The nature and degree of the biological activity shown by triadimenol and paclobutrazol are influenced considerably by the absolute configuration of the two asymmetric carbon atoms present in the molecules. The order of the activity of triadimenol enantiomers was found to be: fungitoxicity—1S, 2R > 1R, 2R > 1R, 2S > 1S, 2S; inhibition of gibberellin biosynthesis—1R, 2S > 1S, 2S > 1R, 2R > 1S, 2R; and inhibition of plant sterol biosynthesis—1R, 2S≈1S, 2R > 1R, 2R > 1S, 25. The relative activity of paclobutrazol enantiomers was: fungitoxicity— 2R, 3R > 2S, 3R > 2R, 3S≈2S, 3S; inhibition of gibberellin biosynthesis—2S, 3S > 2R, 3S > 2R, 3R > 2S, 3R; and inhibition of plant sterol biosynthesis—2R, 3S > 2R, 3R > 2S, 3S > 2S, 3R. These data indicate that the R configuration at the chiral carbon bearing the hydroxyl group is the prime determinant for fungitoxicity whereas enantiomers having the S configuration at this carbon are effective inhibitors of gibberellin biosynthesis. This agrees well with published data for the structurally related vinylazoles. Plant sterol C‐14a demethylation was not always inhibited by enantiomers having high fungicidal activity thus indicating that the structural features necessary for binding to the plant C‐14a demethylase may differ from those needed for the analogous fungal enzyme.

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The Magnaporthe grisea ERG2 gene, encoding delta 8-->delta 7 sterol isomerase, was isolated from a genomic library by heterologous hybridization to a fragment of the Ustilago maydis ERG2 gene. The isolated gene contained a reading frame of 745 bp which encoded a protein of 221 amino acids. The coding region was interrupted by a single putative 79-bp-long intron. The deduced amino-acid sequence exhibited similarity to the ERG2 gene products of U. maydis and of Saccharomyces cerevisiae, particularly in the central region of the proteins. The NH2-terminal of all three proteins contained a long stretch of amino acids that were strongly hydrophobic, suggesting that they may function by anchoring the protein to a membrane surface. The M. grisea ERG2 gene complemented a U. maydis deletion mutant in which the ERG2 gene had been removed using a one-step gene replacement procedure. The delta 8-->delta 7 sterol isomerase produced by the M. grisea ERG2 gene exhibited a level of sensitivity to the sterol biosynthesis inhibitor, tridemorph, similar to that of the enzyme derived from the U. maydis ERG2 gene.

Ustilago maydis mutants resistant to polyene antibiotics were screened for defects in ergosterol biosynthesis. Slowgrowing mutants recovered after selecting for amphotericin B resistance were devoid of ergosterol and accumulated the methylated sterols, 14a-methylfecosterol, obtusifoliol and eburid, indicating that these isolates were impaired in C-14 sterol demethylation and were similar to the Erg40 mutant of U. m y & By contrast, nystatin-and pimaricin-resistant isolates which exhibited reduced growth rates showed a dysfunction in C-8 sterol isomerization. In these mutants (Erg2) ergosterol was replaced by the A*-sterols, ergosta-5,8,22-trienol, ergosta-8,22-dienol, fecosterol and ergost-8-enol. Analysis of a random sample of polyene-resistant isolates that grew normally revealed that although most retained a typical wild-type sterol profile, two of the isolates failed to accumulate ergosterol. The msjor sterols detected in these isolates were ergosta-7,22-dienol and ergosta-7-eno1, suggesting a lesion in C-5 sterol desaturation in these mutants (Erg3). Of four Erg2 mutants recovered, one mutant, selected on nystatin, contained low but detectable amounts of ergosterol. Ergosterol was not detected immediately after selection in the three other pimaricin-resistant Erg2 mutants. Although the growth of three of the Erg2 mutants remained unchanged during non-selective culture, one mutant reverted and began to grow at a greater rate than the rest; analysis of the sterols produced by this strain revealed that ergosterol was now present, but at lower concentrations than those in the wild-type strain. No changes in the type of sterol formed were observed in the other slower-growing Erg2 mutants even after prolonged culture. All the Erg2 mutants exhibited morphological abnormalities; sporidia were swollen and distorted, and the inability of sporidia to separate after cell division led to the development of highly branched, multicellular groups of cells.