Oxazolones as potent inhibitors of 11beta-hydroxysteroid dehydrogenase type 1

2,5,5-Trisubstituted oxazolones were identified as potent inhibitors of 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1). The synthesis, structure-activity relationship and metabolic stability of these compounds are presented.     

Host-parasite kinship in a female-philopatric bird population: evidence from relatedness trend analysis

Conspecific brood parasitism (CBP), an alternative reproductive tactic where some females lay eggs in the nests of other females of the same species, occurs in many animals with egg care. It is particularly common in waterfowl, for reasons that are debated. Many waterfowl females nest near their birthplace, making it likely that some local females are relatives. We analyse brood parasitism in a Hudson Bay population of common eiders, testing predictions from two alternative hypotheses on the role of relatedness in CBP. Some models predict host-parasite relatedness, others predict that parasites avoid close relatives as hosts. To distinguish between the alternatives, we use a novel approach, where the relatedness of host-parasite pairs is tested against the spatial population trend in pairwise relatedness. We estimate parasitism, nest take-over and relatedness with protein fingerprinting and bandsharing analysis of egg albumen, nondestructively sampled from each new egg in the nest throughout the laying period. The results refute the hypothesis that parasites avoid laying eggs in the nests of related hosts, and corroborate the alternative of host-parasite relatedness. With an estimated r of 0.12-0.14, females laying eggs in the same nest are on average closer kin than nesting neighbour females. Absence of a population trend in female pairwise relatedness vs. distance implies that host-parasite relatedness is not only an effect of strong natal philopatry: some additional form of kin bias is also involved.     

Inhibitors of type III secretion in Yersinia: design, synthesis and multivariate QSAR of 2-arylsulfonylamino-benzanilides

Compound 1, 2-(benzo[1,2,5]thiadiazole-4-sulfonylamino)-5-chloro-N-(3,4-dichloro-phenyl)-benzamide, was identified as a putative type III secretion inhibitor in Yersinia, and the compound thus has a potential to be used to prevent or treat bacterial infections. A set of seven analogues was synthesized and evaluated in a type III secretion dependent reporter-gene assay with viable bacterial to give basic SAR. A second set of 19 compounds was obtained by statistical molecular design in the building block and product space and by subsequent synthesis. Evaluation in the reporter-gene assay showed that the compounds ranged from non-active to compounds more potent than 1. Based on the data multivariate QSAR models were established and the final Hi-PLS model showed good correlation between experimentally determined % inhibition and the calculated % inhibition of the reporter-gene signal.     

Clavulanic acid dehydrogenase: structural and biochemical analysis of the final step in the biosynthesis of the beta-lactamase inhibitor clavulanic acid

The ultimate step in the biosynthesis of the medicinally important beta-lactamase inhibitor clavulanic acid is catalyzed by clavulanic acid dehydrogenase (CAD). CAD is responsible for the NAPDH-dependent reduction of the unstable intermediate clavulanate-9-aldehyde to yield clavulanic acid. Here, we report biochemical and structural studies on CAD. Biophysical analyses demonstrate that CAD exists as dimeric and tetrameric species in solution. The reaction performed by CAD was shown to be reversible, allowing the use of clavulanic acid for activity analyses. The crystal structure of CAD was solved using single-wavelength anomalous diffraction with a seleno-methionine derivative. The structure reveals that the individual monomers comprise a single domain possessing the Rossmann fold, characteristic of dinucleotide-binding enzymes. The monomers are arranged as tetramers, similar to other tetrameric members of the short-chain dehydrogenase/reductase family. The structure of the unreactive complex of CAD with clavulanic acid and NADPH suggests how CAD is able to catalyze the reduction of clavulanate-9-aldehyde without fragmentation of the bicyclic beta-lactam ring structure. The relative positions of NADPH and clavulanic acid, in the active site, together with the presence of the latter in an eclipsed conformation, rationalizes previous labeling studies demonstrating that the incorporation of the C5 pro-R, but not pro-S, hydrogen of ornithine/arginine into the C9 position of clavulanic acid occurs with overall inversion of configuration.     

A cluster of carboxylic groups in PsbO protein is involved in proton transfer from the water oxidizing complex of Photosystem II

The hypothesis presented here for proton transfer away from the water oxidation complex of Photosystem II (PSII) is supported by biochemical experiments on the isolated PsbO protein in solution, theoretical analyses of better understood proton transfer systems like bacteriorhodopsin and cytochrome oxidase, and the recently published 3D structure of PS II (Pdb entry 1S5L). We propose that a cluster of conserved glutamic and aspartic acid residues in the PsbO protein acts as a buffering network providing efficient acceptors of protons derived from substrate water molecules. The charge delocalization of the cluster ensures readiness to promptly accept the protons liberated from substrate water. Therefore protons generated at the catalytic centre of PSII need not be released into the thylakoid lumen as generally thought. The cluster is the beginning of a localized, fast proton transfer conduit on the lumenal side of the thylakoid membrane. Proton-dependent conformational changes of PsbO may play a role in the regulation of both supply of substrate water to the water oxidizing complex and the resultant proton transfer.     

Traces of copper ions deplete glutathione in human hepatoma cell cultures with low cysteine content

BACKGROUND: Cell death induced by intracellular glutathione depletion has been reported to be dependent on the presence of trace amounts of extracellular copper ions. Since little is known about the relationship between glutathione depletion and copper homeostasis, we have in the present study further investigated the role of low amounts of copper ions in glutathione depletion. METHODS: Glutathione turnover was investigated in HeLa and hepatoma cell cultures with normal and low cysteine content in the presence of copper ions (1 and 10micromol/L) and two other glutathione-stimulating agents (lipoic acid and mercury ions). RESULTS: Copper ions (10micromol/L) caused relatively small increases in total amount of glutathione (the sum of the intracellular and the extracellular amount of glutathione) in HeLa and hepatoma cell cultures with normal cysteine levels (420nmol/mL) compared to control cell cultures, whereas lipoic acid and mercury ions strongly increased total glutathione in both types of cell cultures. Lower amount of total glutathione was observed in cell cultures with a lower cysteine levels (84nmol/mL), which is similar to that in human plasma. A strongly decreased total amount of glutathione in the presence of copper ions was observed in hepatoma cell cultures with lower cysteine levels, whereas the other agents showed effects similar to those described for cell cultures with normal cysteine levels. CONCLUSION: Glutathione synthesis in hepatoma cell cultures is probably more sensitive to a low cysteine level than HeLa cell cultures, and the presence of copper ions further decreases the availability of cysteine probably by increasing the disulfide binding to cysteine residues in extracellular proteins, which causes a further decrease of total glutathione.     

Small potent ligands to the insulin-regulated aminopeptidase (IRAP)/AT(4) receptor.

Angiotensin IV analogs encompassing aromatic scaffolds replacing parts of the backbone of angiotensin IV have been synthesized and evaluated in biological assays. Several of the ligands displayed high affinities to the insulin-regulated aminopeptidase (IRAP)/AT(4) receptor. Displacement of the C-terminal of angiotensin IV with an o-substituted aryl acetic acid derivative delivered the ligand 4, which exhibited the highest binding affinity (K(i) = 1.9 nM). The high affinity of this ligand provides support to the hypothesis that angiotensin IV adopts a gamma-turn in the C-terminal of its bioactive conformation.Ligand (4) inhibits both human IRAP and aminopeptidase N-activity and induces proliferation of adult neural stem cells at low concentrations. Furthermore, ligand 4 is degraded considerably more slowly in membrane preparations than angiotensin IV. Hence, it might constitute a suitable research tool for biological studies of the (IRAP)/AT(4) receptor.     

Internet‐delivered psychological treatments: from innovation to implementation

Internet interventions, and in particular Internet‐delivered cognitive behaviour therapy (ICBT), have existed for at least 20 years. Here we review the treatment approach and the evidence base, arguing that ICBT can be viewed as a vehicle for innovation. ICBT has been developed and tested for several psychiatric and somatic conditions, and direct comparative studies suggest that therapist‐guided ICBT is more effective than a waiting list for anxiety disorders and depression, and tends to be as effective as face‐to‐face CBT. Studies on the possible harmful effects of ICBT are also reviewed: a significant minority of people do experience negative effects, although rates of deterioration appear similar to those reported for face‐to‐face treatments and lower than for control conditions. We further review studies on change mechanisms and conclude that few, if any, consistent moderators and mediators of change have been identified. A recent trend to focus on knowledge acquisition is considered, and a discussion on the possibilities and hurdles of implementing ICBT is presented. The latter includes findings suggesting that attitudes toward ICBT may not be as positive as when using modern information technology as an adjunct to face‐to‐face therapy (i.e., blended treatment). Finally, we discuss future directions, including the role played by technology and machine learning, blended treatment, adaptation of treatment for minorities and non‐Western settings, other therapeutic approaches than ICBT (including Internet‐delivered psychodynamic and interpersonal psychotherapy as well as acceptance and commitment therapy), emerging regulations, and the importance of reporting failed trials.     

Exploring a Pool‐seq‐only approach for gaining population genomic insights in nonmodel species

Developing genomic insights is challenging in nonmodel species for which resources are often scarce and prohibitively costly. Here, we explore the potential of a recently established approach using Pool‐seq data to generate a de novo genome assembly for mining exons, upon which Pool‐seq data are used to estimate population divergence and diversity. We do this for two pairs of sympatric populations of brown trout (Salmo trutta): one naturally sympatric set of populations and another pair of populations introduced to a common environment. We validate our approach by comparing the results to those from markers previously used to describe the populations (allozymes and individual‐based single nucleotide polymorphisms [SNPs]) and from mapping the Pool‐seq data to a reference genome of the closely related Atlantic salmon (Salmo salar). We find that genomic differentiation (F_ST) between the two introduced populations exceeds that of the naturally sympatric populations (F_ST = 0.13 and 0.03 between the introduced and the naturally sympatric populations, respectively), in concordance with estimates from the previously used SNPs. The same level of population divergence is found for the two genome assemblies, but estimates of average nucleotide diversity differ ( ≈ 0.002 and  ≈ 0.001 when mapping to S. trutta and S. salar, respectively), although the relationships between population values are largely consistent. This discrepancy might be attributed to biases when mapping to a haploid condensed assembly made of highly fragmented read data compared to using a high‐quality reference assembly from a divergent species. We conclude that the Pool‐seq‐only approach can be suitable for detecting and quantifying genome‐wide population differentiation, and for comparing genomic diversity in populations of nonmodel species where reference genomes are lacking.