Krysan DJ and Didone L (2008) A high-throughput screening assay for small molecules that disrupt yeast cell integrity. J Biomol Screen 13(7):657-64
Abstract: Lead compounds for antifungal drug development are urgently needed because invasive fungal infections are an important cause of morbidity and mortality in immunocompromised patients. Here, a high-throughput screening assay for small molecules that cause yeast cell lysis is described. The assay is based on the detection of the intracellular enzyme adenylate kinase in the culture medium as a reporter of yeast cell lysis. Features of the assay protocol include 1) the ability to detect cell lysis at drug concentrations that cause no apparent growth defect, 2) specificity for fungicidal molecules, 3) a simple 1-plate, add-and-read protocol using a commercially available adenylate kinase assay kit, 4) short, 5-h incubation time, and 5) low cost. The assay is applicable to the model yeast Saccharomyces cerevisiae and to Candida albicans, the most common human fungal pathogen. The adenylate kinase assay is validated in a pilot screen of 4505 compounds. Consistent with its specificity for fungicidal molecules, the largest class of molecules identified in 2 libraries of known bioactive molecules targeted the plasma membrane. Fungistatic compounds are not detected by the assay. Adenylate kinase-based screening appears to be a useful approach to the direct identification of small molecules that kill yeast cells. (Journal of Biomolecular Screening XXXX:xx-xx).
|Status: Published||Type: Journal Article||PubMed ID: 18626115|
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