Premium
Molecular and functional analysis of the lipopolysaccharide biosynthesis locus wlb from Bordetella pertussis , Bordetella parapertussis and Bordetella bronchiseptica
Author(s) -
Allen Andrew G.,
Thomas Richard M.,
Cadisch Joanna T.,
Maskell Duncan J.
Publication year - 1998
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1046/j.1365-2958.1998.00878.x
Subject(s) - biology , bordetella bronchiseptica , bordetella pertussis , locus (genetics) , mutant , bordetella , genetics , microbiology and biotechnology , gene , bacteria
The Bordetella pertussis wlb locus ( wlb pe , formerly bpl ) is required for the biosynthesis of a trisaccharide that, when attached to the B. pertussis lipopolysaccharide (LPS) core (band B), generates band A LPS. The equivalent loci in Bordetella bronchiseptica ( wlb br ) and Bordetella parapertussis ( wlb pa ) were identified and cloned. The wlb br and wlb pa loci differ from wlb pe in that they lack the insertion sequence that defines the right‐hand terminus of wlb pe . Deletion of 12 kb of DNA containing the whole wlb locus (Δ wlb ) by allelic exchange in each of the three bordetellae had no effect on band B biosynthesis, whereas band A biosynthesis was prevented in B. pertussis and B. bronchiseptica . In B. bronchiseptica and B. parapertussis , Δ wlb mutants also lacked O‐antigen. Reintroduction of the wlb pe or wlb br loci on a shuttle vector into the three Δ wlb mutants restored the wild‐type LPS phenotype in the B. pertussis and B. bronchiseptica mutants. In the case of B. parapertussis , which normally does not synthesize an apparent band A structure, introduction of the wlb pe or wlb br loci now enabled the generation of band A. This suggests that the attachment point for band A trisaccharide on the LPS core is present in B. parapertussis , and further suggests that the wild‐type wlb pa locus is not fully functional. Introduction of the wlb pa locus into the Δ wlb pe , Δ wlb br and Δ wlb pa mutants had interesting consequences. The B. bronchiseptica and B. parapertussis recipients were now able to biosynthesize O‐antigen, but no band A was generated. In the B. pertussis recipient, a truncated band A was expressed consistent with a mutation in the wlbH gene, but on Western blotting the expression of a small amount of full‐length band A was also seen. Evidence that the wlbH pa protein is not fully functional was provided by the failure of the wlb pa locus to fully complement a B. pertussis wlbH (Δ wlbH pe ) mutant. This was supported by DNA sequence data showing that a single amino acid, conserved between homologous proteins from a range of bacteria, is altered in the B. parapertussis WlbH protein.