Sordarin Inhibits Fungal Protein Synthesis by Blocking Translocation Differently to Fusidic Acid

Domínguez, Juan M.; Gómez-Lorenzo, Marı́a G.; Martin, J.J.

doi:10.1074/jbc.274.32.22423

Cited by 69 publications

(41 citation statements)

References 29 publications

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“…The sordarin derivatives apparently block the interaction of the elongation factor in the ribosome (2,11) in a way similar to but distinguishable from fusidic acid (4). In agreement with this mode of action, a drug-binding site has been recently located in a yeast eukaryotic elongation factor 2 (EF2)-sordarin crystal structure (10) in a previously proposed position based on the mapping of sordarin resistance mutations in the elongation factor modeled structure (2).…”

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confidence: 56%

Ribosomal P0 Protein Domain Involved in Selectivity of Antifungal Sordarin Derivatives

Santos

Gabriel

Remacha

et al. 2004

Antimicrob Agents Chemother

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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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confidence: 56%

Ribosomal P0 Protein Domain Involved in Selectivity of Antifungal Sordarin Derivatives

Santos

Gabriel

Remacha

et al. 2004

Antimicrob Agents Chemother

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“…Cerythromycin (9 pmol; concentration = 9 3 10 À8 M) was mixed with 0-160 pmol (1.6 3 10 À6 M) ribosomes from the indicated sources. Ribosome concentrations were calculated from A 260 measurement and assuming that A 260 = 1 in a 10-mm cuvette corresponds to 23 pmol ribosomes per milliliter for E. coli (Haselkorn et al 1963) and 16 pmol per milliliter for yeast (Domínguez et al 1999 À7 M E. coli ribosomes or 4 3 10 À7 M yeast ribosomes. In all cases, erythromycin binding was measured by passing the mixture through nitrocellulose filters.…”

Section: Methodsmentioning

confidence: 99%

Mutation from guanine to adenine in 25S rRNA at the position equivalent to E. coli A2058 does not confer erythromycin sensitivity in Sacchromyces cerevisae

Bommakanti

Lindahl²,

Zengel³

2008

RNA

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The macrolide erythromycin binds to the large subunit of the prokaryotic ribosome near the peptidyltransferase center (PTC) and inhibits elongation of new peptide chains beyond a few amino acids. Nucleotides A2058 and A2059 (E. coli numbering) in 23S rRNA play a crucial role in the binding of erythromycin, and mutation of nucleotide A2058 confers erythromycin resistance in both Gram-positive and Gram-negative bacteria. There are high levels of sequence and structural similarity in the PTC of prokaryotic and eukaryotic ribosomes. However, eukaryotic ribosomes are resistant to erythromycin and the presence of a G at the position equivalent to E. coli nucleotide A2058 is believed to be the reason. To test this hypothesis, we introduced a G to A mutation at this position of the yeast Saccharomyces cerevisiae 25S rRNA and analyzed sensitivity toward erythromycin. Neither growth studies nor erythromycin binding assays on mutated yeast ribosomes indicated any erythromycin sensitivity in mutated yeast strains. These results suggest that the identity of nucleotide 2058 is not the only determinant responsible for the difference in erythromycin sensitivity between yeast and prokaryotes.

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“…(Justice et al, 1998). Recent studies by other workers suggest that sordarin prevents the transition of the ribosome from the PRE-eEF2-GTP to POST-eEF2-GTP state in Candida albicans (Dominguez et al, 1999). Despite the extensive homology between the eEF2s, there is no significant binding of sordarin to either plant or mammalian eEF2 (Dominguez et al, 1998 ;Justice et al, 1998), thus making sordarin a fungal-specific inhibitor of translation.…”

Section: Introductionmentioning

confidence: 99%

Species-specific inhibition of fungal protein synthesis by sordarin: identification of a sordarin-specificity region in eukaryotic elongation factor 2 The GenBank accession numbers for the sequences reported in this manuscript are AF107286–AF107291, AF292693 and AF248644.

Shastry

Nielsen

et al. 2001

Microbiology

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The sordarin class of natural products selectively inhibits fungal protein synthesis by impairing the function of eukaryotic elongation factor 2 (eEF2). Mutations in Saccharomyces cerevisiae eEF2 or the ribosomal stalk protein rpP0 can confer resistance to sordarin, although eEF2 is the major determinant of sordarin specificity. It has been shown previously that sordarin specifically binds S. cerevisiae eEF2 while there is no detectable binding to eEF2 from plants or mammals, despite the high level of amino acid sequence conservation among these proteins. In both whole-cell assays and in vitro translation assays, the efficacy of sordarin varies among different species of pathogenic fungi. To investigate the basis of sordarin's fungal selectivity, eEF2 has been cloned and characterized from several sordarin-sensitive and -insensitive fungal species. Results from in vivo expression of Candida species eEF2s in S. cerevisiae and in vitro translation and growth inhibition assays using hybrid S. cerevisiae eEF2 proteins demonstrate that three amino acid residues within eEF2 account for the selectivity of this class of compounds. It is also shown that the corresponding residues at these positions in human eEF2 are sufficient to confer sordarin insensitivity to S. cerevisiae identical to that observed with mammalian eEF2.

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Sordarin Inhibits Fungal Protein Synthesis by Blocking Translocation Differently to Fusidic Acid

Cited by 69 publications

References 29 publications

Ribosomal P0 Protein Domain Involved in Selectivity of Antifungal Sordarin Derivatives

Ribosomal P0 Protein Domain Involved in Selectivity of Antifungal Sordarin Derivatives

Mutation from guanine to adenine in 25S rRNA at the position equivalent to E. coli A2058 does not confer erythromycin sensitivity in Sacchromyces cerevisae

Species-specific inhibition of fungal protein synthesis by sordarin: identification of a sordarin-specificity region in eukaryotic elongation factor 2 The GenBank accession numbers for the sequences reported in this manuscript are AF107286–AF107291, AF292693 and AF248644.

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