Aspergillus fumigatus SidF is a dual substrate acyltransferase involved in biosynthesis of both fusarinine- and ferrichrome-type siderophores

ABSTRACT The human pathogen Aspergillus fumigatus produces fusarinine-type (FusTS) and ferrichrome-type siderophores (FchTS), both of which have been shown to be crucial for virulence of this mold. After the common first siderophore biosynthetic step, SidA-catalyzed hydroxylation of ornithine, the pathway splits. For FusTS biosynthesis, SidF incorporates an anhydromevalonyl group, while for FchTS biosynthesis, SidL and an as yet unknown enzyme incorporate an acetyl group. The transacylases SidF and SidL share only limited similarity in their C-terminal GNAT (Gcn5-related N-acetyltransferases) motif-containing domains. SidF is transcriptionally induced by iron limitation and localizes to peroxisomes, whereas SidL is a cytosolic enzyme with largely iron-independent expression. Here, we discovered that simultaneous inactivation of both SidF and SidL abolished the biosynthesis of both FusTS and FchTS and caused a growth defect under iron limitation, similar to the inactivation of SidA. Biosynthesis of both FusTS and FchTS depended on both the unique N-terminal and the GNAT motif-containing C-terminal SidF domains. In conclusion, SidF is the hitherto unknown FchTS biosynthetic enzyme: in contrast to SidL, SidF is a bifunctional enzyme accepting acetyl-CoA and anhydromevalonyl-CoA as substrates for biosynthesis of both FusTS and FchTS. Furthermore, this study revealed interdependence of FusTS and FchTS production, and that the peroxisomal localization of FusTS enzymes is important for optimizing FusTS production at the expense of FchTS. Phylogenetic analyses supported the relevance of these findings to other fungal species and revealed overlapping but distinct consensus sequences for the GNAT motifs of SidL and SidF, most likely reflecting their different substrate specificities. IMPORTANCE Adaptation to the host niche is key for any pathogenic organism. Aspergillus fumigatus is a major fungal pathogen causing 90% of invasive aspergillosis cases, which is associated with a high mortality rate. Siderophore-mediated iron acquisition has been shown to be essential for virulence of A. fumigatus and other fungal pathogens. In recent years, the hyphal siderophore biosynthetic pathway has been largely elucidated with exception of a single unknown enzyme, which we identified here as SidF. In contrast to another siderophore biosynthetic acyltransferase, SidL, SidF is a bifunctional enzyme accepting different substrates. As simultaneous inactivation of SidF and SidL, which share a common protein domain and a common substrate, blocks the biosynthesis of all siderophores, simultaneous targeting of SidF and SidL may allow development of new antifungal drugs. Phylogenetic analyses supported the relevance of these findings to other fungal species Moreover, this study clarified the rational for partial peroxisomal localization of siderophore biosynthesis and their metabolic interdependence. The human pathogen Aspergillus fumigatus produces fusarinine-type (FusTS) and ferrichrome-type siderophores (FchTS), both of which have been shown to be crucial for virulence of this mold. After the common first siderophore biosynthetic step, SidA-catalyzed hydroxylation of ornithine, the pathway splits. For FusTS biosynthesis, SidF incorporates an anhydromevalonyl group, while for FchTS biosynthesis, SidL and an as yet unknown enzyme incorporate an acetyl group. The transacylases SidF and SidL share only limited similarity in their C-terminal GNAT (Gcn5-related N-acetyltransferases) motif-containing domains. SidF is transcriptionally induced by iron limitation and localizes to peroxisomes, whereas SidL is a cytosolic enzyme with largely iron-independent expression. Here, we discovered that simultaneous inactivation of both SidF and SidL abolished the biosynthesis of both FusTS and FchTS and caused a growth defect under iron limitation, similar to the inactivation of SidA. Biosynthesis of both FusTS and FchTS depended on both the unique N-terminal and the GNAT motif-containing C-terminal SidF domains. Taken together, SidF is the hitherto unknown FchTS biosynthetic enzyme: in contrast to SidL, SidF is a bifunctional enzyme accepting acetyl-CoA and anhydromevalonyl-CoA as substrates for biosynthesis of both FusTS and FchTS. Moreover, this study revealed interdependence of FusTS and FchTS production, and that peroxisomal localization of FusTS enzymes is important for optimizing FusTS production at the expense of FchTS.