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

Acta Crystallogr F Struct Biol Commun. 2016 Mar;72(Pt 3):172-8. doi: 10.1107/S2053230X16000753. Epub 2016 Feb 19.

Abstract

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.

Keywords: PPX/GppA family; Zymomonas mobilis; crystallization; exopolyphosphatase.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acid Anhydride Hydrolases / chemistry*
  • Acid Anhydride Hydrolases / isolation & purification
  • Bacterial Proteins / chemistry*
  • Bacterial Proteins / isolation & purification
  • Chromatography, Gel
  • Crystallization
  • Crystallography, X-Ray
  • Ultracentrifugation
  • Zymomonas / enzymology*

Substances

  • Bacterial Proteins
  • Acid Anhydride Hydrolases
  • exopolyphosphatase