Institute of Microbiology and Hygiene, Charité-Universitätsmedizin Berlin, Dorotheenstrasse 96, 10117 Berlin, Germany.
Chemistry. 2010 Mar 15;16(11):3536-44. doi: 10.1002/chem.200902205.
Lyme disease (LD) is the most common tick-borne disease in Europe, North America, and Asia. The etiologic agents of LD are spirochetes of the group Borrelia burgdorferi sensu lato, which possess a lipid content of 25-30% of the dry weight. The major glycolipid cholesteryl 6-O-acyl-beta-D-galactopyranoside (ACGal), present in B. burgdorferi sensu stricto, B. afzelii, and B. garinii, is a specific and highly prevalent antigen frequently recognized by antibodies in late-stage LD. Here we report a convenient route for the chemical synthesis of ACGal by employing a combination of chemical synthesis steps with enzymatic transformations. This synthesized molecule was compared with bacterial extracts by immunoblots with patient sera, confirming the preserved antigenicity. Next, a glycolipid library derived from the native molecules with variations in the fatty acyl moiety and derivatives in which the cholesterol has been replaced was designed and synthesized. The chemical structures were confirmed by 1D and 2D NMR spectroscopy and mass spectrometry. The native and synthetic glycolipids were utilized in immunoblots to determine the epitope recognized by antibodies in patient sera. By this method we could demonstrate that galactose, cholesterol, and a fatty acid with a minimal chain length of four carbon atoms comprises the essential structure for recognition by antibodies. Finally, this finding allowed the synthesis of a functionalized ACGal with an omega-mercapto group at the fatty acid and a facile protection and deprotection strategy. This antigenic hapten can be conjugated to a carrier protein to effect immunization against Lyme disease.
莱姆病（LD）是欧洲、北美和亚洲最常见的蜱传疾病。LD 的病原体是疏螺旋体属伯氏疏螺旋体亚种，其干重的脂质含量为 25-30%。主要糖脂胆固醇 6-O-酰基-β-D-半乳糖吡喃糖苷（ACGal）存在于伯氏疏螺旋体亚种、阿氏疏螺旋体和伽氏疏螺旋体中，是一种特异性和高流行的抗原，经常被晚期 LD 患者的抗体识别。在这里，我们报告了一种通过化学合成步骤与酶转化相结合来合成 ACGal 的方便途径。通过与患者血清的免疫印迹比较，该合成分子与细菌提取物相比较，证实了其保留的抗原性。接下来，设计并合成了一个来源于天然分子的糖脂文库，其中脂肪酸部分存在变化，胆固醇被取代的衍生物也存在变化。通过 1D 和 2D NMR 光谱和质谱确认了化学结构。利用天然和合成糖脂进行免疫印迹，以确定患者血清中的抗体识别的表位。通过这种方法，我们可以证明半乳糖、胆固醇和具有最小四个碳原子链长的脂肪酸是抗体识别的必需结构。最后，这一发现允许合成带有脂肪酸末端巯基和简单保护和脱保护策略的功能化 ACGal。这种抗原半抗原可以与载体蛋白缀合，以针对莱姆病进行免疫接种。
