The structures of Escherichia coli inorganic pyrophosphatase complexed with Ca2+ or CaPPi at atomic resolution and their mechanistic implications

Two structures of Escherichia coli soluble inorganic pyrophosphatase (EPPase) complexed with calcium pyrophosphate (CaPPi-EPPase) and with Ca2+ (Ca2+-EPPase) have been solved at 1.2 and 1.1 Å resolution, respectively. In the presence of Mg2+, this enzyme cleaves pyrophosphate (PPi) into two molecules of orthophosphate (Pi). This work has enabled us to locate PPi in the active site of the inorganic pyrophosphatases family in the presence of Ca2+, which is an inhibitor of EPPase. Upon PPi binding, two Ca2+ at M1 and M2 subsites move closer together and one of the liganded water molecules becomes bridging. The mutual location of PPi and the bridging water molecule in the presence of inhibitor cation is catalytically incompetent. To make a favourable PPi attack by this water molecule, modelling of a possible hydrolysable conformation of PPi in the CaPPi-EPPase active site has been performed. The reasons for Ca2+ being the strong PPase inhibitor and the role in catalysis of each of four metal ions are the mechanistic aspects discussed on the basis of the structures described.