Endo-beta-1,3-galactanase from winter mushroom Flammulina velutipes.

Arabinogalactan proteins are proteoglycans found on the cell surface and in the cell walls of higher plants. The carbohydrate moieties of most arabinogalactan proteins are composed of ß-1,3-galactan main chains and ß-1,6-galactan side chains, to which other auxiliary sugars are attached. For the present study, an endo-ß-1,3-galactanase, designated FvEn3GAL, was first purified and cloned from winter mushroom Flammulina velutipes. The enzyme specifically hydrolyzed ß-1,3-galactan, but did not act on ß-1,3-glucan, ß-1,3:1,4-glucan, xyloglucan, and agarose. It released various ß-1,3-galactooligosaccharides together with Gal from ß-1,3-galactohexaose in the early phase of the reaction, demonstrating that it acts on ß-1,3-galactan in an endo-fashion. Phylogenetic analysis revealed that FvEn3GAL is member of a novel subgroup distinct from known glycoside hydrolases such as endo-ß-1,3-glucanase and endo-ß-1,3:1,4-glucanase in glycoside hydrolase family 16. Point mutations replacing the putative catalytic Glu residues conserved for enzymes in this family with Asp abolished activity. These results indicate that FvEn3GAL is a highly specific glycoside hydrolase 16 endo-ß-1,3-galactanase.

Effect of the addition of calcium hydroxide on the hydrothermal-mechanochemical treatment of Eucalyptus.

The effect of Ca(OH)2 addition on optimization of hydrothermal-mechanochemical pretreatment, which combines hydrothermal and milling treatments, was examined. The highest glucose yield of 90% was achieved in the ball-milled specimen previously treated at 170°C in the presence of 20% Ca(OH)2 per substrate weight. The specific surface area of the substrate was closely correlated with glucose yield, and a larger specific surface area was obtained when treating the specimen at 170°C in the presence of Ca(OH)2 compared to treatment at 170°C without Ca(OH)2. Although the Ca(OH)2-treated specimen was relatively unaffected by delignification, the cleavage of the ester bonds between lignin and hemicellulose was confirmed by FT-IR. This suggests that Ca(OH)2 weakens the substrate structure by loosening the bonds between lignin and hemicellulose as the mechanism to increase the specific surface area regardless of the high lignin content, facilitating the fibrillation of fibers with mechanical milling.

Primary structure of dihydrofolate reductase and mitochondrial ribosomal protein L36 genes from the basidiomycete Coprinus cinereus.

We amplified and sequenced the dihydrofolate reductase (DHFR) gene of the basidiomycete Coprinus cinereus. Downstream of the DHFR coding region, a mitochondrial (mt) ribosomal protein L36 (RPL36) gene was discovered in the opposite orientation to DHFR gene. Putative polyadenylation signals of the two genes overlapped, both containing the 8-bp palindrome 5'-aatatatt-3'. The finding that C. cinereus DHFR gene is closely clustered with a mt protein gene strongly suggests that C. cinereus DHFR is closely related to mt function and evolution. The amino acid sequence of C. cinereus DHFR is most homologous to eukaryotic proteins such as Cryptococcus neoformans and Pneumocystis carinii DHFRs. However, the sequence of C. cinereus mt RPL36 closely resembles RPL36 of bacteria and cyanobacteria such as Synechocystis sp. and Escherichia coli. This result strongly supports the serial endosymbiotic theory of the development of ancestral eukaryotes, and suggests that C. cinereus mt RPL36 gene originated from the ancestral eubacterial genome.

Addition of alkali to the hydrothermal-mechanochemical treatment of Eucalyptus enhances its enzymatic saccharification.

The effects of alkali on hydrothermal-mechanochemical treatment (hydrothermal treatment combined with wet-milling) were examined with the aim of improving pretreatment of lignocellulosic biomass before enzymatic saccharification. After enzymatic saccharification, the highest glucose yield was obtained by autoclaving at 170°C in the presence of 20% NaOH per substrate weight. The wood fiber was unraveled into finer nanofibers by hydrothermal-mechanochemical treatment, thus increasing the specific surface area of the substrate from 11 to 132m(2)/g. Adding 20% NaOH to the treatment further increased the specific surface area of the already fibrillated substrate by 76% (232m(2)/g) due to lignin removal and ester bond cleavage between lignin and hemicellulose. This increase in specific surface area was closely related to the increase in enzymatic digestibility; therefore, NaOH addition may have enhanced the effect of hydrothermal-mechanochemical treatment.

Transient left ventricular apical ballooning developing after the Central Niigata Prefecture Earthquake: two case reports.

Two patients presented with transient left ventricular apical ballooning (takotsubo cardiomyopathy) induced by emotional stress caused by the Central Niigata Prefecture Earthquake in 2004. These patients complained of chest pain immediately after the earthquake. In patient 1, electrocardiography (ECG) showed slight ST elevation in leads V5 to V6 and 1 mm ST depression in lead III. Serial ECG revealed inverted giant T waves in leads V3 to V6 and inverted T waves in leads I, II, aVL and aVF 13 days after the earthquake occurred. Patient 2 also complained of chest pain right after the earthquake, but consulted a doctor 15 days after the earthquake occurred. ECG showed inverted giant T wave in leads V1 to V6 and inverted T waves in leads I, II and aVL. Transthoracic echocardiography showed hypokinesis of the apical area of the left ventricle with normokinesis in the basal area in both patients. Coronary angiography showed no stenotic segments and coronary spasms were not induced by provocative testing. Serial cardiac radionuclide single photon emission computed tomography of myocardial functional sympathetic innervation using iodine-123-metaiodobenzyl-guanidine (MIBG) and thiallium-201 (201Tl) showed an MIBG uptake defect and increased wash-out in the apical area, but only mild decrease of apical 201Tl uptake. Due to strong emotional stress, earthquakes may induce transient left ventricular apical ballooning (takotsubo cardiomyopathy).