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Complementation of a Saccharomyces cerevisiae ERG11/CYP51 (sterol 14?-demethylase) doxycycline-regulated mutant and screening of the azole sensitivity of Aspergillus fumigatus isoenzymes CYP51A and CYP51B.

ABSTRACT: Aspergillus fumigatus sterol 14?-demethylase isoenzymes CYP51A and CYP51B were heterologously expressed in a Saccharomyces cerevisiae mutant (YUG37-erg11), wherein native ERG11/CYP51 expression is controlled using a doxycycline-regulatable promoter. When cultured in the presence of doxycycline, recombinant YUG37-pcyp51A and YUG37-pcyp51B yeasts were able to synthesize ergosterol and grow; a control strain harboring reverse-oriented cyp51A could not. YUG37-pcyp51A and YUG37-pcyp51B constructs showed identical sensitivity to itraconazole, posaconazole, clotrimazole, and voriconazole. Conversely, YUG37-pcyp51A withstood 16-fold-higher concentrations of fluconazole than YUG37-pcyp51B (8 and 0.5 ?g ml?¹, respectively).

SUBMITTER: Martel CM 

PROVIDER: S-EPMC2976139 | biostudies-literature | 2010 Nov

REPOSITORIES: biostudies-literature

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Complementation of a Saccharomyces cerevisiae ERG11/CYP51 (sterol 14α-demethylase) doxycycline-regulated mutant and screening of the azole sensitivity of Aspergillus fumigatus isoenzymes CYP51A and CYP51B.

Martel Claire M CM   Parker Josie E JE   Warrilow Andrew G S AG   Rolley Nicola J NJ   Kelly Steven L SL   Kelly Diane E DE  

Antimicrobial agents and chemotherapy 20100823 11

Aspergillus fumigatus sterol 14α-demethylase isoenzymes CYP51A and CYP51B were heterologously expressed in a Saccharomyces cerevisiae mutant (YUG37-erg11), wherein native ERG11/CYP51 expression is controlled using a doxycycline-regulatable promoter. When cultured in the presence of doxycycline, recombinant YUG37-pcyp51A and YUG37-pcyp51B yeasts were able to synthesize ergosterol and grow; a control strain harboring reverse-oriented cyp51A could not. YUG37-pcyp51A and YUG37-pcyp51B constructs sho  ...[more]

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