Tetracycline-inducible expression of individual secreted aspartic proteases in Candida albicans allows isoenzyme-specific inhibitor screening.

Détails

ID Serval
serval:BIB_0C47BB583005
Type
Article: article d'un périodique ou d'un magazine.
Collection
Publications
Institution
Titre
Tetracycline-inducible expression of individual secreted aspartic proteases in Candida albicans allows isoenzyme-specific inhibitor screening.
Périodique
Antimicrobial Agents and Chemotherapy
Auteur(s)
Staib P., Lermann U., Blass-Warmuth J., Degel B., Würzner R., Monod M., Schirmeister T., Morschhäuser J.
ISSN
0066-4804
Statut éditorial
Publié
Date de publication
2008
Peer-reviewed
Oui
Volume
52
Numéro
1
Pages
146-156
Langue
anglais
Résumé
The yeast Candida albicans possesses a gene family that encodes secreted aspartic proteases (Saps), which are important for the virulence of this human fungal pathogen. Inhibitors of the Saps could therefore be used as novel antimycotic agents for the treatment of C. albicans infections. In the present study, we established a bioassay which allows testing of the activity of potential protease inhibitors against specific Sap isoenzymes by their ability to inhibit protease-dependent growth of C. albicans. In a medium containing bovine serum albumin (BSA) as the sole source of nitrogen, C. albicans specifically expresses the Sap2p isoenzyme, which degrades the BSA and thereby enables the fungus to grow. As the other SAP genes are not significantly expressed under these conditions, mutants lacking SAP2 are unable to utilize BSA as a nitrogen source and cannot grow in such a medium. To investigate whether forced expression of SAP genes other than SAP2 would also allow growth on BSA, we constructed a set of strains expressing each of the 10 SAP genes from a tetracycline-inducible promoter in a sap2Delta mutant background. Expression of Sap1p, Sap2p, Sap3p, Sap4p, Sap5p, Sap6p, Sap8p, and a C-terminally truncated, secreted Sap9p restored the growth of the sap2Delta mutant with different efficiencies. This set of strains was then used to test the activities of various aspartic protease inhibitors against specific Sap isoenzymes by monitoring growth on BSA in the presence of the inhibitors. While pepstatin blocked the activity of all of the Saps tested, the human immunodeficiency virus protease inhibitors ritonavir and saquinavir inhibited growth of the strains expressing Sap1p to Sap3p and Sap1p, respectively, but not that of strains expressing other Saps. Therefore, the strain set can be used to test the activity of new protease inhibitors against individual C. albicans Sap isoenzymes by their ability to block the growth of the pathogen.
Mots-clé
Aspartic Endopeptidases/biosynthesis, Aspartic Endopeptidases/genetics, Candida albicans/drug effects, Candida albicans/enzymology, Culture Media, Enzyme Induction, Humans, Isoenzymes/antagonists &amp, inhibitors, Microbial Sensitivity Tests/methods, Mutation, Pepstatins/pharmacology, Promoter Regions, Genetic, Protease Inhibitors/pharmacology, Ritonavir/pharmacology, Saquinavir/pharmacology, Tetracycline/pharmacology
Pubmed
Web of science
Création de la notice
25/01/2008 16:47
Dernière modification de la notice
20/08/2019 12:33
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