Differential active site loop conformations mediate promiscuous activities in the lactonase SsoPox

PLoS One. 2013 Sep 23;8(9):e75272. doi: 10.1371/journal.pone.0075272. eCollection 2013.


Enzymes are proficient catalysts that enable fast rates of Michaelis-complex formation, the chemical step and products release. These different steps may require different conformational states of the active site that have distinct binding properties. Moreover, the conformational flexibility of the active site mediates alternative, promiscuous functions. Here we focused on the lactonase SsoPox from Sulfolobus solfataricus. SsoPox is a native lactonase endowed with promiscuous phosphotriesterase activity. We identified a position in the active site loop (W263) that governs its flexibility, and thereby affects the substrate specificity of the enzyme. We isolated two different sets of substitutions at position 263 that induce two distinct conformational sampling of the active loop and characterized the structural and kinetic effects of these substitutions. These sets of mutations selectively and distinctly mediate the improvement of the promiscuous phosphotriesterase and oxo-lactonase activities of SsoPox by increasing active-site loop flexibility. These observations corroborate the idea that conformational diversity governs enzymatic promiscuity and is a key feature of protein evolvability.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Catalytic Domain / genetics*
  • Hydrolases / genetics
  • Hydrolases / metabolism*
  • Models, Molecular*
  • Molecular Structure
  • Mutation / genetics
  • Phosphoric Triester Hydrolases / genetics
  • Phosphoric Triester Hydrolases / metabolism*
  • Protein Conformation*
  • Substrate Specificity
  • Sulfolobus solfataricus / enzymology*
  • Sulfolobus solfataricus / genetics


  • Hydrolases
  • Phosphoric Triester Hydrolases

Grant support

This work was granted by Direction Générale de l'Armement (DGA), France (REI. 2009 34 0045) and Vaincre La Mucoviscidose, France. JH and GG are PhD students granted by DGA. ME is a fellow supported by the IEF Marie Curie program (grant No. 252836). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.