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Title: CHARACTERIZATION OF THE USTILAGO MAYDIS SID2 GENE ENCODING A MULTIDOMAIN PEPTIDE SYNTHETASE IN THE FERRICHROME BIOSYNTHETIC GENE CLUSTER

Author
item YUAN, M - UNIVERSITY OF WISCONSIN
item GENTIL, GUILLAUME - UNIVERSITY OF WISCONSIN
item Budde, Allen
item Leong, Sally

Submitted to: Journal of Bacteriology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 4/12/2001
Publication Date: 4/12/2001
Citation: YUAN, M.W., GENTIL, G.D., BUDDE, A.D., LEONG, S.A. CHARACTERIZATION OF THE USTILAGO MAYDIS SID2 GENE ENCODING A MULTIDOMAIN PEPTIDE SYNTHETASE IN THE FERRICHROME BIOSYNTHETIC GENE CLUSTER. JOURNAL OF BACTERIOLOGY. 2001. 183:4040-4051.

Interpretive Summary: Iron is an essential trace element. Most microorganisms take up iron using a system comprised of extracellular iron-binding compounds called siderophores and transport system that recognizes these iron-laden compounds and takes them into cells. Siderophores are known to be important for virulence in many mammalian pathogens as a few plant pathogens. We have isolated a new gene involved in the process of siderophore biosynthesis in Ustilago maydis, a fungal pathogen that causes corn smut disease. The gene product is involved in the synthesis of the peptide backbone of the siderophore ferrichrome. We hope to exploit this gene to create a biotransformation system that can give rise to new antimicrobial drugs for control of human, animal and plant diseases. Such a system would generate new antimicrobial compounds in microorganisms based on the genes which are engineered into these microbes. If used in organisms that are useful in biofuel synthesis, these novel compounds may provide added value to these fermentations. This work will be of interest to biochemists and medical workers who seek new means of controlling iron toxicity in plants and people.

Technical Abstract: I.Ustilago maydis, the causal agent of corn smut disease, acquires and transports ferric ion by producing the extra-cellular, cyclic peptide, hydroxamate siderophores ferrichrome and ferrichrome A. Ferrichrome biosynthesis likely proceeds by hydroxylation and acetylation of L-ornithine and later steps likely involve covalently bound thioester intermediates on a multimodular, nonribosomal peptide synthetase. sid1 encodes L-ornithine N5-oxygenase, which catalyzes hydroxylation of L-ornithine, the first committed step of ferrichrome and ferrichrome A biosynthesis in U. maydis. In this report we characterize sid2, another biosynthetic gene in the pathway, by gene complementation, gene replacement, DNA sequence and Northern hybridization analysis. Nucleotide sequencing has revealed that sid2 is located 3.7-kb upstream of sid1 and encodes an intronless polypeptide of 3947 amino acids with three iterated modules of an approximate length of 1,000 amino acids each. Multiple motifs characteristic of the non-ribosomal peptide synthetase protein family were identified in each module. A corresponding iron-regulated sid2 transcript of 11 kb was detected by Northern hybridization analysis. By contrast, constitutive accumulation of this large transcript was observed in a mutant carrying a disruption of urbs1, a zinc-finger, GATA family transcription factor previously shown to regulate siderophore biosynthesis in Ustilago. Multiple GATA motifs are present in the intergenic region between sid1 and sid2, suggesting bi-directional transcription regulation by urbs1 of this pathway.