Helix swapping between two α/β barrels: crystal structure of phosphoenolpyruvate mutase with bound Mg2+–oxalate

nature and, when linked to macromolecules in eukaryotes, they might play a role in cell signaling. The first obligatory step in the biosynthesis of phosphonates is the formation of a carbon–phosphorus bond by converting phosphoenolpyruvate (PEP) to phosphonopyruvate (P-pyr), a reaction that is catalyzed by PEP mutase. The PEP mutase functions as a tetramer and requires magnesium ions (Mg2+). Results: The crystal structure of PEP mutase from the mollusk Mytilus edulis, bound to the inhibitor Mg2+–oxalate, has been determined using multiwavelength anomalous diffraction, exploiting the selenium absorption edge of a selenomethionine-containing protein. The structure has been refined at 1.8 Å resolution. PEP mutase adopts a modified α/β barrel fold, in which the eighth α helix projects away from the α/β barrel instead of packing against the β sheet. A tightly associated dimer is formed, such that the two eighth helices are swapped, each packing against the β sheet of the neighboring molecule. A dimer of dimers further associates into a tetramer. Mg2+–oxalate is buried close to the center of the barrel, at the C-terminal ends of the β strands. Conclusions: The tetramer observed in the crystal is likely to be physiologically relevant. Because the Mg2+–oxalate is inaccessible to solvent, substrate binding and dissociation might be accompanied by conformational changes. A mechanism involving a phosphoenzyme intermediate is proposed, with Asp58 acting as the nucleophilic entity that accepts and delivers the phosphoryl group. The active-site architecture and the chemistry performed by PEP mutase are different from other α/β-barrel proteins that bind pyruvate or PEP, thus the enzyme might represent a new family of α/β-barrel proteins