Genetic Engineering Activates Biosynthesis of Aromatic Fumaric Acid Amides in the Human Pathogen Aspergillus fumigatus

Abstract

ABSTRACTTheAspergillus fumigatusnonribosomal peptide synthetase FtpA is among the few of this species whose natural product has remained unknown. Both FtpA adenylation domains were characterizedin vitro. Fumaric acid was identified as preferred substrate of the first and bothl-tyrosine andl-phenylalanine as preferred substrates of the second adenylation domain. Genetically engineeredA. fumigatusstrains expressed eitherftpAor the regulator geneftpR, encoded in the same cluster of genes, under the control of the doxycycline-inducible tetracycline-induced transcriptional activation (tet-on) cassette. These strains produced fumaryl-l-tyrosine and fumaryl-l-phenylalanine which were identified by liquid chromatography and high-resolution mass spectrometry. Modeling of the first adenylation domainin silicoprovided insight into the structural requirements to bind fumaric acid as peptide synthetase substrate. This work adds aromatic fumaric acid amides to the secondary metabolome of the important human pathogenA. fumigatuswhich was previously not known as a producer of these compounds.