The fumiquinazolines (FQs) comprise a related, sequentially generated family of bioactive peptidyl alkaloids that are signature metabolites of Aspergillus fumigatus. The FQ framework is built by nonribosomal peptide synthetase (NRPS) machinery with anthranilate as a key non-proteinogenic amino acid building block. Despite being prevalent across the species, its gene cluster has not been characterized. Prior bioinformatic analysis coupled with heterologous expression of the putative A. fumigatus proteins termed here FmqA -FmqD led to the identification of a four-enzymatic process that builds increasingly complex FQ scaffolds. Briefly, FmqA, a trimodular NRPS condenses alanine, tryptophan, and anthranilic acid to form fumiquinazoline F (FQF). The tandem action of a flavoprotein (FmqB) and a monomodular NRPS (FmqC) converts FQF to fumiquinazoline A (FQA). Finally, FmqD, a FAD-dependent oxidoreductase converts FQA to the heptacyclic fumiquinazoline C (FQC). Interestingly, FmqD contains an N-terminus signal peptide predicted for extracellular transport. This study is aimed at providing in vivo validation to the FQ biosynthetic framework and characterizing how cellular localization of FmqD affects production of FQC in A. fumigatus. We found that the conidial metabolite, FQC, is the predominant FQ moiety in two wild type isolates and is selectively accumulated in the conidia. Targeted single gene deletions of FmqA through FmqD coupled with metabolomic profiling of the single biosynthetic gene mutants supported previous biochemical prediction of FQ biosynthesis. Fluorescent microscopy of mutants bearing a C-terminal FmqDGFP fusion showed that FmqD is localized to the cell wall of the fungus and this localization is abolished when the signal peptide is removed. Future studies will elucidate if cell wall localization of FmqD is crucial for FQC production.
Full conference title:
27th Fungal Genetics Conference
- Fungal Genetics Conference 27th (2013)