| Structural highlights
Function
[SIDA_ASPFU] Catalyzes the conversion of L-ornithine to N(5)-hydroxyornithine, the first step in the biosynthesis of all hydroxamate-containing siderophores, such as the secreted triacetylfusarinine C (TAFC) involved in iron uptake and the intracellular iron storage compound desferriferricrocin (DFFC). Highly specific for its substrate, only hydrolyzing l-ornithine. Has preference for NADPH over NADH, NADPH playing a role in stabilization of the C4a-hydroperoxyflavin intermediate. Essential for virulence.[1] [2] [3] [4]
Publication Abstract from PubMed
The SidA ornithine hydroxylase from Aspergillus fumigatus is a fungal disease drug target involved in the production of hydroxamate containing siderophores, which are used by the pathogen to sequester iron. SidA is an N-monooxygenase that catalyzes the NADPH-dependent hydroxylation of L-ornithine through a multistep oxidative mechanism, utilizing a C4a-hydroperoxyflavin intermediate. Here we present four new crystal structures of SidA in various redox- and ligation states, including the first structure of oxidized SidA without NADP(H) or L-ornithine bound (resting state). The resting state structure reveals a new "out" active site conformation characterized by large rotations of the FAD isoalloxazine around the C1'-C2' and N10-C1' bonds, coupled to a 10 A movement of the Tyr-loop. Additional structures show that either flavin reduction or the binding of NADP(H) is sufficient to drive the FAD to the "in" conformation. The structures also reveal protein conformational changes associated with the binding of NADP(H) and L-ornithine. Some of these residues were probed using site-directed mutagenesis. Docking was used to explore the active site of the "out" conformation. These calculations identified two potential ligand-binding sites. Altogether, our results provide new information about conformational dynamics in flavin-dependent monooxygenases. Understanding the different active site conformations that appear during the catalytic cycle may allow fine tuning of inhibitor discovery efforts.
Trapping conformational states of a flavin-dependent N-monooxygenase in crystallo reveals protein and flavin dynamics.,Campbell AC, Stiers KM, Martin Del Campo JS, Mehra-Chaudhary R, Sobrado P, Tanner JJ J Biol Chem. 2020 Jul 28. pii: RA120.014750. doi: 10.1074/jbc.RA120.014750. PMID:32723870[5]
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.
See Also
References
- ↑ Schrettl M, Bignell E, Kragl C, Joechl C, Rogers T, Arst HN Jr, Haynes K, Haas H. Siderophore biosynthesis but not reductive iron assimilation is essential for Aspergillus fumigatus virulence. J Exp Med. 2004 Nov 1;200(9):1213-9. Epub 2004 Oct 25. PMID:15504822 doi:http://dx.doi.org/jem.20041242
- ↑ Hissen AH, Wan AN, Warwas ML, Pinto LJ, Moore MM. The Aspergillus fumigatus siderophore biosynthetic gene sidA, encoding L-ornithine N5-oxygenase, is required for virulence. Infect Immun. 2005 Sep;73(9):5493-503. PMID:16113265 doi:http://dx.doi.org/10.1128/IAI.73.9.5493-5503.2005
- ↑ Chocklett SW, Sobrado P. Aspergillus fumigatus SidA is a highly specific ornithine hydroxylase with bound flavin cofactor. Biochemistry. 2010 Aug 10;49(31):6777-83. doi: 10.1021/bi100291n. PMID:20614882 doi:http://dx.doi.org/10.1021/bi100291n
- ↑ Romero E, Fedkenheuer M, Chocklett SW, Qi J, Oppenheimer M, Sobrado P. Dual role of NADP(H) in the reaction of a flavin dependent N-hydroxylating monooxygenase. Biochim Biophys Acta. 2012 Jun;1824(6):850-7. doi: 10.1016/j.bbapap.2012.03.004. , Epub 2012 Mar 27. PMID:22465572 doi:http://dx.doi.org/10.1016/j.bbapap.2012.03.004
- ↑ Campbell AC, Stiers KM, Martin Del Campo JS, Mehra-Chaudhary R, Sobrado P, Tanner JJ. Trapping conformational states of a flavin-dependent N-monooxygenase in crystallo reveals protein and flavin dynamics. J Biol Chem. 2020 Jul 28. pii: RA120.014750. doi: 10.1074/jbc.RA120.014750. PMID:32723870 doi:http://dx.doi.org/10.1074/jbc.RA120.014750
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