Molecular Evaluation of the Plasma Membrane Proton Pump fromAspergillus fumigatus

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Abstract
The gene encoding the plasma membrane proton pump (H+-ATPase) ofAspergillus fumigatus,PMA1, was characterized fromA. fumigatusstrain NIH 5233 and clinical isolate H11-20. An open reading frame of 3,109 nucleotides with two introns near the N terminus predicts a protein consisting of 989 amino acids with a molecular mass of approximately 108 kDa. The predictedA. fumigatusenzyme is 89 and 51% identical to H+- ATPases ofAspergillus nidulansandSaccharomyces cerevisiae, respectively. TheA. fumigatus PMA1is a typical member of the P-type ATPase family that contains 10 predicted transmembrane segments and conserved sequence motifs TGES, CSDKTGT, MLTGD, and GDGVN within the catalytic region. The enzyme represents 2% of the total plasma membrane protein, and it is characteristically inhibited by orthovanadate, with a 50% inhibitory concentration of ∼1.8 μM. H+-ATPases fromAspergillusspp. contain a highly acidic insertion region of 60 amino acids between transmembrane segments 2 and 3, which was confirmed for the membrane-assembled enzyme with a peptide-derived antibody. An increasingA. fumigatus PMA1copy number confers enhanced growth in low-pH medium, consistent with its role as a proton pump. These results provide support for the development of theA. fumigatusH+-ATPase as a potential drug discovery target.

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