L-ornithine N5 monooxygenase
| L-ornithine N5 monooxygenase | |||||||||
|---|---|---|---|---|---|---|---|---|---|
 Ornithine monooxygenase tetramer, Aspergillus fumigatus | |||||||||
| Identifiers | |||||||||
| EC no. | 1.14.13.196 | ||||||||
| Databases | |||||||||
| IntEnz | IntEnz view | ||||||||
| BRENDA | BRENDA entry | ||||||||
| ExPASy | NiceZyme view | ||||||||
| KEGG | KEGG entry | ||||||||
| MetaCyc | metabolic pathway | ||||||||
| PRIAM | profile | ||||||||
| PDB structures | RCSB PDB PDBe PDBsum | ||||||||
| |||||||||
L-ornithine N5 monooxygenase (EC 1.14.13.195[1] or EC 1.14.13.196[2]) is an enzyme which catalyzes one of the following chemical reactions:
L-ornithine + NADPH + O2 N(5)-hydroxy-L-ornithine + NADP+ + H2O[1][3] L-ornithine + NAD(P)H + O2 N(5)-hydroxy-L-ornithine + NAD(P)+ + H2O [2][4]
The three ligands of this enzyme are L-ornithine (substrate), FAD (cofactor), and NADPH or NAD(P)H (cofactor and electron donor).[5]
Enzyme classification
[edit]L-ornithine N5 monooxygenase is classified under two EC numbers - EC1.14.13.195[1] and EC 1.14.13.196.[2] The first number, 1, identifies the enzyme as an oxidoreductase.[1][2] The subsequent 14 refers to the fact that this enzyme acts "on paired donors, with incorporation or reduction of molecular oxygen".[1][2] The 13 identifies this enzyme as using NADH or NAD(P)H as one donor, while incorporating one atom of oxygen onto the other.[1][2] This is why there are two EC numbers for this enzyme - one ends with 195 referring to NADPH as the donor, while the 196 refers to NAD(P)H as the donor.[1][2]
Structure
[edit]L-ornithine N5 monooxygenase adopbts an oxidoreductase Rossmann fold tertiary structure that binds FAD and NADP cofactors.[6]
Crystallographic structures have been solved for this class of enzymes from Aspergillus fumigatus.[7] These structures reflect structural changes which take place when the enzyme binds combinations of ligands, including ornithine and NADP.[7] Additional structures have also been solved for strain Af293.[8][9] These structures reflect different redox and ligation states.[5][8][9] The following table briefly describes these crystal structures:
| PDB Accession Code | Description | A. fumigatus strain | Source |
|---|---|---|---|
| 4NZH | r279a mutant | parent | [7] |
| 4B63 | bound to NADP and ornithine | " | [7] |
| 4B64 | bound to NADP and lysine | " | [7] |
| 4B65 | reduced and bound to NADPH | " | [7] |
| 4B66 | reduced and bound to NADP and arginine | " | [7] |
| 4B67 | re-oxidized and bound to NADP and ornithine | " | [7] |
| 4B68 | re-oxidized and bound to NADP and arginine | " | [7] |
| 4B69 | bound to ornithine | " | [7] |
| 5CKU | mutant N323A bound to NADP and ornithine | Af293 | [8] |
| 6X0H | oxidized | " | [5] |
| 6X0I | oxidized and bound to NADP+ | " | [5] |
| 6X0J | reduced and bound to NADP+ and L-ornithine | " | [5] |
| 6X0K | reduced and bound to L-ornithine | " | [5] |
| 7JVK | resting state | " | [9] |
| 7JVL | M101A variant complexed with NADP+ | " | [9] |
In A. fumigatus, the enzyme is named Af SidA for siderophore biosynthesis protein A.[7] It has three domains for ornithine (substrate), FAD (cofactor), and NAD(P)H (cofactor and electron donor).[5] The enzyme is a homotetramer.[7]
N-hydroxylating flavin-containing monooxygenase (NMO) enzymes such as this target the nucleophilic terminal amine groups of primary aliphatic amines such as L-ornithine.[7] The enzyme operates via a multistep oxidative mechanism which has a C4a-hydroperoxyflavin intermediate.[5] SidA stabilizes this intermediate and keeps NADP+ bound throughout the remainder of the catalytic cycle because it is necessary for intermediate stabilization.[7] The nicotinamide-ribose moiety and H-bonding between the main chain and residues Lys107, Asn293, and Ser469 position the L-ornithine alpha carbon such that its side chain amino group can be hydroxylated by the C4a-(hydro)peroxyflavin.[7] Unlike many other NMOs, A. fumigatus SidA strictly acts on ornithine.[7] Interactions with arginine increase interactivity between the reduced flavin and oxygen.[7]
The active site is located within a cleft at the interface between the three domains on each subunit. SidA has a resting state (6X0H) in which neither L-ornithine nor NAD(P)H is bound.[5] This resting state has an "out" active site caused by large rotations of the FAD isoalloxazine and a 10-Å movement of the Tyrosine loop.[5] Either flavin reduction or NAD(P)H binding drives the active site to the "in" conformation (6X0I).[5]
SidA demonstrates typical kinetics when saturated with L-ornithine.[10] Inhibition is caused by high concentrations of NADPH and NADH.[10] There is an 8-fold increase in catalytic efficiency for NADPH compared to NADH.[10] NADP+ is a competitive inhibitor with respect to NADPH.[10]
Function
[edit]This enzyme is widely distributed, especially among eukaryotes, being found in Fungi, Metazoa, Protista, Viridiplantae, Choanoflagellates, and Icththyosporeans.[11] Among Bacteria, it is found in Kutzneria sp. 744,[3][4] and an ornithine hydroxylase from Pseudomonas aeruginosa has a similar structure and 41% amino acid similarity to that of A. nidulans.[7]
In addition to being found in the non-pathogenic fungi such as Aspergillus nidulans, it is also found in many fungal pathogens such as Aspergillus fumigatus, Botrytis cinerea, Fusarium oxysporum, Magnaporthe oryzae, Sclerotinia sclerotiorum, Spizellomyces punctatus, and Ustilago maydis.[11]
In A. fumigatus, it is classified as a flavoprotein because FAD is a cofactor. It catalyzes the FAD and NADPH-dependent hydroxylation of L-ornithine in biosynthesis of the ferrichrome siderophores triacetylfusarinine and desferriferricrocin.[3][4][10] It is produced primarily under iron-limited conditions.[10] Siderophores are also important for virulence.[7]
In Kutzneria sp. 744, this enzyme is involved in the biosynthesis of piperazate, which contributes to the biosynthesis of kutzneride antifungal antibiotics.[3][4]
References
[edit]- ^ a b c d e f g "EC 1.14.13.195". IUBMB Enzyme Nomenclature. International Union of Biochemistry and Molecular Biology.
- ^ a b c d e f g "EC 1.14.13.196". IUBMB Enzyme Nomenclature. International Union of Biochemistry and Molecular Biology.
- ^ a b c d "EC 1.14.13.195". KEGG Enzyme. Kyoto Encyclopedia of Genes and Genomes (KEGG). Retrieved 2022-10-22.
- ^ a b c d "EC 1.14.13.195". KEGG Enzyme. Kyoto Encyclopedia of Genes and Genomes (KEGG). Retrieved 2022-10-22.
- ^ a b c d e f g h i j k Campbell AC, Stiers KM, Martin Del Campo JS, Mehra-Chaudhary R, Sobrado P, Tanner JJ (September 2020). "Trapping conformational states of a flavin-dependent N-monooxygenase in crystallo reveals protein and flavin dynamics". The Journal of Biological Chemistry. 295 (38): 13239–13249. doi:10.1074/jbc.RA120.014750. PMC 7504930. PMID 32723870.
- ^ Paul CE, Eggerichs D, Westphal AH, Tischler D, van Berkel WJ (November 2021). "Flavoprotein monooxygenases: Versatile biocatalysts". Biotechnology Advances. 51: 107712. doi:10.1016/j.biotechadv.2021.107712. PMID 33588053.
- ^ a b c d e f g h i j k l m n o p q r s Franceschini S, Fedkenheuer M, Vogelaar NJ, Robinson HH, Sobrado P, Mattevi A (September 2012). "Structural insight into the mechanism of oxygen activation and substrate selectivity of flavin-dependent N-hydroxylating monooxygenases". Biochemistry. 51 (36): 7043–7045. doi:10.1021/bi301072w. PMID 22928747.
- ^ a b c Robinson R, Qureshi IA, Klancher CA, Rodriguez PJ, Tanner JJ, Sobrado P (November 2015). "Contribution to catalysis of ornithine binding residues in ornithine N5-monooxygenase". Archives of Biochemistry and Biophysics. 585: 25–31. doi:10.1016/j.abb.2015.09.008. PMC 6467063. PMID 26375201.
- ^ a b c d Campbell AC, Robinson R, Mena-Aguilar D, Sobrado P, Tanner JJ (December 2020). "Structural Determinants of Flavin Dynamics in a Class B Monooxygenase". Biochemistry. 59 (48): 4609–4616. doi:10.1021/acs.biochem.0c00783. hdl:10919/107583. PMID 33226785.
- ^ a b c d e f Chocklett SW, Sobrado P (August 2010). "Aspergillus fumigatus SidA is a highly specific ornithine hydroxylase with bound flavin cofactor". Biochemistry. 49 (31): 6777–6783. doi:10.1021/bi100291n. PMID 20614882.
- ^ a b Zdobnov E. "Query 2679143". OrthoDB. Retrieved 2022-10-22.
