Allen A G, Thomas R M, Cadisch J T, Maskell D J
Department of Clinical Veterinary Medicine, University of Cambridge, UK.
Mol Microbiol. 1998 Jul;29(1):27-38. doi: 10.1046/j.1365-2958.1998.00878.x.
The Bordetella pertussis wlb locus (wlbpe, 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 (wlbbr) and Bordetella parapertussis (wlbpa) were identified and cloned. The wlbbr and wlbpa loci differ from wlbpe in that they lack the insertion sequence that defines the right-hand terminus of wlbpe. Deletion of 12 kb of DNA containing the whole wlb locus (delta 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, delta wlb mutants also lacked O-antigen. Reintroduction of the wlbpe or wlbbr loci on a shuttle vector into the three delta 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 wlbpe or wlbbr 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 wlbpa locus is not fully functional. Introduction of the wlbpa locus into the delta wlbpe, delta wlbbr and delta wlbpa 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 wlbHpa protein is not fully functional was provided by the failure of the wlbpa locus to fully complement a B. pertussis wlbH (delta wlbHpe) 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.
百日咳博德特氏菌的wlb基因座（wlbpe，以前称为bpl）是一种三糖生物合成所必需的基因座，该三糖连接到百日咳博德特氏菌脂多糖（LPS）核心（B带）上时会产生A带LPS。已鉴定并克隆了支气管败血博德特氏菌（wlbbr）和副百日咳博德特氏菌（wlbpa）中的等效基因座。wlbbr和wlbpa基因座与wlbpe的不同之处在于它们缺乏定义wlbpe右手末端的插入序列。通过等位基因交换在三种博德特氏菌中删除包含整个wlb基因座的12 kb DNA（δwlb）对B带生物合成没有影响，而百日咳博德特氏菌和支气管败血博德特氏菌中的A带生物合成受到抑制。在支气管败血博德特氏菌和副百日咳博德特氏菌中，δwlb突变体也缺乏O抗原。将穿梭载体上的wlbpe或wlbbr基因座重新导入三个δwlb突变体中，恢复了百日咳博德特氏菌和支气管败血博德特氏菌突变体中的野生型LPS表型。对于通常不合成明显A带结构的副百日咳博德特氏菌，引入wlbpe或wlbbr基因座现在能够产生A带。这表明副百日咳博德特氏菌中存在A带三糖在LPS核心上的附着点，并且进一步表明野生型wlbpa基因座功能不完全。将wlbpa基因座导入δwlbpe、δwlbbr和δwlbpa突变体中产生了有趣的结果。支气管败血博德特氏菌和副百日咳博德特氏菌受体现在能够生物合成O抗原，但没有产生A带。在百日咳博德特氏菌受体中，表达了与wlbH基因突变一致的截短A带，但在蛋白质免疫印迹中也观察到少量全长A带的表达。wlbpa基因座未能完全互补百日咳博德特氏菌wlbH（δwlbHpe）突变体，这证明了wlbHpa蛋白功能不完全。DNA序列数据支持了这一点，该数据表明在一系列细菌的同源蛋白之间保守的单个氨基酸在副百日咳博德特氏菌WlbH蛋白中发生了改变。
