Reduction of Fluorinated Cyclopropene by Nitrogenase

Reduction of the first known halogen-containing substrate by nitrogenase (N2ase), 3,3-difluorocyclopropene (DFCP), was investigated. Reduction requires both N2ase proteins (MoFe and Fe protein), ATP, and an exogenous reductant (dithionite, DT), as with N2 and known alternative substrates of the enzyme. Two major products providing evidence for reductive C-F bond cleavage were confirmed, propene (P1, requiring 6e(-)/6H(+)) and 2-fluoropropene (P2, 4e(-)/4H(+)). Both were identified by GC-MS and NMR spectroscopy, and had the same Km constants (0.022 atm, 5.4 mM). Reduction of 1,2-dideuterated DFCP (d2-DFCP) further revealed that (i) in both P1 and P2, two deuterium atoms are retained, one on carbon-1 and one on carbon-3, indicating that C═C bond cleavage rather than C-C bond cleavage is involved during DFCP reduction at least to P2 (assuming no F migration); (ii) no selectivity was observed in formation of cis and trans isomers of 1,3-d2-2-fluoropropene, whereas cis-1,3-d2-propene is the predominant 1,3-d2-propene product, indicating that one of the bound reduction intermediates on the pathway to propene is constrained geometrically. A reduction mechanism, consistent with hydride transfer as a key step, is discussed. Reductive C-F bond cleavage is an ability of N2ase that further demonstrates the unique and remarkable scope of its catalytic prowess.