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首页> 外文期刊>Journal of bacteriology >Effect of wzx (rfbX) Mutations on A-Band and B-Band Lipopolysaccharide Biosynthesis in Pseudomonas aeruginosa O5
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Effect of wzx (rfbX) Mutations on A-Band and B-Band Lipopolysaccharide Biosynthesis in Pseudomonas aeruginosa O5

机译：wzx（rfbX）突变对铜绿假单胞菌O5中A波段和B波段脂多糖生物合成的影响

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The wbp cluster of Pseudomonas aeruginosaO5 encodes a number of proteins involved in biosynthesis of the heteropolymeric and Wzy-dependent B-band O antigen, including Wzy, the O-antigen polymerase, and Wzz, the regulator of O-antigen chain length. A gene (formerly wbpF), contiguous with wzy in the wbp cluster, is predicted to encode a highly hydrophobic protein with multiple membrane-spanning domains. This secondary structure is consistent with that of Wzx (RfbX), the putative O-antigen unit translocase or “flippase.” Insertion of a Gmr cassette at two separate sites within the putativewzx gene led in both cases to the loss of B-band lipopolysaccharide (LPS) O-antigen production. To our knowledge, this is the first report of the successful generation of chromosomalwzx gene replacement mutations. Surprisingly, inactivation of wzx also led to a marked delay in production of the ATP-binding cassette–transporter-dependent, d-rhamnose homopolymer, A-band LPS. This effect on A-band LPS synthesis was alleviated by supplying multiple copies of WbpL in trans. WbpL, a WecA (Rfe) homologue, was shown recently to be essential for the initiation of both A-band and B-band LPS synthesis in P. aeruginosa O5 (H. L. Rocchetta, L. L. Burrows, J. C. Pacan, and J. S. Lam, Mol. Microbiol. 28:1103–1119, 1998). These results suggest that the delay in A-band LPS production may arise from insufficient access to WbpL when the completed B-band O unit is not successfully translocated to the periplasm. Without adequate WbpL, A-band LPS synthesis is delayed. A subset of wzx mutants appeared to have accumulated second-site mutations which either restored the normal expression of A-band LPS or abolished A-band expression completely. Complementation studies showed that all of the additional mutations affecting LPS synthesis that were characterized in this study were located within the B-band LPS genes.
机译：铜绿假单胞菌 O5的 wbp 簇编码许多参与杂聚和依赖Wzy的B带O抗原生物合成的蛋白质，包括Wzy，O抗原聚合酶和Wzz，O抗原链长的调节剂。预测在 wbp 簇中与 wzy 相邻的基因（以前是 wbpF ）编码具有多个跨膜结构域的高度疏水蛋白。这种二级结构与Wzx（RfbX）（推定的O抗原单元转位酶或“脂肪酶”）的结构一致。在两种情况下，在假定的 wzx 基因内两个不同的位点插入Gm r 盒会导致B波段脂多糖（LPS）O抗原产生的损失。据我们所知，这是成功生成染色体 wzx 基因替代突变的第一份报告。出乎意料的是， wzx 的失活也导致了ATP结合盒依赖于转运蛋白的d-鼠李糖均聚物A波段LPS的生产显着延迟。通过在 trans 中提供WbpL的多个副本，减轻了对A波段LPS合成的影响。最近发现，WbpL是一种WecA（Rfe）同源物，对于在 P中启动A波段和B波段LPS合成至关重要。铜绿O5（H. L. Rocchetta，L. L. Burrows，J. C. Pacan和J. S. Lam，Mol。Microbiol。28：1103-1119，1998）。这些结果表明，当完整的B波段O单元未成功转移到周质时，A波段LPS产生的延迟可能是由于对WbpL的访问不足。没有足够的WbpL，A波段LPS合成会延迟。 wzx 突变体的一个子集似乎具有积累的第二位突变，这些突变要么恢复了A波段LPS的正常表达，要么完全废除了A波段的表达。补充研究表明，在这项研究中表征的所有其他影响LPS合成的突变都位于B波段LPS基因内。

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