While most GRE contain only the conserved Gly and Cys residues as obvious active site elements, some are equipped with additional components required for activity. In particular, the PFL family contains two consecutive conserved Cys in place of just one in all other GRE (Cys418 and 419 in Escherichia coli), which leads to an extended hydrogen atom transfer cascade with two thiyl intermediates instead of the single one involved in other GRE. Both Cys are essential for the mechanism, as Cys419 is needed as 'radical hub' thiyl, which is involved in various intermediary partial reactions, while Cys418 binds covalently to the carbonyl carbon of pyruvate to activate the substrate for C-C cleavage to a formyl radical and an acetyl thioester, which is transferred to CoA . A second exception is known in ARNR because ribonucleotide reduction requires an additional redox step in addition to the radicaldependent activation reaction. Therefore, in addition to the bound ribonucleotide triphosphate, the active site accommodates an additional formate molecule as a reductant, which is intimately involved in the reaction mechanism . Moreover, the enzyme contains an Zn-binding domain missing in other GRE. Finally, the enzymes of the BSS and aromatic acid decarboxylase families contain additional subunits carrying FeS-clusters. In case of BSS, two small subunits with one Fe4S4 cluster each have been observed , whereas 4-hydroxyphenylacetate decarboxylase contains one small subunit with two Fe4S4 clusters . In either case, the functions of these subunits are still unclear. The glycyl radical has been characterized by EPR spectroscopy in many of these enzymes and they show remarkably similar spectroscopic features (Table ). The remaining H-atom at C of the glycyl radical residue causes a characteristic pronounced hyperfine splitting of the observed EPR spectra of 1.5 mT (1.7 mT for radiation-induced glycyl radicals in small model substrates ). In PFL and BSS, this hydrogen atom has been reported to be rapidly exchanged to deuterium in D2O-based solvents, while such an exchange did not occur in ARNR . For PFL, the activation reaction has been shown to be highly enantiospecific for the removal of pro-S hydrogen of glycine , while one of its two conserved cysteines (Cys419) was shown to be involved in the observed H/D isotope exchange of the glycyl radical . However, the detailed mechanism of this process is still unknown for any GRE.