One-carbon chemistry of oxalate oxidoreductase captured by X-ray crystallography

Abstract

Thiamine pyrophosphate (TPP)-dependent oxalate oxidoreductase (OOR) metabolizes oxalate, generating two molecules of CO 2 and two low-potential electrons, thus providing both the carbon and reducing equivalents for operation of the Wood−Ljungdahl pathway of acetogenesis. Here we present structures of OOR in which two different reaction intermediate bound states have been trapped: the covalent adducts between TPP and oxalate and between TPP and CO 2 . These structures, along with the previously determined structure of substrate-free OOR, allow us to visualize how active site rearrangements can drive catalysis. Our results suggest that OOR operates via a bait-and-switch mechanism, attracting substrate into the active site through the presence of positively charged and polar residues, and then altering the electrostatic environment through loop and side chain movements to drive catalysis. This simple but elegant mechanism explains how oxalate, a molecule that humans and most animals cannot break down, can be used for growth by acetogenic bacteria.oxalate | oxidoreductase | carbon dioxide | thiamine pyrophosphate

A Reverse TCA Cycle 2-Oxoacid:Ferredoxin Oxidoreductase that Makes C-C Bonds from CO2

“…Prior to our work, structures of OFORs have been limited to three different types, two PFORs (MtPFOR 7 and the cognate enzyme from Desulfovibiro africanus [DaPFOR] [20][21][22] ), an oxaloacetate oxidoreductase (OOR) from M. thermoacetica (MtOOR), 23,24 and two OFORs from Sulfolobus tokodaii of cryptic function (StOFORs). 25 Even with these structures in hand, many open questions about OFOR chemistry remain, including the issues of electron transfer described above.…”

A structural phylogeny for understanding 2-oxoacid oxidoreductase function

“…Crystal structures have been determined of two members of the OFOR superfamily: the PFOR from Desulfovibrio africanus ( Da PFOR) [24–26] and the OOR from M. thermoacetica ( Mt OOR) [21,22]. Both enzymes have similar overall structures despite the fact that Da PFOR is an α 2 homodimer and Mt OOR is a dimer of trimers (αγβ) 2 (Figure 1d,e,g).…”

“…Following TPP-adduct formation, however, this additional positive charge may hinder the next mechanistic steps, which are decarboxylation and electron transfer from TPP to the [4Fe-4S] clusters. Interestingly, crystal structures of OOR reveal that in the presence of substrate, a conformational change of a loop (aka the Switch loop) occurs that repositions Phe117α out of the active site (Figure 3b) and moves into the active site a negatively charged residue, Asp116α (Figure 3c) [22]. With this new loop conformation, Asp116α is proximal to the TPP-adduct, such that it could drive decarboxylation and/or electron transfer (Figure 3c).…”

“…With a second crystal structure of an OFOR finally determined a decade and a half after the first [21,22], we can now examine structure-function relationships in this important enzyme superfamily. Here we review what is known about the molecular basis of OFOR activity, focusing on PFOR and OOR.…”

Properties of Intermediates in the Catalytic Cycle of Oxalate Oxidoreductase and Its Suicide Inactivation by Pyruvate

“…18 In the OOR structure with carboxyl-TPP bound (PDB entry 5EXE), ∼50% of the switch loop is in the Asp-in conformation. In forming this Asp-in conformation, Arg109α swings away from its ionic interactions with carboxyl-TPP, observed in the Asp-out state, and Asp112α moves within H-bonding distance of the carboxyl group of carboxyl-TPP (Figure 9).…”