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Applied and Environmental Microbiology, March 2004, p. 1744-1748, Vol. 70, No. 3
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.3.1744-1748.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Controlled Autolysis and Enzyme Release in a Recombinant Lactococcal Strain Expressing the Metalloendopeptidase Enterolysin A

Rita M. Hickey,1,2 R. Paul Ross,1,3* and Colin Hill2,3

Teagasc Dairy Products Research Centre, Moorepark, Fermoy,1 Microbiology Department, University College Cork,2 Alimentry Pharmabiotic Centre, County Cork, Ireland3

Received 27 June 2003/ Accepted 24 November 2003

This study concerns the exploitation of the lytic enzyme enterolysin A (EntL), produced by Enterococcus faecalis strain DPC5280, to elicit the controlled autolysis of starter lactococci. EntL, a cell wall metalloendopeptidase secreted by some E. faecalis strains, can kill a wide range of gram-positive bacteria, including lactococci. The controlled expression of entL, which encodes EntL, was achieved using a nisin-inducible expression system in a lactococcal host. Zymographic analysis of EntL activity demonstrated that active enzyme is produced by the recombinant lactococcal host. Indeed, expression of EntL resulted in almost complete autolysis of the host strain 2 h after induction with nisin. Model cheese experiments using a starter strain in addition to the inducible enterolysin-producing strain showed a 27-fold increase in activity with respect to the release of lactate dehydrogenase in the strain overexpressing EntL, demonstrating the potential of EntL production in large-scale cheese production systems. Indeed, the observation that a wide range of lactic bacteria are sensitive to EntL suggests that EntL-induced autolysis has potential applications with a variety of lactic acid bacteria and could be a basis for probiotic delivery systems.

* Corresponding author. Mailing address: Teagasc Dairy Products Research Centre, Moorepark, Fermoy, Co. Cork, Ireland. Phone: 353-25-42229. Fax: 353-25-42340. E-mail: pross{at}moorepark.teagasc.ie.

Applied and Environmental Microbiology, March 2004, p. 1744-1748, Vol. 70, No. 3
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.3.1744-1748.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.





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