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Applied and Environmental Microbiology, June 2009, p. 3554-3563, Vol. 75, No. 11
0099-2240/09/$08.00+0     doi:10.1128/AEM.02919-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Identification of a Gene Cluster for the Biosynthesis of a Long, Galactose-Rich Exopolysaccharide in Lactobacillus rhamnosus GG and Functional Analysis of the Priming Glycosyltransferase{triangledown} ,{dagger}

Sarah Lebeer,1 Tine L. A. Verhoeven,1 Grégory Francius,2 Geert Schoofs,1 Ivo Lambrichts,3 Yves Dufrêne,2 Jos Vanderleyden,1 and Sigrid C. J. De Keersmaecker1*

Centre of Microbial and Plant Genetics, Katholieke Universiteit Leuven, Kasteelpark Arenberg 20, B-3001 Leuven, Belgium,1 Unité de Chimie des Interfaces, Université Catholique de Louvain, Croix du Sud 2/18, B-1348 Louvain-la-Neuve, Belgium,2 Laboratory of Histology, BIOMED Research Institute, University Hasselt, Agoralaan Gebouw D, 3590 Diepenbeek, Belgium3

Received 22 December 2008/ Accepted 27 March 2009

Cell surface polysaccharides have an established role as virulence factors in human bacterial pathogens. Less documented are the biosynthesis and biological functions of surface polysaccharides in beneficial bacteria. We identified a gene cluster that encodes the enzymes and regulatory and transporter proteins for the different steps in the biosynthesis of extracellular polysaccharides (EPS) of the well-documented probiotic strain Lactobacillus rhamnosus GG. Subsequent mutation of the welE gene, encoding the priming glycosyltransferase within this cluster, and comparative phenotypic analyses of wild-type versus mutant strains confirmed the specific function of this gene cluster in the biosynthesis of high-molecular-weight, galactose-rich heteropolymeric EPS molecules. The phenotypic analyses included monomer composition determination, estimation of the polymer length of the isolated EPS molecules, and single-molecule force spectroscopy of the surface polysaccharides. Further characterization of the welE mutant also showed that deprivation of these long, galactose-rich EPS molecules results in an increased adherence and biofilm formation capacity of L. rhamnosus GG, possibly because of less shielding of adhesins such as fimbria-like structures.

* Corresponding author. Mailing address: Centre of Microbial and Plant Genetics, K.U. Leuven, Kasteelpark Arenberg 20, 3001 Leuven, Belgium. Phone: 32 16 321631. Fax: 32 16 321966. E-mail: sigrid.dekeersmaecker{at}biw.kuleuven.be

{triangledown} Published ahead of print on 3 April 2009.

{dagger} Supplemental material for this article may be found at http://aem.asm.org/.

Applied and Environmental Microbiology, June 2009, p. 3554-3563, Vol. 75, No. 11
0099-2240/09/$08.00+0     doi:10.1128/AEM.02919-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.



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