The quorum-quenching lactonase from Alicyclobacter acidoterrestris: purification, kinetic characterization, crystallization and crystallographic analysis

Acta Crystallogr F Struct Biol Commun. 2017 Aug 1;73(Pt 8):476-480. doi: 10.1107/S2053230X17010640. Epub 2017 Jul 26.

Abstract

Lactonases comprise a class of enzymes that hydrolyze lactones, including acyl-homoserine lactones (AHLs); the latter are used as chemical signaling molecules by numerous Gram-negative bacteria. Lactonases have therefore been demonstrated to quench AHL-based bacterial communication. In particular, lactonases are capable of inhibiting bacterial behaviors that depend on these chemicals, such as the formation of biofilms or the expression of virulence factors. A novel representative from the metallo-β-lactamase superfamily, named AaL, was isolated from the thermoacidophilic bacterium Alicyclobacter acidoterrestris. Kinetic characterization proves AaL to be a proficient lactonase, with catalytic efficiencies (kcat/Km) against AHLs in the region of 105 M-1 s-1. AaL exhibits a very broad substrate specificity. Its structure is expected to reveal the molecular determinants for its substrate binding and specificity, as well as to provide grounds for future protein-engineering projects. Here, the expression, purification, characterization, crystallization and X-ray diffraction data collection of AaL at 1.65 Å resolution are reported.

Keywords: Alicyclobacter acidoterrestris; lactonases; quorum quenching; quorum sensing; thermophiles.

MeSH terms

  • Amino Acid Sequence
  • Bacterial Proteins / chemistry*
  • Bacterial Proteins / genetics
  • Bacterial Proteins / metabolism
  • Carboxylic Ester Hydrolases / chemistry*
  • Carboxylic Ester Hydrolases / genetics
  • Carboxylic Ester Hydrolases / metabolism
  • Cloning, Molecular
  • Crystallization
  • Crystallography, X-Ray
  • Escherichia coli / genetics
  • Escherichia coli / metabolism
  • Firmicutes / chemistry*
  • Firmicutes / metabolism
  • Gene Expression
  • Genetic Vectors / chemistry
  • Genetic Vectors / metabolism
  • Kinetics
  • Quorum Sensing / genetics*
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism

Substances

  • Bacterial Proteins
  • Recombinant Proteins
  • Carboxylic Ester Hydrolases
  • gluconolactonase