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The endotoxic shock syndrome is characterized by systemic inflammation, multiple organ damage, circulatory collapse and death. Systemic release of tumor necrosis factor (TNF)-alpha and other cytokines purportedly mediates this process. However, the primary tissue target remains unidentified. The present studies provide evidence that endotoxic shock results(More)
A proline-directed serine/threonine ceramide-activated protein (CAP) kinase mediates transmembrane signaling through the sphingomyelin pathway. CAP kinase reportedly initiates proinflammatory TNF alpha action by phosphorylating and activating Raf-1. The present studies delineate kinase suppressor of Ras (KSR), identified genetically in Caenorhabditis(More)
A high-resolution structure of a ligand-bound, soluble form of human monoglyceride lipase (MGL) is presented. The structure highlights a novel conformation of the regulatory lid-domain present in the lipase family as well as the binding mode of a pharmaceutically relevant reversible inhibitor. Analysis of the structure lacking the inhibitor indicates that(More)
Prior investigations document that proliferative signaling cascades, under some circumstances, initiate apoptosis, although mechanisms that dictate the final outcome are largely unknown. In COS-7 cells, ceramide signals Raf-1 activation through Ras (Zhang, Y., Yao, B., Delikat, S., Bayoumy, S., Lin, X. H., Basu, S., McGinley, M., Chan-Hui, P. Y.,(More)
MAPK-activated protein kinase-2 (MAPKAPK2) regulates the synthesis of tumor necrosis factor and other cytokines and is a potential drug target for inflammatory diseases. Five protein constructs were produced in 4-10mg quantities per liter of culture media using baculovirus-infected insect cells and characterized for kinase activity, thermal stability, and(More)
Diacylglycerol acyltransferase (DGAT) catalyzes the final step in triglyceride (TG) synthesis. There are two isoforms, DGAT1 and DGAT2, with distinct protein sequences and potentially different physiological functions. To date, the ability to determine clear functional differences between DGAT1 and DGAT2, especially with respect to hepatic TG synthesis, has(More)
Chymases (EC 3.4.21.39) are mast cell serine proteinases that are variably expressed in different species and, in most cases, display either chymotryptic or elastolytic substrate specificity. Given that chymase inhibitors have emerged as potential therapeutic agents for treating various inflammatory, allergic, and cardiovascular disorders, it is important(More)
Divergence of substrate specificity within the context of a common structural framework represents an important mechanism by which new enzyme activity naturally evolves. We present enzymological and x-ray structural data for hamster chymase-2 (HAM2) that provides a detailed explanation for the unusual hydrolytic specificity of this rodent alpha-chymase. In(More)
A parallel approach to designing crystallization constructs for the c-FMS kinase domain was implemented, resulting in proteins suitable for structural studies. Sequence alignment and limited proteolysis were used to identify and eliminate unstructured and surface-exposed domains. A small library of chimeras was prepared in which the kinase insert domain of(More)