A possible model of benzimidazole binding to beta-tubulin disclosed by invoking an inter-domain movement.

Although it is well established that benzimidazole (BZMs) compounds exert their therapeutic effects through binding to helminth beta-tubulin and thus disrupting microtubule-based processes in the parasites, the precise location of the benzimidazole-binding site on the beta-tubulin molecule has yet to be determined. In the present study, we have used previous experimental data as cues to help identify this site. Firstly, benzimidazole resistance has been correlated with a phenylalanine-to-tyrosine substitution at position 200 of Haemonchus contortus beta-tubulin isotype-I. Secondly, site-directed mutagenesis studies, using fungi, have shown that other residues in this region of the protein can influence the interaction of benzimidazoles with beta-tubulin. However, the atomic structure of the alphabeta-tubulin dimer shows that residue 200 and the other implicated residues are buried within the protein. This poses the question: how might benzimidazoles interact with these apparently inaccessible residues? In the present study, we present a mechanism by which those residues generally believed to interact with benzimidazoles may become accessible to the drugs. Furthermore, by docking albendazole-sulphoxide into a modelled H. contortus beta-tubulin molecule we offer a structural explanation for how the mutation conferring benzimidazole resistance in nematodes may act, as well as a possible explanation for the species-specificity of benzimidazole anthelmintics.

Nitric Oxide Produced in Response to Engagement of beta 2 Integrins on Human Neutrophils Activates the Monomeric GTPases Rap1 and Rap2 and Promotes Adhesion

We found that engagement of beta 2 integrins on human neutrophils increased the levels of GTP-bound Rap1 and Rap2. Also, the activation of Rap1 was blocked by PP1, SU6656, LY294002, GF109203X, or BAPTA-AM, which indicates that the downstream signaling events in Rap1 activation involve Src tyrosine kinases, phosphoinositide 3-kinase, protein kinase C, and release of calcium. Surprisingly, the integrin-induced activation of Rap2 was not regulated by any of the signaling pathways mentioned above. However, we identified nitric oxide as the signaling molecule involved in beta 2 integrin-induced activation of Rap1 and Rap2. This was illustrated by the fact that engagement of beta 2 integrins increased the production of nitrite, a stable end-product of nitric oxide. Furthermore, pretreatment of neutrophils with N-monomethyl-L-arginine, or 1400W, which are inhibitors of inducible nitric-oxide synthase, blocked integrin-induced activation of Rap1 and Rap2. Similarly, Rp-8pCPT-cGMPS, an inhibitor of cGMP-dependent serine/threonine kinases, also blunted the integrin-induced activation of Rap GTPases. Also nitric oxide production and its downstream activation of cGMP-dependent serine/threonine kinases were essential for proper neutrophil adhesion by beta 2 integrins. Thus, we made the novel findings that beta 2 integrin engagement on human neutrophils triggers production of nitric oxide and its downstream signaling is essential for activation of Rap GTPases and neutrophil adhesion.

Post-depositional modification of atmospheric dust on a granite building in central Rio de Janeiro: implications for surface induration and subsequent stone decay

Extensive contour scaling of a 200 year old granite church is associated with the breaching of an apparently iron-rich crust and the widespread deposition of atmospheric dust within the canyon-like streetscape of Rio de Janeiro. Contemporary dust, accumulated dust from within the a depression on the building surface, the surface crust and the underlying granite are examined by a combination of total element analysis and sequential extraction, X-ray diffraction and energy dispersive X-ray fluorescence. Results indicate an increase in total organic carbon and a marked decrease in pH within the accumulated dust, and a rapid mobilization of anions and cations from the water-soluble and carbonate phases. It is considered that the latter is linked to salt accumulation within and eventual salt weathering of the granite. Post-depositional alteration of the dust is also linked with the de-silicification of clay minerals (Illite to kaolinite) and the loss of silica from the amorphous Fe/Mn phase of the accumulated dust under the initially saline and progressively more acidic conditions experienced at the stone - atmosphere interface. This mobilization of silica is associated with the formation of what is, in effect, a thin silica-rich surface crust or glaze. Within the glaze, assessory amounts of extractable iron are concentrated within the amorphous and crystalline Fe/Mn phases at levels that are significantly elevated with respect to the underlying granite, but much lower than the equivalent phases of the accumulated dust from which it is principally assumed to derive. The protection afforded to the stone work by the crust is not, however, permanent and within the last 15 years it has been possible to observe a rapid increase in the surface delamination of the church close to street level.

Assembly intermediates in polyketide biosynthesis; enantioselective syntheses of beta-hydroxycarbonyl compounds

A versatile approach for the enantioselective synthesis of functionalised beta-hydroxy N-acetylcysteamine thiol esters has been developed which allows the facile incorporation of isotopic labels. It has been shown that a remarkable reversal of selectivity occurs in the titanium mediated aldol reaction of the acyloxazolidone intermediate using either (S)- or (R)-tert-butyldimethylsilyloxybutanal. The aldol products are valuable intermediates in the synthesis of 4-hydroxy-6-substituted gamma-lactones.

Inactivation of Human beta-Defensins 2 and 3 by Elastolytic Cathepsins

beta-Defensins are antimicrobial peptides that contribute to the innate immune responses of eukaryotes. At least three defensins, human beta-defensins 1, 2, and 3 (HBD-1, -2, and -3), are produced by epithelial cells lining the respiratory tract and are active toward Gram-positive (HBD-3) and Gram-negative (HBD-1, -2, and -3) bacteria. It has been postulated that the antimicrobial activity of defensins is compromised by changes in airway surface liquid composition in lungs of patients with cystic fibrosis (CF), therefore contributing to the bacterial colonization of the lung by Pseudomonas and other bacteria in CF. In this report we demonstrate that HBD-2 and HBD-3 are susceptible to degradation and inactivation by the cysteine proteases cathepsins B, L, and S. In addition, we show that all three cathepsins are present and active in CF bronchoalveolar lavage. Incubation of HBD-2 and -3 with CF bronchoalveolar lavage leads to their degradation, which can be completely (HBD-2) or partially (HBD-3) inhibited by a cathepsin inhibitor. These results suggest that beta-defensins are susceptible to degradation and inactivation by host proteases, which may be important in the regulation of beta-defensin activity. In chronic lung diseases associated with infection, overexpression of cathepsins may lead to increased degradation of HBD-2 and -3, thereby favoring bacterial infection and colonization.

Theorising European Integration: Revisiting Neo-Functionalism and Testing its Suitability for Explaining the Development of EC Competition Policy?

This paper was published in the highly respected, peer reviewed and ISI ranked journal - 'European Integration on-line paper series

Development of a rapid screening test for veterinary sedatives and the beta-blocker carazolol in porcine kidney by ELISA

Sedatives and tranquillisers are frequently used to reduce stress during the transportation of food producing animals. The most widely used classes of sedatives include the butyrophenone azaperone, the phenothiazines acepromazine, propionylpromazine, chlorpromazine and the beta-blocker, carazolol. For regulatory control purposes, tolerances for azaperone and carazolol have been set by the European Union as 100 and 25 mug kg(-1), respectively. Furthermore, the use of the phenothiazines is prohibited and therefore has a zero tolerance. A method for the detection of residues of five tranquillisers and one beta-blocker using a single ELISA plate has been developed. Kidney samples (2.5 g) were extracted with dichloromethane and applied to a competitive enzyme immunoassay using three polyclonal antibodies raised in rabbits against azaperol, propionylpromazine and carazolol conjugates. In sample matrix, the azaperol antibody cross-reacted 28.0% with azaperone and the propionylpromazine antibody cross-reacted 24.9% with acepromazine and 11.7% with chlorpromazine. In the ELISA, the detection capabilities of the six sedatives, azaperol, azaperone, carazolol, acepromazine, chlorpromazine, and propionylpromazine are 5, 15, 5, 5, 20 and 5 mug kg(-1), respectively. The proposed method is a sensitive and rapid multi-residue technique that offers a cost effective alternative to current published procedures, without any concession on the ability to detect sedative misuse.