Cryptococcus neoformans Resistance to Echinocandins: (1,3)β-Glucan Synthase Activity Is Sensitive to Echinocandins
Open Access
- 1 July 2005
- journal article
- Published by American Society for Microbiology in Antimicrobial Agents and Chemotherapy
- Vol. 49 (7), 2851-2856
- https://doi.org/10.1128/aac.49.7.2851-2856.2005
Abstract
(1,3)β-d-Glucan synthase (EC 2.4.1.34. UDP-glucose: 1,3-β-d-glucan 3-β-glucosyltransferase) uses UDP-glucose as substrate and catalyzes the polymerization of glucose ([1,3]-β-linkages) to form the major carbohydrate component of the fungal cell wall. We have optimized in vitro assay conditions for (1,3)β-glucan synthase activity from Cryptococcus neoformans. Cells lysed in 50 mM Tris, pH 7.75, containing 20% glycerol, 2 mM NaF, 1 mM dithiothreitol, 0.1 mM phenylmethylsulfonyl fluoride, 5 mM MgCl2, 0.1% protease and phosphatase inhibitor cocktails, and 60 μM GTPγS produced maximum specific activity in vitro. We tested in vitro C. neoformans (1,3)β-glucan synthase activity against the (1,3)β-glucan synthase inhibitors, caspofungin and cilofungin, and have determined that (1,3)β-glucan synthase activity is very sensitive (apparent Ki of 0.17 ± 0.02 μM and 22 ± 5.7 μM, respectively) to these echinocandins. Taken together with high MICs for C. neoformans (caspofungin MIC, 16 μg/ml; cilofungin MIC, 64 μg/ml), our results indicate that C. neoformans is resistant to caspofungin and cilofungin by a mechanism(s) unrelated to (1,3)β-glucan synthase resistance.Keywords
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