Formate C-acetyltransferase


In enzymology, formate C-acetyltransferase is an enzyme. Pyruvate formate lyase is found in Escherichia coli and other organisms. It helps regulate anaerobic glucose metabolism. Using radical non-redox chemistry, it catalyzes the reversible conversion of pyruvate and coenzyme-A into formate and acetyl-CoA. The reaction occurs as follows:
This enzyme belongs to the family of transferases, specifically those acyltransferases transferring groups other than aminoacyl groups. The systematic name of this enzyme class is acetyl-CoA:formate C-acetyltransferase. Other names in common use include pyruvate formate-lyase, pyruvic formate-lyase, and formate acetyltransferase. This enzyme participates in 3 metabolic pathways: pyruvate metabolism, propanoate metabolism, and butanoate metabolism.

Structural studies

As of late 2007, 8 structures have been solved for this class of enzymes, with PDB accession codes,,,,,,, and.
Pyruvate formate lyase is a homodimer made of 85 kDa, 759-residue subunits. It has a 10-stranded beta/alpha barrel motif into which is inserted a beta finger that contains major catalytic residues. The active site of the enzyme, elucidated by x-ray crystallography, holds three essential amino acids that perform catalysis, three major residues that hold the substrate pyruvate close by, and two flanking hydrophobic residues.
Studies have found structural similarities between the active site of pyruvate formate lyase and that of Class I and Class III ribonucleotide reductase enzymes.

Mechanism

Roles of the three catalytic residues

It has been shown that:
  1. The proposed mechanism for pyruvate formate lyase begins with radical transfer from Gly734 to Cys418, via Cys419.
  2. The Cys418 thiyl radical adds covalently to C2 of pyruvate, generating an acetyl-enzyme intermediate.
  3. The acetyl-enzyme intermediate releases a formyl radical that undergoes hydrogen-atom transfer with Cys419. This generates formate and a Cys419 radical.
  4. coenzyme-A comes in and undergoes hydrogen-atom transfer with the Cys419 radical to generate a coenzyme-A radical.
  5. The coenzyme-A radical then picks up the acetyl group from Cys418 to generate acetyl-CoA, leaving behind a Cys418 radical.
  6. Pyruvate formate lyase can then undergo radical transfer to put the radical back onto Gly734.
Note that each step is reversible.

Regulation

Two additional enzymes regulate the “on” and “off” states of pyruvate formate lyase to regulate anaerobic glucose metabolism: pyruvate formate lyase activase and pyruvate formate lyase deactivase. Activated pyruvate formate lyase allows formation of acetyl-CoA, a small molecule important in the production of energy, when pyruvate is available. Deactivated pyruvate formate lyase, even with substrates present, does not catalyze the reaction.
Pyruvate formate lyase activase is part of the radical SAM superfamily. The enzyme turns pyruvate formate lyase “on” by converting Gly734 into a Gly734 radical via a 5'-deoxyadenosyl radical.
For more information about radical SAM activation and radical SAM enzymes, see the discussion by Wang et al., 2007.
Pyruvate formate lyase deactivase turns pyruvate formate lyase “off” by quenching the Gly734 radical. Furthermore, pyruvate formate lyase is sensitive to molecular oxygen, the presence of which shuts the enzyme off.