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Quorum Sensing pp 275-290 | Cite as

Heterologous Overexpression, Purification, and In Vitro Characterization of AHL Lactonases

  • Pei W. Thomas
  • Walter Fast
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 692)

Abstract

Quorum-quenching enzymes are useful as biochemical tools and possible therapeutic proteins. One of the best-characterized families of these catalysts is the N-acyl-l-homoserine lactone (AHL) lactonases, which rely on a dinuclear metal ion active site to hydrolytically cleave the autoinducer’s lactone bond and inactivate signaling. A detailed understanding of how this enzyme works can help in the design of more selective and efficient reagents. To facilitate these studies, we describe a methodology to heterologously express, purify, and conduct in vitro characterization of several metalloforms of the AHL lactonase from Bacillus thuringiensis (AiiA). These procedures should be applicable to similar enzymes and will facilitate the production of more useful quorum-quenching reagents for biochemical studies and possible therapeutic applications.

Key words

Quorum quenching Lactonase N-acyl-l-homoserine lactone Protein purification Metalloprotein Enzyme kinetics 

Notes

Acknowledgments

This work was supported in part by the Texas Advanced Research Program (Grant 003658-0018-2006) and the Robert A. Welch Foundation (Grant F-1572).

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Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  • Pei W. Thomas
    • 1
  • Walter Fast
    • 1
  1. 1.Division of Medicinal Chemistry, College of PharmacyUniversity of TexasAustinUSA

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