Overproduction, crystallization and X-ray diffraction data analysis of ectoine synthase from the cold-adapted marine bacterium Sphingopyxis alaskensis

Acta Crystallogr F Struct Biol Commun. 2015 Aug;71(Pt 8):1027-32. doi: 10.1107/S2053230X15011115. Epub 2015 Jul 28.


Ectoine biosynthetic genes (ectABC) are widely distributed in bacteria. Microorganisms that carry them make copious amounts of ectoine as a cell protectant in response to high-osmolarity challenges. Ectoine synthase (EctC; EC is the key enzyme for the production of this compatible solute and mediates the last step of ectoine biosynthesis. It catalyzes the ring closure of the cyclic ectoine molecule. A codon-optimized version of ectC from Sphingopyxis alaskensis (Sa) was used for overproduction of SaEctC protein carrying a Strep-tag II peptide at its carboxy-terminus. The recombinant SaEctC-Strep-tag II protein was purified to near-homogeneity from Escherichia coli cell extracts by affinity chromatography. Size-exclusion chromatography revealed that it is a dimer in solution. The SaEctC-Strep-tag II protein was crystallized using the sitting-drop vapour-diffusion method and crystals that diffracted to 1.0 Å resolution were obtained.

Keywords: X-ray analysis; chemical chaperone; compatible solute; cupin; ectoine synthesis; enzyme; osmostress protectant.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adaptation, Physiological
  • Amino Acid Sequence
  • Aquatic Organisms
  • Bacterial Proteins / chemistry*
  • Bacterial Proteins / genetics
  • Cloning, Molecular
  • Cold Temperature
  • Crystallization
  • Crystallography, X-Ray
  • Escherichia coli / genetics
  • Escherichia coli / metabolism
  • Gene Expression
  • Hydro-Lyases / chemistry*
  • Hydro-Lyases / genetics
  • Molecular Sequence Data
  • Recombinant Fusion Proteins / chemistry*
  • Recombinant Fusion Proteins / genetics
  • Sequence Alignment
  • Sphingomonadaceae / chemistry*
  • Sphingomonadaceae / enzymology


  • Bacterial Proteins
  • Recombinant Fusion Proteins
  • Hydro-Lyases
  • ectoine synthase