Capsules are long-chain carbohydrate polymers that envelop the surfaces of many bacteria, protecting them from host immune responses. Capsule biosynthesis enzymes are potential drug targets and valuable biotechnological tools for generating vaccine antigens. Despite their importance, it remains unknown how structurally variable capsule polymers of Gram-negative pathogens are linked to the conserved glycolipid anchoring these virulence factors to the bacterial membrane. Using Actinobacillus pleuropneumoniae as an example, we demonstrate that CpsA and CpsC generate a poly(glycerol-3-phosphate) linker to connect the glycolipid with capsules containing poly(galactosylglycerol-phosphate) backbones. We reconstruct the entire capsule biosynthesis pathway in A. pleuropneumoniae serotypes 3 and 7, solve the X-ray crystal structure of the capsule polymerase CpsD, identify its tetratricopeptide repeat domain as essential for elongating poly(glycerol-3-phosphate) and show that CpsA and CpsC stimulate CpsD to produce longer polymers. We identify the CpsA and CpsC product as a wall teichoic acid homolog, demonstrating similarity between the biosynthesis of Gram-positive wall teichoic acid and Gram-negative capsules.

胶囊是长链碳水化合物聚合物，包裹着许多细菌的表面，保护它们免受宿主免疫反应的影响。胶囊生物合成酶是潜在的药物靶标和产生疫苗抗原的有价值的生物技术工具。尽管它们很重要，但仍不清楚革兰氏阴性病原体的结构可变的胶囊聚合物如何与将这些毒力因子锚定在细菌膜上的保守糖脂相关联。以胸膜肺炎放线杆菌为例，我们证明 CpsA 和 CpsC 生成聚（3-磷酸甘油）连接体，将糖脂与含有聚（半乳糖基甘油-磷酸）主链的胶囊连接起来。我们重建了胸膜肺炎放线菌血清型 3 和 7 的整个荚膜生物合成途径，解析了荚膜聚合酶 CpsD 的 X 射线晶体结构，确定其四三肽重复结构域对于延长聚（甘油-3-磷酸）至关重要，并表明 CpsA CpsC 刺激 CpsD 产生更长的聚合物。我们将 CpsA 和 CpsC 产物鉴定为壁磷壁酸同系物，证明革兰氏阳性壁磷壁酸和革兰氏阴性胶囊的生物合成之间的相似性。