Author: Ghavam, Nazanin
Aspergillus fumigatus is a major fungal pathogen of immunosuppressed patients. Recent studies demonstrated that hydroxamate-siderophore biosynthesis was required for virulence of A. fumigatus. SidA represents an excellent target for development of novel antifungal agents. In A. fumigatus the predominant hydroxamate-siderophore is N’, N’’, N’’’-triacetylfusarinine-C and its biosynthesis is initiated by hydroxylation of ornithine by ornithine-N5-oxygenase (SidA). The purpose of this research was to characterize SidA. AfSidA protein was cloned and expressed in an E. coli expression-vector. SidA was specific for its substrate L-ornithine, and cofactor FAD and preferred NADPH to NADH. L-lysine stimulated NADPH-oxidation but substrate hydroxylation was uncoupled. SidA had Michaelis-Menten kinetics and compared to bacterial ornithine-oxygenases, showed a higher thermotolerance and pH preference but a lower maximal velocity. A 3D-model of SidA was generated based on the crystal structure of phenylacetone-monooxygenase from Thermobifida fusca. Localization studies predicted a possible membrane association for SidA through its N-terminus.
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Thesis advisor: Moore, Margo M.
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