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Identification of cell wall proteins of Bacteroides fragilis to which bacteriophage B40-8 binds specifically

Departament de Microbiologia, Universitat de Barcelona, Av. Diagonal 645, 08028 Barcelona, Spain¹

Author for correspondence: Jorge Frias-Lopez. Tel: +34 393 580 9494. Fax: +34 393 590 9004. e-mail: frias{at}dentaid.es

Bacteriophage infecting Bacteroides fragilis, one of the most abundant bacteria in the human colon, have been proposed as indicators of virological faecal pollution. The first identification of a receptor for a bacteriophage in B. fragilis is reported here. First, resistant mutants were characterized following phage inactivation, and it was shown that cell wall proteins are involved in phage binding. Then the proteins involved were identified by various approaches: (i) comparison of the protein profiles of wild-type B. fragilis HSP40 and phage-resistant mutants; (ii) application of a modification of the virus overlay protein blot assay (VOPBA). At least two proteins of B. fragilis, with apparent molecular masses of 35 ± 5 kDa and 65 ± 5 kDa, bind to B40-8. This result was later confirmed by running a complex consisting of this phage bound to radiolabelled proteins of B. fragilis on an immunoaffinity column loaded with a specific antibody against the phage. Cell proteins retained in the column also coincided with the proteins that differed in the profiles of resistant mutants. Finally, to identify the potential function of these two proteins, their N-terminal sequences were determined and compared to published sequences, but no homologies were found.

Keywords: bacteriophage receptor, Bacteroides fragilis, cell wall proteins, VOPBA

Abbreviations: VOPBA, virus overlay protein blot assay

The PIR accession numbers for the sequences reported in this paper are A59325 (BactA) and B59325 (BactB).

^a Present address: DENTAID, Parc Tecnològic del Vallès, Ronda Can Fatjó, 10, 08290 Cerdanyola, Barcelona, Spain.

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