The Antifungal Action of 1,10-o-phenanthroline and EDTA is Mediated Through Zinc Chelation and Involves Cell Wall Construction  

S. Brul^1*, M. Stratford¹, J. M. van der Vaart², S. K. Dielbandhoesing¹, H. Steels¹, F. M. Klis³, C. T. Verrips<sup>1,2

1</sup>Unilever Research Laboratories Vlaardingen/Colworth, 3133AT Viaardingen The Netherlands MK44 1LQ Sharnbrook, Bedford, UK. 
²Department of Molecular Cell Biology, Utrecht University, 3584 CH Utrecht, The Netherlands
³Department of Molecular Cell Biology, BioCentrum Amsterdam, University of Amsterdam, 1098 SM Amsterdam, The Netherlands

Article history:
Received October 6, 1997
Accepted November 12, 1997

Key words:
Zn²⁺, growth, secretion, cell wall, yeast, fungi

Summary:
Yeast growth was inhibited at 1,10-o-phenanthroline concentrations of 0.1-0.5 mM and EDTA concentrations of 1-2 mM. Growth was most efficiently restored by adding Zn²⁺ to the medium, whereas Fe³⁺ was less effective, and Cu²⁺, Mn²⁺, Ca²⁺, or Mg²⁺ had only a minor or negligible effect. Culturing yeast cells in defined media that lacked individual metal salts showed that zinc ions were indeed needed for growth. These observations indicate that the growth inhibition caused by 1,10-o-phenanthroline and EDTA is largely due to a deficiency in zinc ions. Growth inhibition of filamentous fungi by 1,10-o-phenanthroline and EDTA could also be relieved by adding zinc ions to the medium. Chelation did not prevent the formation of an osmotically stable cell wall in regenerating yeast spheroplasts indicating that the synthesis of β-glucan and chitin were not affected. Regenerating spheroplasts that expressed α-galactosidase secreted equal amounts of α-galactosidasc in the presence of l,10-o-phenanlhroline as control cells indicating that protein synthesis and the secretory pathway were functioning normally in the presence of 1,10-o-phenanthroline. However, regenerating yeast spheroplasts that expressed a cell wall fusion protein containing α-galactosidase as a reporter protein released much less fusion protein into the medium than control cells. We suggest that the processing step responsible for releasing GPl-bound cell wall proteins from the plasma membrane might be affected, resulting in defective cell wall construction. 

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Section Microbiology and Preservation, Unilever Research Labovatorium, Olivier van Noortlaan 120, 3133 AT Vlaardingen, The Netherlands