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Applied and Environmental Microbiology, May 1999, p. 2078-2083, Vol. 65, No. 5
0099-2240/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Identification and Disruption of BetL, a Secondary Glycine Betaine Transport System Linked to the Salt Tolerance of Listeria monocytogenes LO28

Roy D. Sleator,1 Cormac G. M. Gahan,1 Tjakko Abee,2 and Colin Hill1,*

Department of Microbiology and National Food Biotechnology Centre, University College Cork, Cork, Ireland,1 and Department of Food Science, Wageningen Agricultural University, Wageningen, The Netherlands2

Received 30 November 1998/Accepted 5 March 1999

The trimethylammonium compound glycine betaine (N,N,N-trimethylglycine) can be accumulated to high intracellular concentrations, conferring enhanced osmo- and cryotolerance upon Listeria monocytogenes. We report the identification of betL, a gene encoding a glycine betaine uptake system in L. monocytogenes, isolated by functional complementation of the betaine uptake mutant Escherichia coli MKH13. The betL gene is preceded by a consensus sigma B-dependent promoter and is predicted to encode a 55-kDa protein (507 amino acid residues) with 12 transmembrane regions. BetL exhibits significant sequence homologies to other glycine betaine transporters, including OpuD from Bacillus subtilis (57% identity) and BetP from Corynebacterium glutamicum (41% identity). These high-affinity secondary transporters form a subset of the trimethylammonium transporter family specific for glycine betaine, whose substrates possess a fully methylated quaternary ammonium group. The observed Km value of 7.9 µM for glycine betaine uptake after heterologous expression of betL in E. coli MKH13 is consistent with values obtained for L. monocytogenes in other studies. In addition, a betL knockout mutant which is significantly affected in its ability to accumulate glycine betaine in the presence or absence of NaCl has been constructed in L. monocytogenes. This mutant is also unable to withstand concentrations of salt as high as can the BetL+ parent, signifying the role of the transporter in Listeria osmotolerance.

* Corresponding author. Mailing address: Department of Microbiology, University College Cork, Cork, Ireland. Phone: 353-21-902397. Fax: 353-21-903101. E-mail: c.hill{at}ucc.ie.

Applied and Environmental Microbiology, May 1999, p. 2078-2083, Vol. 65, No. 5
0099-2240/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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