Disruption of the gene encoding the secreted acid protease (ACP) in the yeast Candida tropicalis |
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Authors: | Dominique Sanglard Guiseppe Togni Pierre A de Viragh Michel Monod |
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Institution: | Institut für Biotechnologie, ETH-H?nggerberg, Zürich, Switzerland. |
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Abstract: | The gene for the secreted acid protease (ACP), a potential virulence factor of Candida species, was inactivated in Candida tropicalis by gene disruption. The disruption was performed by cotransformation of an ade2 C. tropicalis mutant with a linear DNA fragment carrying a deletion in ACP, and the replicative vector pMK16 which carries a selectable ADE2 gene marker. Few of the transformants exhibited lower protease secretion levels and were shown to have one deleted and one unaffected ACP copy, since C. tropicalis is a diploid yeast. These transformants were rendered homozygotic for this deletion by mild UV-treatment. One of the homozygotic acp deletion mutants obtained was completely devoid of extracellular protease activity and grew poorly on bovine serum albumin-containing medium. This mutant could be complemented by an ACP fragment inserted in pMK16, but also by an acid protease gene isolated from C. parapsilosis. |
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Keywords: | Candida tropicalis Secretion Acid protease (EC 3 4 23 6) Gene disruption Complementation |
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