A Radical Intermediate in Tyrosine Scission to the CO and CN⁻ Ligands of FeFe Hydrogenase

The radical S-adenosylmethionine (SAM) enzyme HydG lyses free L-tyrosine to produce CO and CN⁻ for the assembly of the catalytic H cluster of FeFe hydrogenase. We used electron paramagnetic resonance spectroscopy to detect and characterize HydG reaction intermediates generated with a set of ²H, ¹³C, and ¹⁵N nuclear spin-labeled tyrosine substrates. We propose a detailed reaction mechanism in which the radical SAM reaction, initiated at an N-terminal 4Fe-4S cluster, generates a tyrosine radical bound to a C-terminal 4Fe-4S cluster. Heterolytic cleavage of this tyrosine radical at the C α -C β bond forms a transient 4-oxidobenzyl (4OB*) radical and a dehydroglycine bound to the C-terminal 4Fe-4S cluster. Electron and proton transfer to this 4OB* radical forms p-cresol, with the conversion of this dehydroglycine ligand to Fe-bound CO and CN⁻, a key intermediate in the assembly of the 2Fe subunit of the H cluster.