Rational design of allosteric regulation of homoserine dehydrogenase by a nonnatural inhibitor L-lysine

ACS Synth Biol. 2015 Feb 20;4(2):126-31. doi: 10.1021/sb400133g. Epub 2014 Jan 6.


Allosteric proteins, which can sense different signals, are interesting biological parts for synthetic biology. In particular, the design of an artificial allosteric enzyme to sense an unnatural signal is both challenging and highly desired, for example, for a precise and dynamical control of fluxes of growth-essential but byproduct pathways in metabolic engineering of industrial microorganisms. In this work, we used homoserine dehydrogenase (HSDH) of Corynebacterium glutamicum, which is naturally allosterically regulated by threonine and isoleucine, as an example to demonstrate the feasibility of reengineering an allosteric enzyme to respond to an unnatural inhibitor L-lysine. For this purpose, the natural threonine binding sites of HSD were first predicted and verified by mutagenesis experiments. The threonine binding sites were then engineered to a lysine binding pocket. The reengineered HSD only responds to lysine inhibition but not to threonine. This is a significant step toward the construction of artificial molecular circuits for dynamic control of growth-essential byproduct formation pathway for lysine biosynthesis.

Keywords: allosteric regulation; dynamic flux control; homoserine dehydrogenase; lysine; protein design.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Allosteric Regulation
  • Amino Acid Substitution
  • Binding Sites
  • Corynebacterium / enzymology
  • Drug Design
  • Enzyme Inhibitors / chemistry*
  • Enzyme Inhibitors / metabolism
  • Homoserine Dehydrogenase / antagonists & inhibitors*
  • Homoserine Dehydrogenase / genetics
  • Homoserine Dehydrogenase / metabolism
  • Isoleucine / chemistry
  • Isoleucine / metabolism
  • Kinetics
  • Lysine / chemistry*
  • Lysine / metabolism
  • Molecular Dynamics Simulation
  • Protein Binding
  • Protein Engineering
  • Recombinant Proteins / biosynthesis
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Threonine / chemistry
  • Threonine / metabolism


  • Enzyme Inhibitors
  • Recombinant Proteins
  • Isoleucine
  • Threonine
  • Homoserine Dehydrogenase
  • Lysine