SD0006: a potent, selective and orally available inhibitor of p38 kinase.

SD0006 is a diarylpyrazole that was prepared as an inhibitor of p38 kinase-alpha (p38alpha). In vitro, SD0006 was selective for p38alpha kinase over 50 other kinases screened (including p38gamma and p38delta with modest selectivity over p38beta). Crystal structures with p38alpha show binding at the ATP site with additional residue interactions outside the ATP pocket unique to p38alpha that can confer advantages over other ATP competitive inhibitors.

Development of improved cell lysis, solubilization and imaging approaches for proteomic analyses.

Analysis of complex biochemical processes at the level of the proteome requires methods that quantitatively solubilize cytosolic and membrane bound proteins yet are compatible with isoelectric focusing and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. In addition, it is often necessary to employ several highly sensitive detection methods to identify key proteins that are modified or exhibit a change in expression levels in response to a given experimental stimulus or condition. Methods were developed that efficiently extract tissues or lyse cultured cells and quantitatively solubilize proteins in a single step without the need to shear nucleic acids.

Practical aspects of fluorescent staining for proteomic applications.

SYPRO Orange and SYPRO Ruby staining methods, modified for use with large-format two dimensional (2-D) gels, are compared to the manufacturer's recommended protocols to determine sensitivity and reproducibility of the new methods. This study examines the critical aspects of fixation, washing, and staining to develop an optimized fluorescent staining method. It was determined that careful control of sodium dodecyl sulfate (SDS) levels and pH in the gel was critical for successful staining with SYPRO Orange.

Multiple conformations of a human interleukin-3 variant.

Interleukin-3 (IL-3) is a cytokine that stimulates the proliferation and differentiation of hematopoietic cells. The hyperactive hIL-3 variant SC-55494 was shown to have at least two major conformations by high-resolution NMR spectroscopy. Mutants of SC-55494 were constructed in which alanine was substituted for proline in order to test the hypothesis that proline cis-trans isomerization is the source of the observed conformational heterogeneity, as well as to evaluate the effect of prolyl peptide bond configuration on biological activity.

Large-scale production and characterization of Bacillus thuringiensis subsp. tenebrionis insecticidal protein from Escherichia coli.

Bacillus thuringiensis subsp. tenebrionis insecticidal protein was produced in recombinant Escherichia coli and purified to near homogeneity to provide quantities of protein for safety-assessment studies associated with the registration of transgenic potato plants. The 68-kDa protein is produced naturally by Bacillus thuringiensis subsp.