Crystal structure of Ralstonia eutropha polyhydroxyalkanoate synthase C-terminal domain and reaction mechanisms

Biotechnol J. 2017 Jan;12(1). doi: 10.1002/biot.201600648. Epub 2016 Nov 30.


Polyhydroxyalkanoates (PHAs) are natural polyesters synthesized by numerous microorganisms as energy and reducing power storage materials, and have attracted much attention as substitutes for petroleum-based plastics. Here, we report the first crystal structure of Ralstonia eutropha PHA synthase at 1.8 Å resolution and structure-based mechanisms for PHA polymerization. RePhaC1 contains two distinct domains, the N-terminal (RePhaC1ND ) and C-terminal domains (RePhaC1CD ), and exists as a dimer. RePhaC1CD catalyzes polymerization via non-processive ping-pong mechanism using a Cys-His-Asp catalytic triad. Molecular docking simulation of 3-hydroxybutyryl-CoA to the active site of RePhaC1CD reveals residues involved in the formation of 3-hydroxybutyryl-CoA binding pocket and substrate binding tunnel. Comparative analysis with other polymerases elucidates how different classes of PHA synthases show different substrate specificities. Furthermore, we attempted structure-based protein engineering and developed a RePhaC1 mutant with enhanced PHA synthase activity.

Keywords: Crystal structure; Enzyme mechanism; PHA synthase; Polyhydroxyalkanoates; Ralstonia eutropha.

MeSH terms

  • Acyl Coenzyme A / metabolism
  • Acyltransferases / chemistry*
  • Acyltransferases / genetics
  • Acyltransferases / metabolism*
  • Catalytic Domain
  • Circular Dichroism
  • Crystallography, X-Ray
  • Cupriavidus necator / enzymology*
  • Models, Molecular
  • Molecular Docking Simulation
  • Mutagenesis
  • Protein Domains
  • Spectrometry, Fluorescence
  • Substrate Specificity


  • Acyl Coenzyme A
  • 3-hydroxybutyryl-coenzyme A
  • Acyltransferases
  • poly(3-hydroxyalkanoic acid) synthase