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Abstract

Fatty Acid Photodecarboxylase (FAP) is a recently discovered photoenzyme that catalyzes the conversion of fatty acids into alkane
and CO2 under light, with potential importance in green chemistry applications [1]. Its mechanism was still not fully understood and
partly relied on a low-resolution crystal structure obtained from crystals with a twinning default [1]. Here, we present high-resolution
crystal structures of FAP obtained in the dark and after light illumination at cryogenic temperatures (Figure 1). Combined with
structural, computational, and spectroscopic techniques we are now able to provide a detailed reaction cycle of FAP. The reaction
mechanism starts with an electron transfer from the fatty acid to a photoexcited oxidized flavin cofactor. Decarboxylation yields an
alkyl radical, which is then reduced by back electron transfer and protonation rather than hydrogen atom transfer. Along with flavin
reoxidation by the alkyl radical intermediate, a major fraction of the cleaved CO2 unexpectedly transforms in 100 ns, most likely into
bicarbonate. This is orders of magnitude faster than in solution, which indicates a catalytic step. FT-IR, structural and functional
studies on variants centered on two conserved active site residues (R451 and C432) showed that R451 is essential for substrate
stabilization and proton transfer. Altogether this study provides a detailed characterization of this unique enzyme and reveals a striking
and unanticipated mechanistic complexity [2].