Structure–activity relationship and improved hydrolytic stability of pyrazole derivatives that are allosteric inhibitors of West Nile Virus NS2B-NS3 proteinase

West Nile Virus (WNV) is a potentially deadly mosquito-borne flavivirus which has spread rapidly throughout the world. Currently there is no effective vaccine against flaviviral infections. We previously reported the identification of pyrazole ester derivatives as allosteric inhibitors of WNV NS2B-NS3 proteinase. These compounds degrade rapidly in pH 8 buffer with a half life of 1–2 h. We now report the design, synthesis and in vitro evaluation of pyrazole derivatives that are inhibitors of WNV NS2B-NS3 proteinase with greatly improved stability in the assay medium.     

Comparison of Post-Mine Rehabilitated and Natural Shrubland Communities in Southwestern Australia

Following mineral sand mining near Eneabba, southwestern Australia, rehabilitation managers have the difficult task of restoring shrubland communities of exceptional plant species richness. Species diversity, composition, structure, and key functional attributes in four mined sites rehabilitated 8 (R8) to 24 (R24) years ago were compared with those of typical nearby natural areas classified on the basis of substrate type (Low and High sand Dunes, shallow sand Swales, sand over Laterite, and sand over Limestone). The rehabilitated sites (except R8) had more species (about 140) than natural sites (about 100) in 40 × 40–m plots, with 12–37% species in common with natural sites. Rehabilitated sites were more similar in composition to each other than they were to the natural sites, with two strong colonizers, the fire-killed Acacia blakelyi and the fire-tolerant Melaleuca leuropoma , universally present. Dendrograms and ordinations based on composition and cover showed that rehabilitated sites grouped with each other before they did with the Dune and Swale sites (physically closest), and last with the Laterite and Limestone sites. Plant densities for R16 and R24 were about half those of the High Dune and Limestone, and about a quarter those of the Swale and Laterite. Fire resprouters were under-represented in the rehabilitated sites. Growth form distribution in rehabilitated sites was most similar to those of the dunes, with some woody shrubs up to 2.5 m tall present. Total iron and soil hardness (penetrability) were the only soil factors consistently different (higher) in the rehabilitated sites.     

Eph/Ephrin membrane proteins: a mammalian expression vector pTIg-BOS-Fc allowing rapid protein purification

There is an urgent need for high purity, single chain, fully functional Eph/ephrin membrane proteins. This report outlines the pTIg-BOS-Fc vector and purification approach resulting in rapid increased production of fully functional single chain extracellular proteins that were isolated with high purity and used in structure-function analysis and pre-clinical studies.     

Multiple Shoot Regeneration from Young Shoots of Kenaf (Hibiscus cannabinus)

A method was developed to initiate multiple shoots from the young shoot of kenaf. Young shoots along with the cotyledons were excised from ten-day old aseptically germinated seeds and pre-cultured for two weeks in Murashige and Skoog (MS) medium supplemented with benzyl adenine (BA) or a combination of BA and kinetin. After two weeks in culture, elongated shoots were excised above the cotyledonary nodes and cultured on fresh medium of the same composition. Multiple shoots were initiated within eight weeks. The number of shoots varied among cultivars. The highest number of shoots (11/explant) occured in cultivar Tainung 2 (T2) cultured in MS medium supplemented with 8.8 μM BA. Concentrations of BA higher than 8.8 μM had a negative effect on the number of shoots. Furthermore, callus growth was initiated from which morphologically abnormal shoots were induced. Kinetin had a significant effect only on cultivar Everglades 41 (E41). Shoot elongation and rooting were obtained simultaneously in half strength MS basal medium with no plant growth regulators. About 98% of the rooted plants were grown to maturity under greenhouse conditions. This method was successful with all four genotypes tested. However, significant genotypic variations were observed among the genotypes.     

Carbon dioxide compensation values in citronella and lemongrass

Carbon dioxide compensation values of mature leaves from 10 selections of citronella (Cymbopogon nardus [L.] Rendle) grown at 32/27 or 27/21 C day/night temperatures and three strains of lemongrass (Cymbopogon citratus [D.C.] Stapf. and Cymbopogon flexuosus [D.C.] Stapf.) grown at 8- or 15-hour photoperiods were measured in a controlled environment at 25 C. All leaves had low compensation values but citronella varied from 1.3 to 9.7 μl/liter and lemongrass from 0.7 to 3.5 μl/liter. Lower growing temperature generally resulted in lower compensation values for citronella but there was no consistent photoperiod effect on lemongrass.     

Equilibration of H labeling between body water and free amino acids: Enabling studies of proteome synthesis

Protein synthesis can be estimated by measuring the incorporation of a labeled amino acid into a proteolytic peptide. Although prelabeled amino acids are typically administered, recent studies have tested ²H₂O; the assumption is that there is rapid equilibration of ²H (in body water) with the carbon-bound hydrogens of amino acids before those amino acids are incorporated into a protein(s). We have determined the temporal changes in ²H labeling of body water and amino acids which should build confidence in ²H₂O-based studies of protein synthesis when one aims to measure the ²H labeling of proteolytic peptides.     

Equilibration of (2)H labeling between body water and free amino acids: enabling studies of proteome synthesis

Protein synthesis can be estimated by measuring the incorporation of a labeled amino acid into a proteolytic peptide. Although prelabeled amino acids are typically administered, recent studies have tested (2)H(2)O; the assumption is that there is rapid equilibration of (2)H (in body water) with the carbon-bound hydrogens of amino acids before those amino acids are incorporated into a protein(s). We have determined the temporal changes in (2)H labeling of body water and amino acids which should build confidence in (2)H(2)O-based studies of protein synthesis when one aims to measure the (2)H labeling of proteolytic peptides.     

Quantification of Magnetically Induced Changes in ECM Local Apparent Stiffness

The stiffness of the extracellular matrix (ECM) is known to influence cell behavior. The ability to manipulate the stiffness of ECM has important implications in understanding how cells interact mechanically with their microenvironment. This article describes an approach to manipulating the stiffness ECM, whereby magnetic beads are embedded in the ECM through bioconjugation between the streptavidin-coated beads and the collagen fibers and then manipulated by an external magnetic field. It also reports both analytical results (obtained by formal modeling and numerical simulation) and statistically meaningful experimental results (obtained by atomic force microscopy) that demonstrate the effectiveness of this approach. These results clearly suggest the possibility of creating desired stiffness gradients in ECM in vitro to influence cell behavior.