Because the influenza A virus has an RNA genome, its RNAdependent RNA polymerase, comprising the PA, PB1, and PB2 subunits, is essential for viral transcription and replication. The binding of RNA primers/promoters to the polymerases is an initiation step in viral transcription. In our current study, we reveal the 2.7 Å tertiary structure of the C-terminal RNA-binding domain of PB2 by x-ray crystallography. This domain incorporates lysine 627 of PB2, and this residue is associated with the high pathogenicity and host range restriction of influenza A virus. We found from our current analyses that this lysine is located in a unique ""-shaped structure consisting of a helix and an encircled loop within the PB2 domain. By electrostatic analysis, we identified a highly basic groove along with this loop and found that lysine 627 is located in the loop. A PB2 domain mutant in which glutamic acid is substituted at position 627 shows significantly lower RNA binding activity. This is the first report to show a relationship between RNA binding activity and the pathogenicity-determinant lysine 627. Using the Matras program for protein three-dimensional structural comparisons, we further found that the helix bundles in the PB2 domain are similar to that of activator 1, the 40-kDa subunit of DNA replication clamp loader (replication factor C), which is also an RNA-binding protein. This suggests a functional and structural relationship between the RNA-binding mechanisms underlying both influenza A viral transcription and cellular DNA replication. Our present results thus provide important new information for developing novel drugs that target the primer/promoter RNA binding of viral RNA polymerases.

CS-1008 has a selective toxicity toward tumor cells expressing DR5 and the potential for antitumor efficacy in human malignancies.

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In this article, 35 published studies on life cycle assessment (LCA) of sewage sludge were reviewed for their methodological and technological assumptions. Overall, LCA has been providing a flexible framework to quantify environmental impacts of wastewater and sewage sludge treatment and disposal processes for multiple scales, ranging from process selection to policy evaluation. The results of LCA are, in principle, unique to the goal and scope of each study, reflecting its local conditions and comparison between different LCAs is not intended. Furthermore, the assessments are limited by the methodological development of the life cycle impact assessment (LCIA) and the advancement of research in quantifying environmental emissions associated with wastewater and sewage sludge treatment processes. Thus, large discrepancies were found in the selection of the environmental emissions to be included and how they were estimated in the analysis. In order to reduce these choice uncertainties, consolidation of the modelling approach in the following area are recommended: quantification of fugitive gas emissions and modelling of disposal practices. Besides harmonization of the key technical assumptions, clear documentation of the modelling approach and the uncertainties associating with each assumption is encouraged so as to improve the integrity and robustness of assessment.

In the presence of native DNA the hydrolysis of benzo[a]pyrene-7,8-diol 9,10-epoxide (BPDE) to tetrols (BPT) is markedly accelerated (by a factor of up to approximately 80 at 25 degrees C, pH 7.0, in 5 mM sodium cacodylate buffer solution). When stopped-flow kinetic techniques are utilized, it is shown that the pseudo-first-order hydrolysis rate constant kH is smaller by a factor of approximately 3 in the presence of equivalent concentrations of denatured DNA, by a factor of 8-25 in the presence of nucleotides, and by a factor of 35-45 in the presence of nucleosides (depending on the nucleotide or nucleoside). In the presence of native DNa, kH increases with increasing DNA concentration and reaches a limiting value of kH = 0.684 +/- 0.04 s-1 at DNA concentrations in excess of approximately 5 x 10(-4) M (expressed in concentration of nucleotides). A kinetic model based on (1) rapid formation of a noncovalent BPDE-DNA complex followed by (2) slower hydrolysis of BPDE to BPT at these binding sites is consistent with the experimental data. It is shown furthermore that the DNA concentration dependence of kH and of noncovalent intercalative binding of BPDE to DNA is similar and that addition of magnesium ions (which is known to reduce intercalative binding of planar aromatic molecules to DNA) also reduces kH. These results suggest, but do not necessarily prove, that the DNA binding sites at which the hydrolysis of BPDE (to BPT) is catalyzed are intercalative in nature.