Structural and functional analysis of an oligomeric hydrophobin gene from Claviceps purpurea

SUMMARY Fungal hydrophobins are small hydrophobic proteins containing eight cysteine residues at conserved positions which have the ability to form amphipathic polymers. We have characterized a gene from the phytopathogenic ascomycete Claviceps purpurea, cpph1, which encodes a modular-type hydrophobin. It consists of five units, each showing a significant homology to class II hydrophobins. The units are separated by GN-repeat regions, which could form amphipathic alpha-helices; the amino terminus contains a glycine-rich region which could be involved in attaching the protein to the cell wall. The presence of long direct repeats within cpph1, and the high homology of the three internal modules suggest a recent generation of this gene from a tripartite precursor. Although sequencing of cDNA clones indicated that recombination could be mediated via the direct repeats, the majority of the transcripts appear to be full-sized. This was confirmed by Northern blot analysis, which showed the presence of a full-sized transcript in axenic culture. The high molecular weight pentahydrophobin was detected by Western blot analysis, indicating that CPPH1 is not processed into monomeric subunits. Targeted deletion of cpph1 did not lead to differences in morphology, growth rate, sporulation, or hydrophobicity of spores. Furthermore, the cpph1 deletion mutants showed no reduction in virulence on rye.