Mechanistic Insights into the N-Hydroxylations Catalyzed by the Binuclear Iron Domain of SznF Enzyme: Key Piece in the Synthesis of Streptozotocin.

SznF, a member of the emerging family of heme-oxygenase-like (HO-like) di-iron oxidases and oxygenases, employs two distinct domains to catalyze the conversion of N ω -methyl-L-arginine (L-NMA) into N-nitroso-containing product, which can subsequently be transformed into streptozotocin. Using unrestricted density functional theory (UDFT) with the hybrid functional B3LYP, we have mechanistically investigated the two sequential hydroxylations of L-NMA catalyzed by SznF's binuclear iron central domain. Mechanism B primarily involves the O-O bond dissociation, forming Fe(IV)=O, induced by the H + /e - introduction to the Fe A side of μ-1,2-peroxo-Fe 2 (III/III), the substrate hydrogen abstraction by Fe(IV)=O, and the hydroxyl rebound to the substrate N radical. The stochastic addition of H + /e - to the Fe B side (mechanism C) can transition to mechanism B, thereby preventing enzyme deactivation. Two other competing mechanisms, involving the direct O-O bond dissociation (mechanism A) and the addition of H 2 O as a co-substrate (mechanism D), have been ruled out.