Metabolic Rewiring Improves the Production of the Fungal Active Targeting Molecule Fusarinine C

Abstract

Author: Recently, increasing research in siderophores has been dedicated to their possible medical applications in diagnostics and therapeutics for human pathogenic infections. Fusarinine C (FsC) is a natural hydroxamate siderophore that harbors three amino groups, which allow the easy chemical modification of FsC for the design of novel multifunctional conjugates. However, low production of FsC has hampered its extensive exploitation.Herein, we rewired the FsC biosynthetic pathway in the Aureobasidium melanogenum HN6.2 strain to achieve a self-supplying l-ornithine with component-simplified and enhanced production of extracellular siderophores, for which the FsC accounted for 94%, its final titer being approximately 1.7 g L–1. The convenient acquisition of FsC effectuated our exploitation for its application. We employed in vitro and in vivo assays to show that FsC is an active targeting molecule that acts on the human pathogenic fungi Trichophyton rubrum and Candida albicans; this demonstrates the potential to use FsC for the development of novel antifungal targeting reagents in the future.

Bioproduction of L‐piperazic acid in gram scale using Aureobasidium melanogenum

“…In A. melanogenum, siderophore biosynthesis is the only known metabolic pathway that uses N 5 -hydroxy-L-ornithine as the precursor (Lu et al, 2019a(Lu et al, ,2019b. Previously, we reported that the biosynthesis of ferricrocin and hydroxyferricrocin siderophores in the recombined A. melanogenum strain DOLC19 was eliminated through the deletion of the L-ornithine-N 5transacetylase gene sidL and the FC synthase gene sidC (Lu et al, 2019a(Lu et al, ,2019b; however, the biosynthesis of fusarinine C (FsC) and FsC-derived fusarinine B (FsB) siderophores was still active. To completely block the consumption of N 5 -hydroxy-L-ornithine through the biosynthesis of FsC and FsB in the DOLC19 strain, the first genetic manipulation was the deletion of the anhydromevalonyl-CoA transferase gene sidF, which converts N 5 -hydroxy-L-ornithine into FsC (Hu et al, 2019) (Fig.…”

“…1B, Module 1), yielding SidF-null strain DF1. After the DF1 strain was cultivated in an iron-depleted (modified sucrose peptone; MSP) medium (Lu et al, 2019a(Lu et al, ,2019b for 120 h, high-performance liquid chromatography (HPLC) analysis showed that none of the characteristic peaks of FsC and FsB siderophores produced by the DOLC19 strain (Fig. S1A) could be detected in the fermentation broth of the DF1 strain (Fig.…”

“…Following the development of the strain DFK1, a higher production of L-Piz was pursed to pave the basis for its possible scale-up production as a fine chemical. It should be noted that MSP medium [2.5% sucrose, 0.4% (NH 4 ) 2 SO 4 , 0.3% K 2 HPO 4 , 0.075% citric acid, 0.008% MgSO 4 , 0.0002% ZnSO 4 and 0.17% yeast nitrogen base (YNB) without amino acids, (w/v)] (Lu et al, 2019a(Lu et al, ,2019b was used to establish iron-depleted conditions. As described above, the sidA gene has not been modified; thus, its transcription would be strongly inhibited under the presence of iron in the medium (Chi et al, 2013).…”

“…We previously found that N 5 -hydroxy-L-ornithine is a precursor for the produced hydroxamate siderophores in the yeast-like fungus Aureobasidium melanogenum. This precursor is synthesized via the catalysis of SidA, a homolog of KtzI, using L-ornithine as the substrate (Lu et al, 2019a(Lu et al, ,2019b. In addition, we developed an efficient genome editing system for A. melanogenum (Lu (B) Bioengineering A. melanogenum strain for the production of L-piperazic acid.…”

Transfer of Nitrogen Fixation (nif) Genes to Non‐diazotrophic Hosts

“…Notably, M. thermautotrophicus NifB generated NifX that contained in vivo formed NifB-co. [62] The FeMo (NifDK) tetramer and the active Fe protein from P. polymyxa WLY78 werep roduced in the cytoplasm of S. cerevisiae by expressing a1 5-genen itrogenase biosynthetic pathway ( % 38 kb). [63] Of the 15 genes, 11 genes (nifH, nifD, nifK, nifB, nifE, nifN, nifV, nifX, hesA, groES, groEL)w ere from P. polymyxa WLY78 and 4g enes (nifF, nifJ, nifS, nifU)w ere from K. oxytoca. Each coding region of the 15 genes was codon optimized for S. cerevisiae and then fused with ay east promoter and a yeast terminator,g enerating 15 expressionc assettes.…”