applied and Environmental Microbiology, 77(23), 8350-8354.
The cell wall of the fruiting body of the mushroom Lentinula edodes is degraded after harvesting by enzymes such as β-1,3-glucanase. In this study, a novel endo-type β-1,3-glucanase, GLU1, was purified from L. edodes fruiting bodies after harvesting. The gene encoding it, glu1, was isolated by rapid amplification of cDNA ends (RACE)-PCR using primers designed from the N-terminal amino acid sequence of GLU1. The putative amino acid sequence of the mature protein contained 247 amino acid residues with a molecular mass of 26 kDa and a pI of 3.87, and recombinant GLU1 expressed in Pichia pastoris exhibited β-1,3-glucanase activity. GLU1 catalyzed depolymerization of glucans composed of β-1,3-linked main chains, and reaction product analysis by thin-layer chromatography (TLC) clearly indicated that the enzyme had an endolytic mode. However, the amino acid sequence of GLU1 showed no significant similarity to known glycoside hydrolases. GLU1 has similarity to several hypothetical proteins in fungi, and GLU1 and highly similar proteins should be classified as a novel glycoside hydrolase family (GH128).
A (1→3)-β-glucanase expressed during oat endosperm development.
Martin, D. J. & Somers, D. A. (2004). Journal of Cereal Science, 39(2), 265-272.
In mature kernels of oat (Avena sativa L.) and other cereals, mixed-linked (11→33; 11→34)-β-glucans and arabinoxylans are major structural polysaccharides in cell walls of the endosperm. However, (1→3)-β-glucans are deposited transiently in walls during cellularization of endosperm early in grain development [Planta 202 (1997) 414–426]. The absence of (1→3)-β-glucans in mature endosperm cell walls suggests that (1→3)-β-glucanases are active during endosperm development. To investigate the role of β-glucanases during endosperm development, a (1→3)-β-glucanase cDNA, Oglc13, was isolated from an oat (A. sativa L.) kernel cDNA library. The enzymatic activity of the protein product, OGLC13, expressed from the cDNA in an in vitro expression system, exhibited substrate specificity for (1→3)-β-glucans. Oglc13 transcripts were detected in the endosperm of portions of developing kernels with the highest steady state level of mRNA at 15 days after anthesis (DAA) and not in vegetative tissues. Antibodies raised against OGLC13 immunoprecipitated (1→3)-β-glucanase activity from endosperm extracts of 10 and 15 DAA kernels and milky endosperm extracted from 15 DAA kernels. The OGLC13 antibodies did not precipitate (1→3)-β-glucanase activity from leaf extracts. These results indicated that Oglc13 is a unique (1→3)-β-glucanase expressed early in endosperm development.
Induction of defence responses in roots and mesocotyls of sorghum seedlings by inoculation with Fusarium thapsinum and F. proliferatum, wounding and light.
Huang, L. D. & Backhouse, D. (2005). Journal of Phytopathology, 153(9), 522-529.
The defence reactions of sorghum seedlings 7 days after inoculation with Fusarium thapsinum and F. proliferatum, and interactions with wounding and exposure to light were studied to determine whether responses to these fungi differed from those to abiotic stresses. In non-wounded plants, inoculation with both fungi increased concentrations of anthocyanins and soluble phenolics and activities of peroxidase (POX), chitinase and β-1,3-glucanase in the roots, and increased β-1,3-glucanase activity in the mesocotyls. There was no effect of inoculation on phenylalanine ammonia-lyase (PAL) activity. Wounding by itself increased anthocyanin content of mesocotyls. Wounding also had a variety of interactions with inoculation. Exposure to light had very little effect on any defence response measured. A time course experiment showed that induction of chitinase and β-1,3-glucanase occurred in less than 24 h after inoculation. POX activity increased 2 days after inoculation, followed by a transient increase in PAL activity. The content of anthocyanins and soluble phenolics in roots of inoculated seedlings increased gradually compared with controls over 6 days. The responses of sorghum seedlings to inoculation with F. thapsinum and F. proliferatum were similar to those found by other workers following challenge by necrotrophic pathogens and were different from those induced by wounding and exposure to light.
Bioprospecting in potato fields in the Central Andean Highlands: screening of rhizobacteria for plant growth-promoting properties.
Ghyselinck, J., Velivelli, S. L. S., Heylen, K., O’Herlihy, E., Franco, J., Rojas, M., de Vos, P. & Prestwich, B. D. (2013). Systematic and Applied Microbiology, 36(2), 116-127.
The Central Andean Highlands are the center of origin of the potato plant (Solanum tuberosum). Ages of mutualism between potato plants and soil bacteria in this region support the hypothesis that Andean soils harbor interesting plant growth-promoting (PGP) bacteria. Therefore, the aim of this study was to isolate rhizobacteria from Andean ecosystems, and to identify those with PGP properties. A total of 585 bacterial isolates were obtained from eight potato fields in the Andes and they were screened for suppression of Phytophthora infestans and Rhizoctonia solani. Antagonistic mechanisms were determined and antagonistic isolates were further tested for phosphate solubilization, 1-aminocyclopropane-1-carboxylate (ACC) deaminase activity, and production of NH3- and indole-3-acetic acid (IAA). PGP was studied in healthy and R. solani diseased plantlets under growth room conditions. Performance was compared to the commercial strain B. subtilis FZB24® WG. Isolates were dereplicated with matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS), and identified with 16S rRNA gene sequencing and multi locus sequence analysis (MLSA). A total of 10% of the isolates were effective antagonists, of which many were able to solubilize phosphate, and produce IAA, ACC deaminase, NH3 and hydrogen cyanide (HCN). During growth room experiments, 23 antagonistic isolates were associated with plant growth-promotion and/or disease suppression. Ten isolates had a statistically significant impact on test parameters compared to the uninoculated control. Three isolates significantly promoted plant growth in healthy plantlets compared to the commercial strain, and seven isolates outperformed the commercial strain in in vitro R. solani diseased plantlets.
Green-odour compounds have antifungal activity against the rice blast fungus Magnaporthe oryzae.
Tajul, M. I., Motoyama, T., Hatanaka, A., Sariah, M. & Osada, H. (2012). European Journal of Plant Pathology, 132(1), 91-100.
Four green-odour compounds—trans-2-hexenal, cis-3-hexenol, n-hexanal, and cis-3-hexenal—were applied (0.85 µg ml-1 as vapour) to rice plants in laboratory conditions to observe their biological activity against the phytopathogenic fungus Maganporthe oryzae, which causes rice blast disease worldwide. Two compounds, trans-2-hexenal and cis-3-hexenal, showed remarkable disease suppression efficacy (99.7% and 100% suppression, respectively), while n-hexanal had moderate (86.5%) and cis-3-hexenol had weak (20.8%) disease-suppressing effects. Pre-application and post-application of trans-2-hexenal or cis-3-hexenal had slight effects on blast incidence, suggesting that these compounds had direct effects to suppress M. oryzae infection. In fact, trans-2-hexenal and cis-3-hexenal exhibited a growth suppression effect on M. oryzae. Interestingly, these two compounds inhibited appressorium formation at lower concentrations than the growth suppression. Studies on the hypersensitive response (HR)-like reaction and plant β-1,3-glucanase activity in rice plant confirmed that induced resistance was not the major factor involved in the disease suppression mechanism. Results of this study conclusively showed that trans-2-hexenal and cis-3-hexenal possess potent inhibitory activities against the growth and the appressorium formation of M. oryzae and could be used as antifungal agents to significantly reduce M. oryzae infections in rice.
A novel antifungal Pseudomonas fluorescens isolated from potato soils in Greenland.
Michelsen, C. F. & Stougaard, P. (2011). Current Microbiology, 62(4), 1185-1192.
A rhizobacterium with high antifungal activity was isolated from a potato field at Inneruulalik, South Greenland. Phylogenetic analysis based on multi locus sequence typing showed that the bacterium was affiliated with strains of Pseudomonas fluorescens. The bacterium, denoted as Pseudomonas fluorescens In5, inhibited in vitro a broad range of phytopathogenic fungi, and the antifungal activity increased with decreasing temperature. Microcosm experiments demonstrated that P. fluorescens In5 protected tomato seedlings from Rhizoctonia solani. Transposon mutagenesis showed that the major cause for the antifungal activity of P. fluorescens In5 was a novel non-ribosomal peptide synthase (NRPS) gene. In addition, transposon mutagenesis showed that P. fluorescens In5 also contained a putative quinoprotein glucose dehydrogenase gene, which was involved in growth inhibition of phytopathogenic fungi. Although P. fluorescens In5 contained the capacity to synthesize hydrogen cyanide, β-1,3-glucanase, protease, and chitinase, these did not seem to play a role in the in vitro and microcosm antifungal assays.