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Genetics and Molecular Biology |
Department of Microbiology, Moyne Institute of Preventive Medicine, Trinity College, Dublin 2, Ireland1
Department of Pathology, University of Cambridge, Tennis Court Road, Cambridge CB2 1QP, UK2
Author for correspondence: Timothy J. Foster. Tel: +353 1 6082014. Fax: +353 1 6799294. e-mail: tfoster{at}tcd.ie
Staphylococcus epidermidis strains HB and K28 express surface proteins called Fbe or SdrG, respectively, that have sequence similarity to the clumping factors ClfA and ClfB of Staphylococcus aureus. A mutation in the fbe gene of strain HB was isolated by directed plasmid integration using the broad-host-range temperature-sensitive plasmid pG+Host9 (pVE6155). An internal fragment of fbe was cloned into pG+Host9 and the chimaeric plasmid was mobilized from S. aureus RN4220 to S. epidermidis 9142 by conjugation promoted by plasmid pGO1. The plasmid was then transferred to S. epidermidis strain HB by phage-48-mediated transduction. The plasmid integrated into the chromosomal fbe gene at a frequency of 2·8x10-4. All the survivors tested had a copy of pG+Host9‘fbe’ integrated into the chromosomal fbe gene either as a single copy or as a tandem array. Western immunoblotting showed that the wall-associated Fbe protein was absent in the mutant. Wild-type S. epidermidis HB adhered to immobilized fibrinogen in a dose-dependent and saturable fashion whereas the mutant did not bind. The Fbe proteins of HB and K28 were expressed at a high level in Lactococcus lactis MG1363 using the expression vector pKS80. These strains adhered strongly to immobilized fibrinogen. These results confirm that Fbe is a fibrinogen-binding adhesin.
Keywords: surface protein, Lactococcus lactis, adhesion, MSCRAMM
Abbreviations: MSCRAMM, microbial surface component recognizing adhesive matrix molecule
a These authors made an equal contribution.
b Present address: Plant Bioscience Limited, Norwich Research Park, Colney Lane, Norwich NR4 7UH, UK.
c Present address: Institute of Food Research, Norwich Research Park, Colney, Norwich NR4 7UA, UK.
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