Theoretical study of the nitrosation reaction mechanism of N-nitrosation reaction catalyzed by the cupin domain of SznF enzyme toward the modified L-Arg with methyl nitrogen deprotonation

The SznF enzyme provides a novel approach to synthesize the N-nitrosourea pharmacophore. In this work, the modified L-Arg with methyl nitrogen deprotonation(L-DHMAd), a not yet studied substrate's reaction mechanism of the critical N-nitrosation reaction catalyzed by the cupin domain of SznF enzyme is investigated, through the combined quantum mechanics/molecular mechanics method. Basis on our calculation results, the optimal N-nitrosation reaction pathway can be divided in to three reaction steps:1) superoxo addition with accompanying of Cε-Nω bond cleavage; 2) O1-O2 bond heterolytic cleavage; 3) NMe-Nω bond coupling. The first reaction step is the rate-determining step of the entire optimal reaction with an energy barrier of 20.6 Kcal/mol. Significantly, Cε-Nω bond could be homolytic cleaved spontaneously during the Cε-O2 bond formation in the first reaction step. This appearance mainly attributes to the Cε-Nω single bond of the L-DHMAd, which could be more feasible to cleave to maintain the Cε sp2 hybridization within the superoxo addition. The subsequent reaction step is the O1-O2 bond heterolytic cleavage, which involves a minimum energy crossing point to form the stable intermediate with Fe(IV) = O1 species at quintet state. Finally, NMe and Nω of L-DHMAd are very energetic favorable to coupling to form the N-nitroso product. This is consist with the experimental observations that two nitrogen atoms of the N-nitroso product are both from the same arginine substrate. Our work could contribute to the deeper understanding of the N-nitrosation reaction catalyzed by SznF enzyme, and might enlighten further studies of biomimetic chemistry of SznF enzyme.