CERMAV- Centre national de la recherche scientifique UPR5301 (affiliated to Université Joseph Fourier and ICMG), BP53, 38041 Grenoble, France ; Département de Chimie Moléculaire, UMR- Centre national de la recherche scientifique 5250 & ICMG FR 2607, Université Joseph Fourier, BP 53, 38041 Grenoble, France.
PLoS One. 2013 Aug 19;8(8):e71149. doi: 10.1371/journal.pone.0071149. eCollection 2013.
Recent advances in glycobiology revealed the essential role of lectins for deciphering the glycocode by specific recognition of carbohydrates. Integrated multiscale approaches are needed for characterizing lectin specificity: combining on one hand high-throughput analysis by glycan array experiments and systematic molecular docking of oligosaccharide libraries and on the other hand detailed analysis of the lectin/oligosaccharide interaction by x-ray crystallography, microcalorimetry and free energy calculations. The lectins LecB from Pseudomonas aeruginosa and BambL from Burkholderia ambifaria are part of the virulence factors used by the pathogenic bacteria to invade the targeted host. These two lectins are not related but both recognize fucosylated oligosaccharides such as the histo-blood group oligosaccharides of the ABH(O) and Lewis epitopes. The specificities were characterized using semi-quantitative data from glycan array and analyzed by molecular docking with the Glide software. Reliable prediction of protein/oligosaccharide structures could be obtained as validated by existing crystal structures of complexes. Additionally, the crystal structure of BambL/Lewis x was determined at 1.6 Å resolution, which confirms that Lewis x has to adopt a high-energy conformation so as to bind to this lectin. Free energies of binding were calculated using a procedure combining the Glide docking protocol followed by free energy rescoring with the Prime/Molecular Mechanics Generalized Born Surface Area (MM-GBSA) method. The calculated data were in reasonable agreement with experimental free energies of binding obtained by titration microcalorimetry. The established predictive protocol is proposed to rationalize large sets of data such as glycan arrays and to help in lead discovery projects based on such high throughput technology.
糖生物学的最新进展揭示了凝集素在通过特异性识别碳水化合物来破译糖码方面的重要作用。为了表征凝集素的特异性，需要采用综合多尺度方法：一方面结合糖芯片实验的高通量分析和聚糖文库的系统分子对接，另一方面结合 X 射线晶体学、微量热法和自由能计算对凝集素/寡糖相互作用进行详细分析。铜绿假单胞菌来源的 LecB 和鲍曼不动杆菌来源的 BambL 是致病菌用于入侵靶宿主的毒力因子的一部分。这两种凝集素没有相关性，但都识别岩藻糖基化寡糖，如 ABH(O)和 Lewis 表位的组织血型寡糖。使用糖芯片的半定量数据对特异性进行了表征，并通过 Glide 软件进行分子对接进行了分析。通过与现有复合物晶体结构的验证，可以获得可靠的蛋白质/寡糖结构预测。此外，还确定了 BambL/Lewis x 的晶体结构，分辨率为 1.6 Å，这证实了 Lewis x 必须采用高能构象才能与该凝集素结合。通过结合 Glide 对接方案和 Prime/分子力学广义 Born 表面积 (MM-GBSA) 方法进行自由能重评分的程序计算了结合自由能。计算数据与通过滴定微量热法获得的实验结合自由能吻合良好。所建立的预测方案旨在合理化糖芯片等大型数据集，并帮助基于这种高通量技术的先导发现项目。
