Purification and properties of pyrophosphatase of Acinetobacter johnsonii 210A and its involvement in the degradation of polyphosphate

Inorganic pyrophosphatase (E.C. 3.6.1.1) of Acinetobacter johnsonii 210A wa s purified 200-fold to apparent homogeneity. The enzyme catalyzed the hydro lysis of inorganic pyrophosphate and triphosphate to orthophosphate. No act ivity was observed with other polyphosphates and a wide variety of organic phosphate esters. The molecular mass of the enzyme was estimated to be 141 kDa by gelfiltration. Sodium dodecyl sulfate-polyacrylamide gel electrophor esis indicated a subunit composition of six identical polypeptides with a m olecular mass of 23 kDa. The cation Mg2+ was required for activity, the act ivity with Mn2+, Co2+ and Zn2+ was 48, 48 and 182% of the activity observed with Mg2+, respectively. The enzyme was heat-stable and inhibited by fluor ide and iodoacetamide. The analysis of the kinetic properties of the enzyme revealed an apparent K-m for pyrophosphate of 0.26 mM. In A. johnsonii 210 A, pyrophosphatase may be involved in the degradation of high-molecular pol yphosphates under anaerobic conditions: (i) it catalyses the further hydrol ysis of pyrophosphate and triphosphate formed from high-molecular weight po lyphosphates by the action of exopolyphosphatase, and (ii) it abolishes the inhibition of polyphosphate: AMP phosphotransferase-mediated degradation b y pyrophosphate and triphosphate.