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Essential Genes as Broad-Spectrum Antifungal Targets
K S Dobb 1 , M Birch 1 , M J Bromley 2 , S Kaye 1 , L Stateva 3 , J D Oliver 1
1 F2G Ltd, Lankro Way, Manchester, UK. 2 Manchester Academic Health Centre,
The University of Manchester UK. 3 Faculty of Life Sciences, The University of
Manchester, UK
Aspergillus fumigatus and Candida albicans both cause serious, systemic disease
in vulnerable patients. Current antifungal drugs have many limitations and there is
a need for new classes of antifungal drugs. This study aims to investigate genes
essential for the growth of A. fumigatus and C. albicans as antifungal targets.
In silico analysis of a set of essential genes from A. fumigatus revealed those with
C. albicans homologs. To assess selectivity those with human homologs were also
identified. A subset of the A. fumigatus essential gene set was chosen for
essentiality studies in C. albicans.
One gene found to be essential in both A. fumigatus and C. albicans was
Phosphopantetheinyl transferase b (PPTb). This gene is involved in mitochondrial
fatty acid synthesis and catalyses the transfer of a phosphopantetheine group from
coenzyme A (CoA) to acyl carrier protein (ACP). To identify inhibitors of Pptb a
fluorescence polarisation (FP) assay was developed. The transfer of fluorophore
labelled phosphopantetheine group from CoA to ACP can be measured by a
change in FP value. As Pptb catalyses this transfer inhibition of this enzyme can
also be detected by a change in FP values. A statistically robust FP assay was
achieved using C. albicans ACP and proteins and fluorescently labelled CoA. Single
concentration and IC50 screening of a small molecule compound library has
identified inhibitors of Pptb. Other work on these “hits” has included antifungal
susceptibility testing (minimum inhibitory concentration, MIC) and mammalian cell
toxicology testing (MCTs). PPTb has so far shown good potential as a broadspectrum
antifungal target and future work will aim to further characterise these
small molecule inhibitors for their potential as drug leads.
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