Laboratory of Experimental Immunotherapy, Department of Immunology, Stephan Angeloff Institute of Microbiology, Bulgarian Academy of Sciences, Sofia, Bulgaria.
Laboratory of Clinical Immunology, Department of Clinical Hematology, Sofiamed University Hospital, Sofia, Bulgaria.
Front Immunol. 2019 Dec 3;10:2796. doi: 10.3389/fimmu.2019.02796. eCollection 2019.
Specific antibody reactivities are routinely used as biomarkers, but the antibody repertoire reactivity (igome) profiles are still neglected. Here, we propose rationally designed peptide arrays as efficient probes for these system level biomarkers. Most IgM antibodies are characterized by few somatic mutations, polyspecificity, and physiological autoreactivity with housekeeping function. Previously, probing this repertoire with a set of immunodominant self-proteins provided a coarse analysis of the respective repertoire profiles. In contrast, here, we describe the generation of a peptide mimotope library that reflects the common IgM repertoire of 10,000 healthy donors. In addition, an appropriately sized subset of this quasi-complete mimotope library was further designed as a potential diagnostic tool. A 7-mer random peptide phage display library was panned on pooled human IgM. Next-generation sequencing of the selected phage yielded 224,087 sequences, which clustered in 790 sequence clusters. A set of 594 mimotopes, representative of the most significant sequence clusters, was shown to probe symmetrically the space of IgM reactivities in patients' sera. This set of mimotopes can be easily scaled including a greater proportion of the mimotope library. The trade-off between the array size and the resolution can be explored while preserving the symmetric sampling of the mimotope sequence and reactivity spaces. BLAST search of the non-redundant protein database with the mimotopes sequences yielded significantly more immunoglobulin J region hits than random peptides, indicating a considerable idiotypic connectivity of the targeted igome. The proof of principle predictors for random diagnoses was represented by profiles of mimotopes. The number of potential reactivity profiles that can be extracted from this library is estimated at more than 10. Thus, a quasi-complete IgM mimotope library and a scalable representative subset thereof are found to address very efficiently the dynamic diversity of the human public IgM repertoire, providing informationally dense and structurally interpretable IgM reactivity profiles.
特异性抗体反应通常被用作生物标志物，但抗体库反应（免疫组）谱仍然被忽视。在这里，我们提出了合理设计的肽阵列作为这些系统水平生物标志物的有效探针。大多数 IgM 抗体的特点是体细胞突变少、多特异性和与管家功能的生理性自身反应。以前，用一组免疫优势自身蛋白探测这个库提供了对各自库谱的粗略分析。相比之下，在这里，我们描述了生成一个反映 10000 名健康供体共同 IgM 免疫组的肽模拟文库。此外，还进一步设计了该准完整模拟文库的适当大小子集作为潜在的诊断工具。一个 7 -mer 随机肽噬菌体展示文库被用于人 IgM 的池化。对所选噬菌体的下一代测序产生了 224087 个序列，这些序列聚类为 790 个序列簇。一组 594 个模拟物，代表最重要的序列簇，被证明可以对称地探测患者血清中 IgM 反应的空间。这个模拟物集可以很容易地扩展，包括更大比例的模拟物库。在保留模拟物序列和反应性空间的对称采样的同时，可以探索阵列大小和分辨率之间的权衡。用模拟物序列对非冗余蛋白质数据库进行 BLAST 搜索，得到的免疫球蛋白 J 区命中数明显多于随机肽，表明靶向 igome 的免疫型连接相当可观。随机诊断的原理预测因子由模拟物的图谱表示。可以从这个库中提取的潜在反应性图谱的数量估计超过 10 个。因此，一个准完整的 IgM 模拟文库和一个可扩展的代表性子集被发现可以非常有效地解决人类公共 IgM 免疫组的动态多样性，提供信息密集且结构可解释的 IgM 反应性图谱。
