Nitric oxide (NO●) is important in cellular signaling and the innate immune response. It is synthesized by NO-synthases (NOS) from L-arginine in two steps through the intermediary Nω-hydroxy-L-arginine (NHA). In both steps P450-like reaction cycles are coupled with the redox cycle of the cofactor tetrahydrobiopterin (H4B). The mechanism of the second step, which is particularly elusive, is studied by DFT calculations. Our calculations show that the stabilizing effects of spin delocalization between substrate, cofactor and ferriheme-bound activated oxygen species along the reaction cycle direct the mechanism through the low-barrier pathway toward the intended products (Figure 1), whilst disruption of the prearranged balance of the active orbitals changes the reaction route leading to uncoupling. The pathway starts with proton transfer from one of heme propionates to the ferrous-dioxy FeIIO2 species (I) coupled with hole transfer from the bound dioxygen to the H4B though the filled bridge π-HOMO of the NHA oxime leading to the ferric-hydroperoxo complex (Cpd 0; II). Following the conformational transition within Cpd 0 resulting in the adjacent location of the OH-group of Cpd 0 and the guanidinium C of the NHA, the reaction proceeds through shallow reaction intermediates. First, proton-coupled electron transfer (PCET) from the NHA to the proximal O of Cpd 0 triggers peroxide bond cleavage (III) leading to the closed-shell π coordination complex of HO– and the NHA-derived iminoxy cation (IV), which turns into the adduct of protonated L-citrulline and NO– (V). Electron tunneling from this adduct to the H4B+● and PT to the ferriheme HO– complex releases the intended products (VI). Consequent formation and dissociation of the ferriheme nitrosyl complex completes the reaction cycle. Results are consistent with experimental data and support the functional role of heme propionates in PCET events in NOS.

Theoretical studies of the mechanism of the second half-reaction step of nitric oxide synthase: Nondynamical electron correlation drives the reaction to the intended products

253rd National Meeting (2017)

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