Institute of Microbiology, ETH Zurich, Department of Biology, HCI F407, Wolfgang-Pauli-Str. 10, CH-8093 Zurich, Switzerland.
Carbohydr Res. 2013 Dec 15;382:58-64. doi: 10.1016/j.carres.2013.10.002. Epub 2013 Oct 10.
NMR spectroscopy can detect biomolecules like lipopolysaccharide directly on the surface of the cell, thus avoiding isolation and purification, and providing a more realistic description than the one derived from in vitro studies. Here we present a high-resolution magic-angle spinning NMR study of the O-antigen of Salmonella enterica serovar Typhimurium (S. Typhimurium) performed directly on the cells showing the alteration of its acetylation state over time. The O-antigen region of S. Typhimurium consists of the repeating unit [→2)-α-d-Manp-(1→4)-α-l-Rhap-(1→3)-α-d-Galp-(1→] where Man stands for mannose, Rha for rhamnose, and Gal for galactose. Man is substituted with abequose (Abe) O-acetylated at carbon 2. Our studies revealed that the appearance of de-O-acetylated O-antigen in the stationary growth phase is due to the de-O-acetylation of already synthesized O-acetylated O-antigen and that this reaction is caused by the metabolism-induced basic pH of the growth medium. The labile O-acetylation of the O-antigen we observed in S. Typhimurium generates non-stoichiometric O-acetylation states and therefore changes the nature of an immunogenic epitope.
NMR 光谱学可以直接在细胞表面检测生物分子，如脂多糖，从而避免了分离和纯化，并且提供了比体外研究更真实的描述。在这里，我们直接在细胞上进行了沙门氏菌肠炎亚种（S. Typhimurium）O-抗原的高分辨率魔角旋转 NMR 研究，显示了其乙酰化状态随时间的变化。S. Typhimurium 的 O-抗原区域由重复单元 [→2）-α-d-Manp-（1→4）-α-l-Rhap-（1→3）-α-d-Galp-（1→] 组成，其中 Man 代表甘露糖，Rha 代表鼠李糖，Gal 代表半乳糖。Man 被 abequose（Abe）取代，在 2 号碳上 O-乙酰化。我们的研究表明，在静止生长阶段出现去 O-乙酰化 O-抗原是由于已经合成的 O-乙酰化 O-抗原的去 O-乙酰化，并且该反应是由生长介质中代谢诱导的碱性 pH 值引起的。我们在 S. Typhimurium 中观察到的 O-抗原的不稳定 O-乙酰化产生了非化学计量的 O-乙酰化状态，因此改变了免疫原性表位的性质。
