Mechanism and Catalytic Site Atlas

Introduction

The reversible isomerisation reaction between M6P and F6P catalysed by PMI proceeds through a proton transfer between the two oxygen atoms O1 and O2 and a hydrogen transfer between the two carbon atoms C1 and C2 of the substrate and is thought to proceed through a “proton transfer mechanism” via a 1,2-cis-enediol(ate) high-energy intermediate.

After the ligand has been bound, it undergoes ring opening which is assisted by the zinc-bound water molecule and Gln111, displacement of electrons and protontropy between the oxygen atoms O1 and O5 induce cleavage of the C1--O5 bond and generate the open form of M6P. This ring opening is followed by conformational change of the substrate and water displacement: following displacement of the zinc-bound water molecule, zinc coordination of the O1 oxygen atom of M6P allows favourable interaction of its hydroxyl group on C2 with the nearby Lys136. The neutral Lys136-NH2 residue thereafter abstracts the hydrogen on C2 of M6P to yield the first 1,2-cis-enediolate HEI stabilised through interaction with Zn(II) and Lys136-NH. This is followed by proton transfer between O1 and O2, yielding the second 1,2-cis-enediolate HEI. Then the linear F6P is formed by protonation of the C1 carbon atom of the HEI on its Si face by Lys136-NH, regenerating the Lys136. Finally ring closure occurs after the entry of a new water molecule and cyclisation, β-D-fructofuranose 6-phosphate (β-F6P) is released from the active site upon binding of β-M6P.