Purification, crystallization and X-ray crystallographic analysis of a putative exopolyphosphatase from Zymomonas mobilis

Exopolyphosphatase (PPX) enzymes degrade inorganic polyphosphate (poly‐P), which is essential for the survival of microbial cells in response to external stresses. In this study, a putative exopolyphosphatase from Zymomonas mobilis (ZmPPX) was crystallized. Crystals of the wild‐type enzyme diffracted to 3.3 Å resolution and could not be optimized further. The truncation of 29 amino acids from the N‐terminus resulted in crystals that diffracted to 1.8 Å resolution. The crystals belonged to space group C2, with unit‐cell parameters a = 122.0, b = 47.1, c = 89.5 Å, α = γ = 90, β = 124.5°. An active‐site mutant that crystallized in the same space group and with similar unit‐cell parameters diffracted to 1.56 Å resolution. One molecule was identified per asymmetric unit. Analytical ultracentrifugation confirmed that ZmPPX forms a dimer in solution. It was confirmed that ZmPPX possesses exopolyphosphatase activity against a synthetic poly‐P substrate. A putative exopolyphosphatase (PPX) from Z. mobilis (ZmPPX) was expressed, purified and crystallized. PPX enzymes regulate intracellular inorganic polyphosphate levels in microbial cells in response to external stresses. N‐terminal truncation of ZmPPX based on primary‐sequence analysis resulted in an improvement in diffraction from 3.3 to 1.8 Å resolution.