Effects of 1-ethynylpyrene and related inhibitors of P450 isozymes upon benzo[a]pyrene metabolism by liver microsomes

The effects of three aryl acetylenes, 1-ethynylpyrene (EP), 2-ethynylnaphthalene (EN) and 3-ethynylperylene (EPE), upon the metabolism of benzo[a]pyrene (BaP) by microsomes isolated from rat liver were investigated. These aryl acetylenes all inhibited the total metabolism of BaP. Formation of BaP 7,8-dihydrodiol and BaP tetrol products by microsomal preparations from rats that had been pretreated with 3-methylcholanthrene (3MC) were preferentially inhibited. The effects of EP upon the metabolism of BaP 7,8-dihydrodiol by microsomes from rat liver were also studied. This aryl acetylene strongly inhibited the formation of BaP tetrols from BaP 7,8-dihydrodiol by liver microsomes both from untreated rats and from rats pretreated with 3MC, but enhanced the conversion of the BaP dihydrodiol into other metabolites.     

Cyclo(His-Pro) up-regulates heme oxygenase 1 via activation of Nrf2-ARE signalling

Paraquat (1,1'-dimethyl-4,4'-bipyridinium), a widely used non-selective herbicide, is a redox cycling agent with adverse effects on dopamine systems. Epidemiological data have shown that exposure to paraquat is one of the several risk factors for Parkinson's disease. We have already shown that cyclo(His-Pro), an endogenous cyclic dipeptide produced by the cleavage of the thyrotropin releasing hormone, has a cytoprotective effect through a mechanism involving Nrf2 activation that decreases production of reactive oxygen species and increases glutathione synthesis. Using primary neuronal cultures and PC12 cells as targets of paraquat neurotoxicity, we addressed whether and how cyclo(His-Pro) causes cellular protective response against paraquat-mediated cell death. We found that cyclo(His-Pro) attenuated reactive oxygen species production, and prevented glutathione depletion by up-regulating Nrf2 gene expression, triggering its nuclear accumulation and activating the expression of heme oxygenase1. These protective effects were abolished by RNA interference-mediated Nrf2 knock down whereas were unaffected by RNA interference-mediated Keap1 knock down. Inhibition of heme oxygenase activity decreased cyclo(His-Pro)-induced neuroprotection. These results suggest that cyclo(His-Pro), acting as a selective activator of the brain modulable Nrf2 pathway, may be a promising candidate as neuroprotective agent that act through induction of phase II genes.     

Molecular Probing of the HPV-16 E6 Protein Alpha Helix Binding Groove with Small Molecule Inhibitors

The human papillomavirus (HPV) HPV E6 protein has emerged as a central oncoprotein in HPV-associated cancers in which sustained expression is required for tumor progression. A majority of the E6 protein interactions within the human proteome use an alpha-helix groove interface for binding. The UBE3A/E6AP HECT domain ubiquitin ligase binds E6 at this helix-groove interface. This enables formation of a trimeric complex with p53, resulting in destruction of this tumor suppressor. While recent x-ray crystal structures are useful, examples of small molecule probes that can modulate protein interactions at this interface are limited. To develop insights useful for potential structure-based design of ligands for HPV E6, a series of 2,6-disubstituted benzopyranones were prepared and tested as competitive antagonists of E6-E6AP helix-groove interactions. These small molecule probes were used in both binding and functional assays to evaluate recognition features of the E6 protein. Evidence for an ionic functional group interaction within the helix groove was implicated by the structure-activity among the highest affinity ligands. The molecular topographies of these protein-ligand interactions were evaluated by comparing the binding and activities of single amino acid E6 mutants with the results of molecular dynamic simulations. A group of arginine residues that form a rim-cap over the E6 helix groove offer compensatory roles in binding and recognition of the small molecule probes. The flexibility and impact on the overall helix-groove shape dictated by these residues offer new insights for structure-based targeting of HPV E6.     

Inactivation and oxidative modification of Cu,Zn superoxide dismutase by stimulated neutrophils: the appearance of new catalytically active structures.

Bovine superoxide dismutase (SOD) was inactivated during incubation with phorbol myristate acetate-stimulated neutrophils. In addition, stimulated neutrophils were able to disrupt the SOD structure. Inactivation and structural damage were dependent on the action of hypochlorous acid, an oxidant generated by the myeloperoxidase-hydrogen peroxide-chloride system of neutrophils. Incubation of SOD with stimulated neutrophils lead to long-wavelength fluorescence (ex, 350 nm; em, 450 nm) and the appearance of new structural forms with other isoelectric points. These additional forms possess catalytic activity. Generation of catalytically active new forms of SOD demonstrates the inaccessibility of the active centre of SOD to hypochlorite and may be a reason for the successful application of SOD during anti-inflammatory therapy.     

Species loss leads to community closure

Global extinction of a species is sadly irreversible. At a local scale, however, extinctions may be followed by re-invasion. We here show that this is not necessarily the case and that an ecological community may close its doors for re-invasion of species lost from it. Previous studies of how communities are assembled have shown that there may be rules for that process and that limitations are set to the order by which species are introduced and put together. Instead of focusing on the assembly process we randomly generated simple competitive model communities that were stable and allowed for two to 10 coexisting species. When a randomly selected single species was removed from the community, the cascading species loss was recorded and frequently the resulting community was more than halved. Cascading extinctions have previously been recorded, but we here show that the relative magnitude of the cascade is dependent on community size (and not only trophic structure) and that the reintroduction of the original species lost often is impossible. Hence, species loss does not simply leave a void potentially refilled, but permanently alters the entire community structure and consequently the adaptive landscape for potential re-invaders.