Majority of Actinomadura clinical isolates from sputa or bronchoalveolar lavage fluid in Japan belongs to the cluster of Actinomadura cremea and Actinomadura nitritigenes, and the description of Actinomadura chibensis sp. nov

In Japan during 1996-2004, 21 actinomycete strains that have madurose as the diagnostic cell-wall sugar and show true branching in their substrate and aerial mycelia were isolated from sputa or bronchoalveolar lavage (BAL) fluid of patients with pulmonary infections or who were suspected of having related infections. Chemotaxonomic studies showed that all the isolates belong to the genus Actinomadura. Among them, six and seven strains were classified respectively into clusters of Actinomadura nitritigenes and Actinomadura cremea based on 16S rDNA analyses because their 16S rDNA similarities to those respective species were greater than 99.5%. To our knowledge, this is first report that strains of above two species were isolated from clinical specimens. Neither Actinomadura madurae nor Actinomadura pelletieri strain was isolated, and one new species, Actinomadura chibensis, was proposed; the remaining seven strains were not assigned into any known species, suggesting the presence of another new Actinomadura species.     

Substrate recognition by unsaturated glucuronyl hydrolase from Bacillus sp. GL1

Bacterial unsaturated glucuronyl hydrolases (UGLs) together with polysaccharide lyases are responsible for the complete depolymerization of mammalian extracellular matrix glycosaminoglycans. UGL acts on various oligosaccharides containing unsaturated glucuronic acid (DeltaGlcA) at the nonreducing terminus and releases DeltaGlcA through hydrolysis. In this study, we demonstrate the substrate recognition mechanism of the UGL of Bacillus sp. GL1 by determining the X-ray crystallographic structure of its substrate-enzyme complexes. The tetrasaccharide-enzyme complex demonstrated that at least four subsites are present in the active pocket. Although several amino acid residues are crucial for substrate binding, the enzyme strongly recognizes DeltaGlcA at subsite -1 through the formation of hydrogen bonds and stacking interactions, and prefers N-acetyl-d-galactosamine and glucose rather than N-acetyl-d-glucosamine as a residue accommodated in subsite +1, due to the steric hindrance.     

Atomic structure of the sweet-tasting protein thaumatin I at pH 8.0 reveals the large disulfide-rich region in domain II to be sensitive to a pH change

Thaumatin, an intensely sweet-tasting plant protein, elicits a sweet taste at 50 nM. Although the sweetness remains when thaumatin is heated at 80 °C for 4 h under acid conditions, it rapidly declines when heating at a pH above 6.5. To clarify the structural difference at high pH, the atomic structure of a recombinant thaumatin I at pH 8.0 was determined at a resolution of 1.0 Å. Comparison to the crystal structure of thaumatin at pH 7.3 and 7.0 revealed the root-mean square deviation value of a Cα atom to be substantially greater in the large disulfide-rich region of domain II, especially residues 154–164, suggesting that a loop region in domain II to be affected by solvent conditions. Furthermore, B-factors of Lys137, Lys163, and Lys187 were significantly affected by pH change, suggesting that a striking increase in the mobility of these lysine residues, which could facilitate a reaction with a free sulfhydryl residue produced via the β-elimination of disulfide bonds by heating at a pH above 7.0. The increase in mobility of lysine residues as well as a loop region in domain II might play an important role in the heat-induced aggregation of thaumatin above pH 7.0.     

Recognition of heteropolysaccharide alginate by periplasmic solute-binding proteins of a bacterial ABC transporter

Alginate is a heteropolysaccharide that consists of β-D-mannuronate (M) and α-L-guluronate (G). The Gram-negative bacterium Sphingomonas sp. A1 directly incorporates alginate into the cytoplasm through the periplasmic solute-binding protein (AlgQ1 and AlgQ2)-dependent ABC transporter (AlgM1-AlgM2/AlgS-AlgS). Two binding proteins with at least four subsites strongly recognize the nonreducing terminal residue of alginate at subsite 1. Here, we show the broad substrate preference of strain A1 solute-binding proteins for M and G present in alginate and demonstrate the structural determinants in binding proteins for heteropolysaccharide recognition through X-ray crystallography of four AlgQ1 structures in complex with saturated and unsaturated alginate oligosaccharides. Alginates with different M/G ratios were assimilated by strain A1 cells and bound to AlgQ1 and AlgQ2. Crystal structures of oligosaccharide-bound forms revealed that in addition to interaction between AlgQ1 and unsaturated oligosaccharides, the binding protein binds through hydrogen bonds to the C4 hydroxyl group of the saturated nonreducing terminal residue at subsite 1. The M residue of saturated oligosaccharides is predominantly accommodated at subsite 1 because of the strict binding of Ser-273 to the carboxyl group of the residue. In unsaturated trisaccharide (ΔGGG or ΔMMM)-bound AlgQ1, the protein interacts appropriately with substrate hydroxyl groups at subsites 2 and 3 to accommodate M or G, while substrate carboxyl groups are strictly recognized by the specific residues Tyr-129 at subsite 2 and Lys-22 at subsite 3. Because of this substrate recognition mechanism, strain A1 solute-binding proteins can bind heteropolysaccharide alginate with different M/G ratios.     

Practical removal of radioactivity from sediment mud in a swimming pool in Fukushima, Japan by immobilized photosynthetic bacteria

About 90% of the radioactive Cs in the sediment mud of a school's swimming pool in Fukushima, Japan was removed by treatment for 3 d using the alginate immobilized photosynthetic bacterium Rhodobcater sphaeroides SSI. Even though batch treatment was carried out 3 times repeatedly, the activity of immobilized cells in removing Cs was maintained at levels of about 84% (second batch) and 78% (third batch). Cs was strongly attached to the sediment mud because, even with HNO(3) treatment at pH of 2.00-1.60 for 24 h, it was not eluted into the water. Furthermore, more than 75% of the Cs could be removed without solubilization with HNO(3). This suggests that the Cs attached to the sediment mud was transformed into immobilized cells via the Cs(+) ion by the negative charge of the immobilized cell surface and/or the potassium transport system of the photosynthetic bacterium.     

Eutrophication of mountain lakes in Japan due to increasing deposition of anthropogenically produced dust

Atmospheric dust has wide-reaching effects, not only influencing climate conditions, but also ecosystems. The eastern region of the Asian continent is one of the largest emitters of dust in the world, and recent economic growth in the region has been accompanied by an increase in anthropogenic emissions. However, the effects of increased Asian dusts on aquatic ecosystems are not well understood. We examined fossil pigments and zooplankton remains from ²¹⁰Pb-dated sediments taken from high mountain lakes of Hourai-Numa and Hachiman-Numa, located in the Towada-Hachimantai National Park of Japan Islands, to uncover historical changes in the phyto- and zooplankton community over the past 100?years. Simultaneously, we measured the geochemical variables of TOC, TN, TP, ??¹³C, ??¹⁵N, and lead isotopes (²⁰⁷Pb/²⁰⁶Pb, ²⁰⁸Pb/²⁰⁶Pb) in the sediments to identify environmental factors causing such changes. As a result, despite few anthropogenic activities in the watersheds, alpine lakes in Japan had increased algal and herbivore plankton biomasses by 3?C6 fold for recent years depending on the surrounding terrestrial vegetation and landscape conditions. Biological and biogeochemical proxies recorded from the lake sediments showed that this eutrophication occurred after the 1990s when P deposition increased as a result of atmospheric loading of dust transported from the Asian continent. The continued increase of anthropogenically produced dust may therefore impart damaging impacts on mountain ecosystems even if they are protected from direct anthropogenic disturbances.     

Differences in Salivary Alpha-Amylase and Cortisol Responsiveness following Exposure to Electrical Stimulation versus the Trier Social Stress Tests

Background

Cortisol is an essential hormone in the regulation of the stress response along the HPA axis, and salivary cortisol has been used as a measure of free circulating cortisol levels. Recently, salivary alpha-amylase (sAA) has also emerged as a novel biomarker for psychosocial stress responsiveness within the sympathetic adrenomedullary (SAM) system.

Principal Findings

We measured sAA and salivary cortisol in healthy volunteers after exposure to the Trier Social Stress Test (TSST) and electric stimulation stress. One hundred forty-nine healthy volunteers participated in this study. All subjects were exposed to both the TSST and electric stimulation stress on separate days. We measured sAA and salivary cortisol levels three times immediately before, immediately after, and 20 min after the stress challenge. The State (STAI-S) and Trait (STAI-T) versions of the Spielberger Anxiety Inventory test and the Profile of Mood State (POMS) tests were administered to participants before the electrical stimulation and TSST protocols. We also measured HF, LF and LF/HF Heart Rate Variability ratio immediately after electrical stimulation and TSST exposure. Following TSST exposure or electrical stimulation, sAA levels displayed a rapid increase and recovery, returning to baseline levels 20 min after the stress challenge. Salivary cortisol responses showed a delayed increase, which remained significantly elevated from baseline levels 20 min after the stress challenge. Analyses revealed no differences between men and women with regard to their sAA response to the challenges (TSST or electric stimulations), while we found significantly higher salivary cortisol responses to the TSST in females. We also found that younger subjects tended to display higher sAA activity. Salivary cortisol levels were significantly correlated with the strength of the applied electrical stimulation.

Conclusions

These preliminary results suggest that the HPA axis (but not the SAM system) may show differential response patterns to distinct kinds of stressors.     

PIP kinases and their role in plant tip growing cells

Phosphatidylinositol (4,5) bisphosphate, [PtdIns(4,5)P2], is a signaling lipid involved in many important processes in animal cells such as cytoskeleton organization, intracellular vesicular trafficking, secretion, cell motility, regulation of ion channels, and nuclear signaling pathways. In the last years PtdIns(4,5)P2 and its synthesizing enzyme, phosphatidylinositol phosphate kinase (PIPK), has been intensively studied in plant cells, revealing a key role in the control of polar tip growth. Analysis of the PIPK members from Arabidopsis thaliana, Oryza sativa and Physcomitrella patens showed that they share some regulatory features with animal PIPKs but also exert plant-specific modes of regulation. This review aims at giving an overview on the PIPK family from Arabidopsis thaliana and Physcomitrella patens. Even though their basic structure, modes of activation and physiological role is evolutionary conserved, modules responsible for plasma membrane localization are distinct for different PIPKs, depending on differences in physiological and/or developmental status of cells, such as polarized and non-polarized.     

Allelic diversification at the <Emphasis Type="Italic">wx</Emphasis> locus in landraces of Asian rice

To examine continuous variation of amylose levels in Asian rice (Oryza sativa) landraces, the five putative alleles (Wx ^a , Wx ⁱⁿ , Wx ^b , Wx ^op , and wx) at the wx locus were investigated in near-isogenic lines (NILs). Apparent amylose levels ranged from 0.5 to 29.9% in the NILs, showing a positive relation with the levels of Wx gene product, granule-bound starch synthase (GBSS) as well as the enzymatic activity per milligram starch granule. Only opaque (Wx ^op ) accessions had an enzymatic activity per GBSS that was reduced to half the level of the others. Nucleotide sequences in the Wx gene were compared among 18 accessions harboring the five different alleles. Each of the Wx alleles had a unique replacement, frame-shift or splice donor site mutation, suggesting that these nucleotide changes could be reflected in phenotype alterations. A molecular phylogenetic tree constructed using the Wx gene indicated that ssp. japonica forms a distinct clade, whereas ssp. indica forms different clades together with the wild progenitor. Unexpectedly, the wx allele of 160 (indica from Taiwan) joined the japonica lineage; however, comparisons using linked genes for two Taiwanese accessions revealed that the wx gene was the product of gene flow from japonica to indica. Therefore, the japonica lineage frequently included Wx ⁱⁿ , Wx ^b and wx, while Wx ^a and Wx ^op were found in the other lineages, strongly suggesting that allelic diversification occurred after divergence of the two subspecies. The present results were discussed in relation to the maintenance of agronomically valuable genes in various landraces.     

Effects of Protoceratium reticulatum yessotoxin on feeding rates of Acartia hudsonica: A bioassay using artificial particles coated with purified toxin

Paralytic shellfish toxins produced by dinoflagellates are known to deter copepod grazing. Dinoflagellate species, including Protoceratium reticulatum, also produce disulfated polyether yessotoxins that were previously referred to as diarrheic shellfish toxins. However, the role of yessotoxins in predator–prey relationships is not yet clear. In the present study, the effects of purified yessotoxin (YTX) on feeding activities of Acartia hudsonica (Copepoda, Calanoida) were experimentally investigated. Polystyrene fluorescent microspheres (10 μm in diameter) colored bright blue or yellow-green were coated with cell extracts of P. reticulatum that do not produce yessotoxins. The bright blue microspheres were further coated with YTX, and the yellow-green microspheres were used as the reference. The microspheres were then given to the copepods separately or in combination to measure clearance rates and feeding selectivity. A. hudsonica was found to feed on the yellow-green microspheres without YTX at twice the rate of the bright blue microspheres with YTX. We also confirmed that microsphere color per se did not affect the feeding rates. The bright blue microspheres adsorbed 1.8–43.3 pg of YTX per microsphere, which is similar to the cell-specific yessotoxin content of toxic P. reticulatum found in natural environments. These results suggest that production of yessotoxin is advantageous for P. reticulatum by deterring predation by copepods.