Proton Delivery in NO Reduction by Fungal Nitric-oxide Reductase

Abstract

Fungal nitric-oxide reductase (NOR) is a heme enzyme that catalyzes the reduction of NO to Nitric oxide (NO)1 serves as a messenger molecule for a variety of biological functions, including neurotransmission, vascular relaxation, and the inhibition of platelet aggregation. In mammalian systems, NO is generated from L-arginine and molecular oxygen (O 2 ), via catalysis by heme-enzyme nitricoxide synthase, whose crystal structure has recently been reported (1-3). Subsequently, the generated NO binds to the heme iron of soluble guanylate cyclase activating the conversion of GTP to cGMP. In addition, the crystal structures of the heme-based NO transport protein, nitrophorin, of a blood sucking insect were recently reported (4,5). NO is also a potential ligand (inhibitor) of many hemoproteins such as myoglobin, hemoglobin, and peroxidase (6 -8). Despite the close relation of NO to hemoproteins, only a small amount of structural information is available for NO adduct of hemoproteins. It is particularly noteworthy that much less is known concerning the ferric-NO (Fe 3ϩ -NO) complex of hemoproteins, and only one crystal structure of cytochrome c peroxidase by Poulos and co-workers (8, 9) and two of the ferric-porphyrin model compounds, Fe Fungal nitric oxide reductase (NOR), a heme enzyme, is involved in the denitrification process by the fungus Fusarium oxysporum (11). In this process, NO is produced from the reduction of NO 2 Ϫ catalyzed by nitrite reductase, which represents an additional NO generating biological system. In order to detoxify the generated NO, fungal NOR converts NO to N 2 O by the reaction (11),Based on spectroscopic and kinetic studies of this reaction, we proposed that the overall enzymatic reaction (Scheme 1) consists of three chemical reactions, Schemes 2-4 (12);

Interaction of Nitric Oxide with Human Heme Oxygenase-1

“…This suggests that coordination of the NO to the heme iron is similar in both hemoproteins, although the broader bandwidth observed with the hHO-1 complex suggests that the NO has greater mobility in that protein. Generally, as is the case for the binding of NO to ferric hemoproteins, these results strongly argue that the NO is bound perpendicular to the heme face rather than at an angle (53)(54)(55)(56), in contrast to its orientation when bound to ferrous hemoproteins. Furthermore, in both met-Mb and ferric hHO-1-heme, the iron in the absence of NO is coordinated to a water molecule.…”

Fungal denitrification and nitric oxide reductase cytochrome P450nor

“…This interaction of Thr243 together with the propionate of haem moving upward restricts the conformation of the nicotinic acid ring so that the pro-R side of C4-hydrogens faces the haem, which is consistent with the pro-R hydrogen-specific hydride transfer [35]. A hydrogen bond network is formed to deliver a proton from solvent to Ser286 that is located in the close vicinity of haem [45]. However, the network is rearranged to form a proton channel upon binding of NAAD, and the bound NADH (NAAD) which is itself involved in the network [43] (figure 7), suggesting that a proton is supplied to the enzymatic reaction via the proton channel before formation of the intermediate (444 nm species; in the second step in figure 5).…”

“…dinucleotide; NAAD) [43]. The structure of the P450nor -NAAD complex is compared with that of the ferric -NO complex of P450nor [45] in figure 6. Little difference was observed between the structures of P450nor in the ferric resting state [46] and in the ferric -NO complex [45].…”

Myeloperoxidase up-regulates the catalytic activity of inducible nitric oxide synthase by preventing nitric oxide feedback inhibition

“…In this pathway, NADPH may interact with MPO, facilitate NO consumption through a mechanism similar to that previously reported for cytochrome P450 NOR and fungal nitric oxide reductase (NOR), and has been extensively studied with regard to its catalytic and structural properties (68,69). Cytochrome P450 NOR and fungal NOR have been known to participate in fungal denitrification by converting NO to N 2 O in a multiple-step sequential mechanism (68,69). Under our current experimental condition, this nonenzymatic pathway has little or no impact on iNOS catalytic activity and function because NO consumption by this system requires the presence of higher concentrations of NADPH.…”