YfiD of Escherichia coli and Y06I of Bacteriophage T4 as Autonomous Glycyl Radical Cofactors Reconstituting the Catalytic Center of Oxygen-Fragmented Pyruvate Formate-Lyase

Structures of benzylsuccinate synthase elucidate roles of accessory subunits in glycyl radical enzyme activation and activity

“…S9), has a repair mechanism to restore enzyme activity upon oxygen damage. In particular, repair involves a "spare part" protein, YfiD, which is posttranslationally activated by PFL-AE and replaces the oxygendamaged glycyl radical domain, yielding active enzyme (26). As more GRE enzymes are characterized, it will be interesting to see if structural features surrounding the glycyl radical domain correlate with the likelihood of oxygen exposure or the presence of repair mechanisms.…”

Structural basis for glycyl radical formation by pyruvate formate-lyase activating enzyme

“…In this circumstance, some organisms produce a short protein with high homology to the last 59 residues of PFL (including G 734 ), called YfiD in E. coli. After activation by PFL-AE, YfiD can associate with cleaved PFL, replacing its lost radical-containing domain and yielding a working heteroenzyme complex (27). In this manner, YfiD acts as a ''spare part,'' restoring activity of oxygen-cleaved PFL with minimal energy expenditure.…”

“…S6 a-c). The best docking model was obtained by using the pept-AE structure and a fragment cut from the PFL model [Protein Data Bank (PDB) ID code 2PFL] with high homology to YfiD (the small protein capable of undergoing activation by PFL-AE) (27). This region of PFL, which was implicated as being involved in the activation reaction on the basis of high structural homology to the corresponding region of the GRE glycerol dehydratase (22), will be referred to as a ''radical domain'' (RD) and encompasses PFL residues 712-759 (Fig.…”

Pyruvate Formate-lyase, Evidence for an Open Conformation Favored in the Presence of Its Activating Enzyme

“…(i) Activated PFL is stable only under strictly anaerobic conditions; in the presence of O 2 , activated PFL is cleaved at the site of the glycyl radical (Gly-734) to form two fragments of 82 kDa (residues 1-733) and 3 kDa. (ii) The O 2 -mediated cleavage of PFL is irreversible and results in complete loss of enzyme activity; however YfiD, a 14-kDa protein from E. coli, can interact with the 82-kDa PFL fragment to recover the full activity of PFL (40). YfiD has substantial sequence homology to the C-terminal portion of PFL (residues 734 -759) and is thought to possibly form a complex with the PFL(1-733) peptide (40).…”

“…(ii) The O 2 -mediated cleavage of PFL is irreversible and results in complete loss of enzyme activity; however YfiD, a 14-kDa protein from E. coli, can interact with the 82-kDa PFL fragment to recover the full activity of PFL (40). YfiD has substantial sequence homology to the C-terminal portion of PFL (residues 734 -759) and is thought to possibly form a complex with the PFL(1-733) peptide (40). These results with YfiD suggest that the C-terminal portion of PFL, which includes Gly-734, can function in a somewhat modular fashion, such that when the protein is cleaved at Gly-734, the YfiD module can "plug in" to regenerate a functional enzyme.…”