Structure of hepatitis C virus polymerase in complex with primer-template RNA

The replication of the hepatitis C viral (HCV) genome is accomplished by the NS5B RNA-dependent RNA polymerase (RdRp), for which mechanistic understanding and structure-guided drug design efforts have been hampered by its propensity to crystallize in a closed, polymerization-incompetent state. The removal of an autoinhibitory β-hairpin loop from genotype 2a HCV NS5B increases de novo RNA synthesis by >100-fold, promotes RNA binding, and facilitated the determination of the first crystallographic structures of HCV polymerase in complex with RNA primer-template pairs. These crystal structures demonstrate the structural realignment required for primer-template recognition and elongation, provide new insights into HCV RNA synthesis at the molecular level, and may prove useful in the structure-based design of novel antiviral compounds. Additionally, our approach for obtaining the RNA primer-template-bound structure of HCV polymerase may be generally applicable to solving RNA-bound complexes for other viral RdRps that contain similar regulatory β-hairpin loops, including bovine viral diarrhea virus, dengue virus, and West Nile virus.     

Tapping the archives: the sterol composition of marine sponge species,as determined non‐invasively from museum‐preserved specimens,reveals biogeographical features

Over 8600 species are currently recorded in the phylum Porifera (sponges). They produce a large diversity of biochemical compounds including sterols, with more than 250 different sterols identified. Some of these sterols are of great interest, due to their use for fingerprinting in ecological and biomarker (molecular fossil) studies. As a large number of identified extant species from biodiversity surveys are housed in museum collections, preserved in ethanol, these present a potentially rich source of identified specimens for comparative lipid analyses. Here, we show that, in at least one species, sterol distributions obtained from the ethanol used to preserve specimens of sponges were representative, and comparable to the sterol distribution obtained from wet‐frozen and from freeze‐dried tissue from the same species. We employed both GC‐MS and two‐dimensional gas chromatography—time‐of‐flight mass spectrometry (GC×GC‐TOFMS), with an improved signal‐to‐noise ratio for even minor constituents. Analysis of two additional specimens of the same species, but of different provenance, resulted in detection of marked differences in sterol composition, which could be attributed to variations in geography, environmental conditions, microbial communities, diet or cryptic speciation. The possibility of using ethanol from identified, preserved museum sponges could drastically increase the number of available samples. This could enable the study of their sterol complements, and the detailed investigation of differences due to geographical and oceanographic, phylogenetic, and other factors in unprecedented detail.     

Novel 5-arylthio-5H-chromenopyridines as a new class of anti-fibrotic agents

Liver fibrosis is a critical wound healing response to chronic liver injury such as hepatitis C virus (HCV) infection. If persistent, liver fibrosis can lead to cirrhosis and hepatocellular carcinoma (HCC). The development of new therapies for preventing liver fibrosis and its progression to cancer associated with HCV infection remains a critical challenge. Identification of novel anti-fibrotic compounds will provide opportunities for innovative therapeutic intervention of HCV-mediated liver fibrosis. We designed and synthesized a focused set of 5-arylthio-5H-chromenopyridines as a new class of anti-fibrotic agents. Liver fibrosis assays demonstrated that the compounds 3a and 3c show inhibitory activity towards human hepatic stellate cells (LX2) activation at 10 μM. The HCV NS3 and NS5A proteins in HCV subgenome-expressing cells were also significantly reduced in cells treated with 3a and 3c, suggesting the possible inhibitory role of the compounds in HCV translation/replication activities. We have also examined the reactivity of these compounds with medicinally-relevant metal compounds such as platinum and gold. The reactivity of these complexes with metals and during Mass Spectrometry suggests that CS bond cleavage is relatively facile.