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Antimicrobial Agents and Chemotherapy, August 2004, p. 2930-2936, Vol. 48, No. 8
0066-4804/04/$08.00+0     DOI: 10.1128/AAC.48.8.2930-2936.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Ribosomal P0 Protein Domain Involved in Selectivity of Antifungal Sordarin Derivatives

C. Santos, M. A. Rodríguez-Gabriel,^, M. Remacha, and J. P. G. Ballesta^*

Centro de Biología Molecular Severo Ochoa, C.S.I.C. and U.A.M., Canto Blanco, Madrid 28049, Spain

Received 23 February 2004/ Returned for modification 1 March 2004/ Accepted 14 March 2004

The ribosomal stalk protein P0 is involved in the susceptibility to the antifungal sordarin derivatives, as reported for a number of Saccharomyces cerevisiae resistant mutants. Mammals and some lower eukaryotes are naturally resistant to these compounds. It is shown here that expression in S. cerevisiae of the ribosomal protein P0 from Homo sapiens and from other sordarin-resistant organisms results in a decrease in the sensitivity of the cells to an agent of this class. To further characterize the P0 region responsible for inducing sordarin resistance, a series of protein chimeras containing complementary regions of the human and yeast P0 proteins were constructed and expressed in yeast. The chimeras complement the absence of the native yeast P0 except in chimeras containing the human P0 carboxyl-terminal domain. Resistance to sordarins was found to be associated with the presence of an HsP0 amino acid sequence comprising P118 to F138, which unexpectedly led to higher resistance than the presence of the complete human P0. A comparison of the corresponding region in P0 from yeast and sordarin-insensitive organisms, followed by site-directed mutagenesis, indicates that residues in positions 119, 124, and 126 have an important role in determining resistance to sordarins. Moreover, since sordarins block the eukaryotic elongation factor 2 (EF2) function, the P0 region affecting sordarin susceptibility must correspond to EF2-interacting domains of the ribosomal stalk protein, which affects the drug-binding site in the elongation factor.

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* Corresponding author. Mailing address: Centro de Biología Molecular Severo Ochoa, C.S.I.C. and U.A.M., Canto Blanco, Madrid 28049, Spain. Phone: 34-914975076. Fax: 34-914974799. E-mail: jpgballesta{at}cbm.uam.es.

C.S. and M.A.R.-G. contributed equally to this study.

Present address: Department of Molecular Biology, The Scripps Research Institute, La Jolla, CA 92037.

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Antimicrobial Agents and Chemotherapy, August 2004, p. 2930-2936, Vol. 48, No. 8
0066-4804/04/$08.00+0     DOI: 10.1128/AAC.48.8.2930-2936.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

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