Unprecedented acetoacetyl-coenzyme A synthesizing enzyme of the thiolase superfamily involved in the mevalonate pathway

Proc Natl Acad Sci U S A. 2010 Jun 22;107(25):11265-70. doi: 10.1073/pnas.1000532107. Epub 2010 Jun 7.


Acetoacetyl-CoA is the precursor of 3-hydroxy-3-methylglutaryl (HMG)-CoA in the mevalonate pathway, which is essential for terpenoid backbone biosynthesis. Acetoacetyl-CoA is also the precursor of poly-beta-hydroxybutyrate, a polymer belonging to the polyester class produced by microorganisms. The de novo synthesis of acetoacetyl-CoA is usually catalyzed by acetoacetyl-CoA thiolase via a thioester-dependent Claisen condensation reaction between two molecules of acetyl-CoA. Here, we report that nphT7, found in the mevalonate pathway gene cluster from a soil-isolated Streptomyces sp. strain, encodes an unusual acetoacetyl-CoA synthesizing enzyme. The recombinant enzyme overexpressed in Escherichia coli catalyzes a single condensation of acetyl-CoA and malonyl-CoA to give acetoacetyl-CoA and CoA. Replacement of malonyl-CoA with malonyl-(acyl carrier protein) resulted in loss of the condensation activity. No acetoacetyl-CoA synthesizing activity was detected through the condensation of two molecules of acetyl-CoA. Based on these properties of NphT7, we propose to name this unusual enzyme of the thiolase superfamily acetoacetyl-CoA synthase. Coexpression of nphT7 with the HMG-CoA synthase gene and the HMG-CoA reductase gene in a heterologous host allowed 3.5-fold higher production of mevalonate than when only the HMG-CoA synthase and HMG-CoA reductase genes were expressed. This result suggests that nphT7 can be used to significantly increase the concentration of acetoacetyl-CoA in cells, eventually leading to the production of useful terpenoids and poly-beta-hydroxybutyrate.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acyl Coenzyme A / metabolism*
  • Amino Acid Sequence
  • Bacterial Proteins / genetics*
  • Bacterial Proteins / physiology
  • Chromatography, High Pressure Liquid
  • Coenzyme A Ligases / genetics*
  • Coenzyme A Ligases / physiology
  • Escherichia coli / metabolism*
  • Gene Expression Regulation, Bacterial*
  • Hydroxybutyrates / metabolism
  • Hydroxymethylglutaryl-CoA Synthase / genetics
  • Kinetics
  • Mevalonic Acid / metabolism*
  • Molecular Sequence Data
  • Multigene Family
  • Polyesters / metabolism
  • Recombinant Proteins / chemistry
  • Sequence Homology, Amino Acid
  • Streptomyces / enzymology*
  • Streptomyces / genetics*
  • Terpenes / chemistry


  • Acyl Coenzyme A
  • Bacterial Proteins
  • Hydroxybutyrates
  • Polyesters
  • Recombinant Proteins
  • Terpenes
  • acetoacetyl CoA
  • poly-beta-hydroxybutyrate
  • Hydroxymethylglutaryl-CoA Synthase
  • Coenzyme A Ligases
  • acetoacetyl-CoA synthase, Streptomyces
  • Mevalonic Acid

Associated data

  • GENBANK/AB540131