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Fig. 7. Expression of the dLBR in the S. cerevisiae erg24 mutant and analysis of synthesized sterols. (A) erg24 mutant cells were transformed with plasmids containing the coding region of dLBR (lane 1), the wild-type ERG24 gene (lane 2) or the plasmid without gene (lane 3). Total proteins of these yeast strains were separated by SDS-PAGE and immunoblotted with dLBR antibodies. The dLBR expressed in yeast forms aggregates in SDS sample buffer; the position of unaggregated dLBR is marked by an arrow. The dLBR antibodies cross-react in addition with a low molecular weight yeast protein present in all three strains. Molecular masses of reference proteins (in kDa) are marked. (B-G) Mass spectrometric analysis of sterols synthesized in S. cerevisiae erg24 mutant cells that had been transformed with a plasmid containing cDNAs of the following genes: S. cerevisiae ERG24 (C); Arabidopsis thaliana FACKEL (D); a plasmid without gene (F, control); and the Drosophila LBR (G, dLBR). Sterols were extracted from cells that had been grown for 24 hours in YPAD medium. The mass spectra of ergosterol (B) and ignosterol (E) are shown as standards.
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