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Applied and Environmental Microbiology, June 2004, p. 3673-3680, Vol. 70, No. 6
0099-2240/04/$08.00+0 DOI: 10.1128/AEM.70.6.3673-3680.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.
Identification and Characterization of the Novel LysM Domain-Containing Surface Protein Sep from Lactobacillus fermentum BR11 and Its Use as a Peptide Fusion Partner in Lactobacillus and Lactococcus
Mark S. Turner,* Louise M. Hafner, Terry Walsh, and Philip M. GiffardInfectious Diseases Program, School of Life Sciences, Faculty of Science, Queensland University of Technology, Brisbane, Australia
Received 25 November 2003/ Accepted 18 February 2004
Examination of supernatant fractions from broth cultures of Lactobacillus fermentum BR11 revealed the presence of a number of proteins, including a 27-kDa protein termed Sep. The amino-terminal sequence of Sep was determined, and the gene encoding it was cloned and sequenced. Sep is a 205-amino-acid protein and contains a 30-amino-acid secretion signal and has overall homology (between 39 and 92% identity) with similarly sized proteins of Lactobacillus reuteri, Enterococcus faecium, Streptococcus pneumoniae, Streptococcus agalactiae, and Lactobacillus plantarum. The carboxy-terminal 81 amino acids of Sep also have strong homology (86% identity) to the carboxy termini of the aggregation-promoting factor (APF) surface proteins of Lactobacillus gasseri and Lactobacillus johnsonii. The mature amino terminus of Sep contains a putative peptidoglycan-binding LysM domain, thereby making it distinct from APF proteins. We have identified a common motif within LysM domains that is shared with carbohydrate binding YG motifs which are found in streptococcal glucan-binding proteins and glucosyltransferases. Sep was investigated as a heterologous peptide expression vector in L. fermentum, Lactobacillus rhamnosus GG and Lactococcus lactis MG1363. Modified Sep containing an amino-terminal six-histidine epitope was found associated with the cells but was largely present in the supernatant in the L. fermentum, L. rhamnosus, and L. lactis hosts. Sep as well as the previously described surface protein BspA were used to express and secrete in L. fermentum or L. rhamnosus a fragment of human E-cadherin, which contains the receptor region for Listeria monocytogenes. This study demonstrates that Sep has potential for heterologous protein expression and export in lactic acid bacteria.
* Corresponding author. Mailing address: Infectious Diseases Program, School of Life Sciences, Faculty of Science, Queensland University of Technology, G.P.O. Box 2434, Brisbane, Queensland 4001, Australia. Phone: (61-7) 3864-2567. Fax: (61-7) 3864-1534. E-mail: ms.turner{at}qut.edu.au.
Applied and Environmental Microbiology, June 2004, p. 3673-3680, Vol. 70, No. 6
0099-2240/04/$08.00+0 DOI: 10.1128/AEM.70.6.3673-3680.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.
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