Elucidation of the complete ferrichrome A biosynthetic pathway in Ustilago maydis.
Iron is an important element for many essential processes in living organisms. To acquire iron, the basidiomycete Ustilago maydis synthesizes the iron-chelating siderophores ferrichrome and ferrichrome A. The chemical structures of these siderophores have been elucidated long time ago but so far only two enzymes involved in their biosynthesis have been described. Sid1, an ornithine monoxygenase, is needed for the biosynthesis of both siderophores, and Sid2, a non-ribosomal peptide synthetase (NRPS), is involved in ferrichrome generation. In this work we identified four novel enzymes, Fer3, Fer4, Fer5 and Hcs1, involved in ferrichrome A biosynthesis in U. maydis. By HPLC-MS analysis of siderophore accumulation in culture supernatants of deletion strains, we show that Fer3, an NRPS, Fer4, an enoyl-coenzyme A (CoA)-hydratase, and Fer5, an acylase, are required for ferrichrome A production. We demonstrate by conditional expression of the hydroxymethyl glutaryl (HMG)-CoA synthase Hcs1 in U. maydis that HMG-CoA is an essential precursor for ferrichrome A. In addition, we heterologously expressed and purified Hcs1, Fer4 and Fer5, and demonstrated the enzymatic activities by in vitro experiments. Thus, we describe the first complete fungal siderophore biosynthetic pathway by functionally characterizing four novel genes responsible for ferrichrome A biosynthesis in U. maydis.
Winterberg
Britta
B
Max Planck Institute for Terrestrial Microbiology, 35043 Marburg, Germany.
Uhlmann
Stefanie
S
Linne
Uwe
U
Lessing
Franziska
F
Marahiel
Mohamed A
MA
Eichhorn
Heiko
H
Kahmann
Regine
R
Schirawski
Jan
J
D000214:Acyl Coenzyme A / Q000378:metabolism; D000581:Amidohydrolases / Q000235:genetics / Q000378:metabolism; D053898:Biosynthetic Pathways / Q000235:genetics*; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D004746:Enoyl-CoA Hydratase / Q000235:genetics / Q000378:metabolism; D005291:Ferrichrome / Q000031:analogs & derivatives* / Q000378:metabolism; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism; D017353:Gene Deletion; D015870:Gene Expression; D023061:Gene Order; D005800:Genes, Fungal; D006904:Hydroxymethylglutaryl-CoA Synthase / Q000235:genetics / Q000378:metabolism; D013058:Mass Spectrometry; D008954:Models, Biological; D015394:Molecular Structure; D010453:Peptide Synthases / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D014588:Ustilago / Q000235:genetics* / Q000378:metabolism*
D000214:Acyl Coenzyme A; D005656:Fungal Proteins; D011994:Recombinant Proteins; C031058:ferrichrome A; C008047:3-hydroxy-3-methylglutaryl-coenzyme A; D005291:Ferrichrome; D006904:Hydroxymethylglutaryl-CoA Synthase; D000581:Amidohydrolases; C019665:amidase; D004746:Enoyl-CoA Hydratase; D010453:Peptide Synthases; C104385:non-ribosomal peptide synthase