Tarnovitski Natalia, Matthews Leslie J, Sui Jianhua, Gershoni Jonathan M, Marasco Wayne A
Department of Cell Research and Immunology, Tel Aviv University, George S. Wise Faculty of Life Science, Israel.
J Mol Biol. 2006 May 26;359(1):190-201. doi: 10.1016/j.jmb.2006.03.008. Epub 2006 Mar 22.
Rapid elucidation of neutralizing antibody epitopes on emerging viral pathogens like severe acute respiratory syndrome (SARS) coronavirus (CoV) or highly pathogenic avian influenza H5N1 virus is of great importance for rational design of vaccines against these viruses. Here we combined screening of phage display random peptide libraries with a unique computer algorithm "Mapitope" to identify the discontinuous epitope of 80R, a potent neutralizing human anti-SARS monoclonal antibody against the spike protein. Using two different types of random peptide libraries which display cysteine-constrained loops or linear 13-15-mer peptides, independent panels containing 42 and 18 peptides were isolated, respectively. These peptides, which had no apparent homologous motif within or between the peptide pools and spike protein, were deconvoluted into amino acid pairs (AAPs) by Mapitope and the statistically significant pairs (SSPs) were defined. Mapitope analysis of the peptides was first performed on a theoretical model of the spike and later on the genuine crystal structure. Three clusters (A, B and C) were predicted on both structures with remarkable overlap. Cluster A ranked the highest in the algorithm in both models and coincided well with the sites of spike protein that are in contact with the receptor, consistent with the observation that 80R functions as a potent entry inhibitor. This study demonstrates that by using this novel strategy one can rapidly predict and identify a neutralizing antibody epitope, even in the absence of the crystal structure of its target protein.
快速阐明新型病毒病原体（如严重急性呼吸综合征（SARS）冠状病毒（CoV）或高致病性禽流感H5N1病毒）上的中和抗体表位，对于合理设计针对这些病毒的疫苗至关重要。在此，我们将噬菌体展示随机肽库筛选与独特的计算机算法“Mapitope”相结合，以鉴定80R（一种针对刺突蛋白的强效中和型抗SARS人单克隆抗体）的不连续表位。使用展示半胱氨酸限制环或线性13 - 15肽的两种不同类型的随机肽库，分别分离出包含42个和18个肽的独立肽组。这些肽在肽库内部或之间以及与刺突蛋白之间没有明显的同源基序，通过Mapitope将其解卷积为氨基酸对（AAPs）并定义了具有统计学意义的对（SSPs）。肽的Mapitope分析首先在刺突的理论模型上进行，随后在真实晶体结构上进行。在两种结构上都预测到了三个簇（A、B和C），且有显著重叠。簇A在两个模型的算法中排名最高，并且与刺突蛋白中与受体接触的位点非常吻合，这与80R作为强效进入抑制剂的观察结果一致。这项研究表明，通过使用这种新策略，即使在其靶蛋白晶体结构缺失的情况下，也能快速预测和鉴定中和抗体表位。
