Isolation and identification of eukaryotic initiation factor 4A as a molecular target for the marine natural product Pateamine A.
ABSTRACT Natural products continue to demonstrate their utility both as therapeutics and as molecular probes for the discovery and mechanistic deconvolution of various cellular processes. However, this utility is dampened by the inherent difficulties involved in isolating and characterizing new bioactive natural products, in obtaining sufficient quantities of purified compound for further biological studies, and in developing bioactive probes. Key to characterizing the biological activity of natural products is the identification of the molecular target(s) within the cell. The marine sponge-derived natural product Pateamine A (PatA) has been found to be an inhibitor of eukaryotic translation initiation. Herein, we describe the methods utilized for identification of the eukaryotic translation initiation factor 4A (eIF4A) as one of the primary protein targets of PatA. We begin by describing the synthesis of an active biotin conjugate of PatA (B-PatA), made possible by total synthesis, followed by its use for affinity purification of PatA binding proteins from cellular lysates. We have attempted to present the methodology as a general technique for the identification of protein targets for small molecules including natural products.
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ABSTRACT: Natural products, and their derivatives and mimics, have contributed to the development of important therapeutics to combat diseases such as infections and cancers over the past decades. The value of natural products to modern drug discovery is still considerable. However, its development is hampered by a lack of a mechanistic understanding of their molecular action, as opposed to the emerging molecule-targeted therapeutics that are tailored to a specific protein target(s). Recent advances in the mass spectrometry-based proteomic approaches have the potential to offer unprecedented insights into the molecular action of natural products. Chemical proteomics is established as an invaluable tool for the identification of protein targets of natural products. Small-molecule affinity selection combined with mass spectrometry is a successful strategy to "fish" cellular targets from the entire proteome. Mass spectrometry-based profiling of protein expression is also routinely employed to elucidate molecular pathways involved in the therapeutic and possible toxicological responses upon treatment with natural products. In addition, mass spectrometry is increasingly utilized to probe structural aspects of natural products-protein interactions. Limited proteolysis, photoaffinity labeling, and hydrogen/deuterium exchange in conjunction with mass spectrometry are sensitive and high-throughput strategies that provide low-resolution structural information of non-covalent natural product-protein complexes. In this review, we provide an overview on the applications of mass spectrometry-based techniques in the identification and characterization of natural product-protein interactions, and we describe how these applications might revolutionize natural product-based drug discovery.Mass Spectrometry Reviews 01/2009; 29(1):126-55. DOI:10.1002/mas.20235 · 8.05 Impact Factor
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ABSTRACT: The survival and proliferation of cancer cells depend largely on the elevation of oncogene expression. Oncogenic expression can be affected by alterations in translation, or protein synthesis. Translation initiation, the rate limiting step of translation, closely regulates protein synthesis which both healthy and cancerous cells require for division and growth. Inhibitors of the eukaryotic translation initiation factor 4F (eIF4F) have been shown to selectively repress the expression of oncogenes in multiple in vitro studies. Although chemotherapeutic application of translation initiation inhibition is in its early stages, promising in vitro results demonstrate the potential of eIF4F inhibitors for use in clinical settings. This review offers a brief overview of the mechanisms that underlie eIF4F inhibitor activity in combating cancer.
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ABSTRACT: Marine organisms have attracted scientific community as a rich source of natural products with unusual structural features and remarkable biological activities. Marine macrolides are a prominent class of natural products characterized by a highly oxygenated polyene backbone containing a macrocyclic lactone as a conformational constraint. Many marine macrolides possess outstanding cell growth antiproliferative properties, making them valuable molecular probes for the investigation of biochemical pathways and promising lead compounds for the development of new antitumor chemotherapeutic agents. In the present review we intend to focus on marine macrolides with potent cytotoxic activity that could be exploited in cancer research and therapy, along with those macrolides currently in clinical trials and/or preclinical development.Anti-cancer agents in medicinal chemistry 03/2009; 9(2):122-37. DOI:10.2174/187152009787313800 · 2.94 Impact Factor