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Long-term phosphorus (P) fertilization results in P accumulation in agricultural soil and increases the risk of P leaching into water bodies. However, evaluating P leaching into groundwater is challenging, especially in clay soil with a high P sorption capacity. This study examined whether the combination of PO 4 oxygen isotope (δ 18 O PO4 ) analysis and the P saturation ratio (PSR) was useful to identify P enrichment mechanisms in groundwater. We investigated the groundwater and possible P sources in Kubi, western Japan, with intensive citrus cultivation. Shallow groundwater had oxic conditions with high PO 4 concentrations, and orchard soil P accumulation was high compared with forest soil. Although the soil had a high P sorption capacity, the PSR was above the threshold, indicating a high risk of P leaching from the surface orchard soil. The shallow groundwater δ 18 O PO4 values were higher than the expected isotopic equilibrium with pyrophosphatase. The high PSR and δ 18 O PO4 orchard soil values indicated that P leaching from orchard soil was the major P enrichment mechanism. The Bayesian mixing model estimated that 76.6% of the P supplied from the orchard soil was recycled by microorganisms. This demonstrates the utility of δ 18 O PO4 and the PSR to evaluate the P source and biological recycling in groundwater.

Recent studies have shown that Escherichia coli often carries a biosynthetic gene cluster termed either the pks island or the clb cluster that allows the production of a genotoxic polyketide-nonribosomal peptide hybrid secondary metabolite called colibactin. While the gene cluster is not always expressed, when the strain that resides in the colon produces the genotoxin, it is suspected to become a risk factor for colorectal cancer. Therefore, there is great interest in devising a simple method for the detection of colibactin-producing strains and understanding the detailed mechanism of how colibactin can induce oncogenesis, to develop convenient early screening methods and possible preventive treatments against colorectal cancer. However, the definitive chemical structure of colibactin remained elusive until recently, primarily due to its low yield and instability. In this review, we will briefly trace the recent studies leading to the identification of the structure of the active intact colibactin. Subsequently, we will describe our efforts toward developing simple methods for detecting colibactin producers, where we established methods based on the conventional polymerase chain reaction and loop-mediated isothermal amplification techniques. We also designed an activity-based fluorogenic probe for detecting colibactin-producing strains that could discern colibactin production levels among the E. coli strains screened. Using the probe, we isolated a wild-type high-colibactin-producing strain from a colorectal cancer tissue sample that proved to be valuable in identifying new colibactin metabolites and structurally characterizing them by nuclear magnetic resonance. Those techniques and the chemical insight they furnished should improve the fight against colorectal cancer.

Light molecules such as H 2 O are the systems in which we can have access to quantum mechanical information on their constituent atoms. Here, we have investigated electron transport through H 2 O@C 60 single molecule transistors (SMTs). The H 2 O@C 60 SMTs exhibit Coulomb stability diagrams that show multiple tunneling-induced excited states below 30 meV. Furthermore, we have performed terahertz (THz) photocurrent spectroscopy on H 2 O@C 60 SMTs and confirmed the same excitations. From comparison between experiment and theory, the excitations observed below 10 meV are identified to be the quantum rotational excitations of the water molecule. Surprisingly, the quantum rotational excitations of both para- and ortho-water molecule are observed simultaneously even for a single water molecule, indicating that the fluctuation between the ortho- and para-water states takes place in a time scale shorter than our measurement time (∼1 min), probably by the interaction between the encapsulated water molecule and conducting electrons.

Colibactin is a polyketide-nonribosomal peptide hybrid secondary metabolite that can form interstrand cross-links in double-stranded DNA. Colibactin-producing Escherichia coli has also been linked to colorectal oncogenesis. Thus, there is a strong interest in understanding the role colibactin may play in oncogenesis. Here, using the high-colibactin-producing wild-type E. coli strain we isolated from a clinical sample with the activity-based fluorescent probe we developed earlier, we were able to identify colibactin 770, which was recently identified and proposed as the complete form of colibactin, along with colibactin 788, 406, 416, 420, and 430 derived from colibactin 770 through structural rearrangements and solvolysis. Furthermore, we were able to trap the degrading mature colibactin species by converting the diketone moiety into quinoxaline in situ in the crude culture extract to form colibactin 860 at milligram scale. This allowed us to determine the stereochemically complex structure of the rearranged form of an intact colibactin, colibactin 788, in detail. Furthermore, our study suggested that we were capturing only a few percent of the actual colibactin produced by the microbe, providing a crude quantitative insight into the inherent instability of this compound. Through the structural assignment of colibactins and their degradative products by the combination of LC-HRMS and NMR spectroscopies, we were able to elucidate further the fate of inherently unstable colibactin, which could help acquire a more complete picture of colibactin metabolism and identify key DNA adducts and biomarkers for diagnosing colorectal cancer.

o -Toluidine ( o -Tol), a monocyclic aromatic amine, causes bladder cancer in humans and experimental animals and is therefore classified as a Group 1 carcinogen (IARC) in which the carcinogenicity of o -Tol is involved in metabolic activation, DNA damage, and DNA adduct formation. In the DNA adduct formation mechanism, o -Tol is metabolized by N -hydroxylation, N -acetoxylation, and then deacetoxylation to produce an electrophilic nitrenium ion, which is able to bind to a DNA base, such as dG-C8. Therefore, dG-C8- o -Tol is thought to be a plausible DNA adduct of o -Tol exposure. However, direct detection of dG-C8- o -Tol in biological samples has not been reported yet. Here, we show that a novel o -Tol metabolite, 2-methyl- N 1 -(2-methylphenyl)benzene-1,4-diamine (MMBD), a dimer by head-to-tail binding, was identified for the first time in o -Tol-exposed rat urine. MMBD was also detected in a reaction of o -Tol and S9 mix, indicating the formation was catalyzed by an enzymatic reaction. Moreover, MMBD showed a potent stronger mutagenicity in N -acetyltransferase overexpressed Salmonella typhimurium strains,and cytotoxicity in human bladder carcinoma T24 cells and human spleen lymphoblastoid TK6 cells compared with o -Tol. Furthermore, a DNA adduct ( m / z 478.1) corresponding to dG-MMBD was detected in the reaction of calf thymus DNA with rat urine containing MMBD, and also in hepatic DNA of rats treated with o -Tol. These results therefore suggested that o -Tol-induced bladder carcinogenesis could be at least partly attributed to MMBD formation. The possible dimerization of monocyclic aromatic amines should be considered in the evaluation of the risk of bladder carcinogenesis after exposure.

Single-crystal SnSe exhibits extremely high thermoelectric properties, and fabrication of SnSe films is promising for practical application and basic research on properties. However, the high thermoelectric properties have not yet been reported in SnSe films and their thermoelectric properties and nanostructure have not yet been analyzed in detail. In the present study, a -axis-oriented epitaxial SnSe films were prepared to discuss the thermoelectric properties of the SnSe films. While the electrical conductivity of the films was orders of magnitude smaller than that in the single crystals at room temperature, surprisingly, the thermoelectric property (power factor) of the films was slightly higher than that in the single crystals at high temperatures (∼300 °C). The SnSe films contained orthorhombic domain boundaries with a spacing of several hundred nanometers. The orthorhombic domain boundaries caused carrier scattering and degraded the mobility of the films at room temperature, but their effect decreased with increasing temperature. Thus, the carrier scattering at domain boundaries results in characteristic temperature dependence of thermoelectric properties in the SnSe films. High thermoelectric properties at high temperatures were successfully achieved in the SnSe films in spite of the existence of domain boundaries, demonstrating the possibility of high-performance of SnSe thermoelectric films.

While high-colibactin-producing Escherichia coli is thought to be associated with colorectal oncogenesis, this study is complicated part due to an inability to isolate colibactin adequately. Here, we created fluorescent probes activated by ClbP, the colibactin-maturing peptidase, to identify high-colibactin-producing strains. Our probe served as a valuable clinical diagnostic tool that allowed simple high-throughput diagnostic screening of clinical samples. Furthermore, the probe also allowed identification of high-colibactin producers that would help advance our understanding of colibactin biosynthesis.

5-HT 3 receptors are members of the Cys-loop family of ligand-gated ion channels, and, like most members of this family, there are multiple subunits that can contribute to functional pentameric receptors. 5-HT 3 A and 5-HT 3 AB receptors have been extensively characterized, but there are few studies on 5-HT 3 AC, 5-HT 3 AD, and 5-HT 3 AE receptors. Here we explore the properties of a range of partial agonists at 5-HT 3 AC, 5-HT 3 AD, and 5-HT 3 AE receptors following expression in Xenopus oocytes. The data show that the characteristics of receptor activation differ in the different heteromeric receptors when they are challenged with 5-HT, m-chlorophenylbiguanide (mCPBG), varenicline, 5-fluorotryptamine (5-FT), or thymol. 5-HT, 5-FT, varenicline, and mCPBG activation of 5-HT 3 AC, 5-HT 3 AD, and 5-HT 3 AE receptors yields similar EC 50 s to homomeric 5-HT 3 A receptors, but maximal responses differ. There are also differences in the levels of potentiation by thymol, which is greater at 5-HT 3 A receptors than 5-HT 3 AB, 5-HT 3 AC, 5-HT 3 AD, or 5-HT 3 AE receptors. Docking thymol into the receptor indicates a different residue in the transmembrane domain could provide an explanation for these data. Overall our study suggests that 5-HT 3 AC, 5-HT 3 AD, and 5-HT 3 AE have distinct pharmacological profiles to those of 5-HT 3 A and 5-HT 3 AB receptors; this is likely related to their distinct roles in the nervous system, consistent with their differential association with various disorders. Thus, these data pave the way for drugs that can specifically target these proteins.

To identify natural products and their associated biosynthetic genes from underutilized, difficult-to-manipulate microbes, chemical screening and bioinformatic analysis were employed to identify secondary metabolites and a potentially associated biosynthetic gene cluster. Subsequently, a heterologous expression system was used to confirm the identity of the gene cluster and the proposed biosynthetic mechanism. This approach successfully identified the curvupallide and spirostaphylotrichin biosynthetic pathways in endophytic fungus Curvularia pallescens and the short-chain pyranonigrin biosynthetic pathway in Aspergillus niger.

We present fractional quantum Hall domain walls confined in a gate-defined wire structure. Our experiments utilize spatial oscillation of domain walls driven by radio frequency electric fields to cause nuclear magnetic resonance. The resulting spectra are discussed in terms of both large quadrupole fields created around the wire and hyperfine fields associated with the oscillating domain walls. This provides the experimental fact that the domain walls survive near the confined geometry despite of potential deformation, by which a localized magnetic resonance is allowed in electrical means.

Graph data are becoming increasingly common in machine learning and data mining, and its application field pervades to bioinformatics and cheminformatics. Accordingly, as a method to extract patterns from graph data, graph mining recently has been studied and developed rapidly. Since the number of patterns in graph data is huge, a central issue is how to efficiently collect informative patterns suitable for subsequent tasks such as classification or regression. In this paper, we consider mining discriminative subgraphs from graph data with multiple labels. The resulting task has important applications in cheminformatics, such as finding common functional groups that trigger multiple drug side effects, or identifying ligand functional groups that hit multiple targets. In computational experiments, we first verify the effectiveness of the proposed approach in synthetic data, then we apply it to drug adverse effect prediction problem. In the latter dataset, we compared the proposed method with L1-norm logistic regression in combination with the PubChem/Open Babel fingerprint, in that the proposed method showed superior performance with a much smaller number of subgraph patterns. Software is available from https://github.com/axot/GLP.

A C29 phytoecdysteroid named amarasterone A (1) has been isolated from Cyathula capitata (Amaranthaceae), Leuzea carthamoides (Asteraceae), and Microsorum scolopendria (Polypodiaceae). We recently isolated amarasterone A from C. officinalis. Amarasterone A has been postulated as a biosynthetic intermediate of cyasterone in Cyathula sp. The stereochemistry at the C-24 and C-25 positions of these amarasterone A samples was investigated by comparing the NMR spectroscopic data with those of stereodefined model compounds, (24R,25S)-, (24R,25R)-, (24S,25S)-, and (24S,25R)-isomers of (20R,22R)-3β-methoxystigmast-5-ene-20,22,26-triol (2a-d), which were synthesized in the present study. Amarasterone A isolated from Cyathula officinalis was determined to be the (24R,25S)-isomer (1a), while amarasterone A from L. carthamoides was found to be the (24R,25R)-isomer (1b). Amarasterone A from M. scolopendria was found to be a mixture of 1a and 1b. The biosynthesis of cyasterone in Cyathula sp. is discussed on the basis of the identical C-24 configuration of sitosterol and amarasterone A.

Aplyronine A (ApA) is a marine natural product that shows potent antitumor activity. While both ApA and ApC, a derivative of ApA that lacks a trimethylserine ester moiety, inhibit actin polymerization in vitro to the same extent, only ApA shows potent cytotoxicity. Therefore, the molecular targets and mechanisms of action of ApA in cells have remained unclear. We report that ApA inhibits tubulin polymerization in a hitherto unprecedented way. ApA forms a 1:1:1 heterotrimeric complex with actin and tubulin, in association with actin synergistically binding to tubulin, and inhibits tubulin polymerization. Tubulin-targeting agents have been widely used in cancer chemotherapy, but there are no previous descriptions of microtubule inhibitors that also bind to actin and affect microtubule assembly. ApA inhibits spindle formation and mitosis in HeLa S3 cells at 100 pM, a much lower concentration than is needed for the disassembly of the actin cytoskeleton. The results of the present study indicate that ApA represents a rare type of natural product, which binds to two different cytoplasmic proteins to exert highly potent biological activities.

The antitumor and apoptogenic macrolide aplyronine A (ApA) is a potent actin-depolymerizing agent. We developed an ApA acetylene analog that bears the aryldiazirine group at the C34 terminus, which formed a covalent bond with actin. With the use of the photoaffinity biotin derivatives of aplyronines A and C, Arp2 and Arp3 (actin-related proteins) were specifically purified as binding proteins along with actin from tumor cell lysate. However, Arp2 and Arp3 did not covalently bind to aplyronine photoaffinity derivatives. Thus, actin-related proteins might indirectly bind to ApA as the ternary adducts of the actin/ApA complex or through the oligomeric actin.

A hybrid compound consisting of aplyronine A and mycalolide B was synthesized, and its biological activities were evaluated. The hybrid compound was found to have somewhat more potent actin-depolymerizing activity than aplyronine A. In contrast, the hybrid compound possessed about 1000-fold less cytotoxicity than aplyronine A. These results indicated that there is no direct correlation between actin-depolymerizing activity and cytotoxicity., (© 2012 American Chemical Society)

New aliphatic polyester-type dendrimers were synthesized using a new AB2-type building block 3, prepared from benzyl acetoacetate and 2 equiv of tert-butyl acrylate by acetoacetic acid ester synthesis. The reiterative [deprotection by HCO2H, then EDCI/DMAP coupling] sequence using divergent growth method gave [G1]-4tBu-[G5]-64tBu dendrimers. 13C NMR relaxation time (T1) measurements on the carboxy carbons show that the extended chain conformations are predominant in CDCl3. [structure: see text]

[see structure]. Synthesis and photochemistry of a new photochemically removable protecting group for alcohols is described. Four carbonates of galactose derivatives (1-4) were synthesized from the corresponding arylmethanols via 4-nitrophenyl carbonate intermediates. Among them, photolysis of anthraquinon-2-ylmethoxycarbonyl (Aqmoc) galactose (1) proceeded with overall photolysis efficiency of 150 (quantum yield 0.10, and molar absorptivity 1500 M(-1) x cm(-1)) and rate constant of approximately 10(6) s(-1). To demonstrate its application to a biologically related molecule, 5'-Aqmoc-adenosine (5) was synthesized and photolyzed to yield adenosine in 91% yield.

Differential scanning calorimetry (DSC) was performed to investigate thermodynamic properties of three carp fast skeletal light meromyosin (LMM) isoforms expressed in Escherichia coli by recombinant DNAs. Three isoforms were the 10 degreesC-, intermediate-, and 30 degreesC-type LMM predominantly expressed in carp acclimated to 10, 20, and 30 degreesC. The isoforms expressed in E. coli by recombinant DNAs exhibited a typical pattern of alpha-helix in CD spectroscopy with two minima at 222 and 208 nm. Moreover, the three isoforms formed paracrystals typical of LMM, suggesting that expressed proteins retained intact structural properties. When the LMM isoforms were subjected to DSC analysis, the 10 degreesC and 30 degreesC types showed endotherms having transition temperatures (Tm) at 35.1 and 39.5 degreesC, respectively, which are responsible for thermal unfolding of alpha-helix. The intermediate type exhibited two comparable endotherms with Tm values at 34.9 and 40.6 degreesC, implying that it has intermediate thermodynamic properties between those of 10 degreesC and 30 degreesC types. However, a chimeric LMM having the 10 degreesC and 30 degreesC type as N- and C-terminal halves, respectively, showed the DSC pattern typical of the whole 30 degreesC-type molecule. On the other hand, another chimeric LMM composed of the N-terminal 30 degreesC type and C-terminal 10 degreesC type gave the pattern of the full 10 degreesC type. These results suggest that thermodynamic properties of the C-terminal half largely account for thermal unfolding of the whole molecule.