基于新抗原受体可变域的赭曲霉毒素A竞争物与VHH抗体互作的亲和力调控机制

抗独特型纳米抗体Nb-Ab2β能竞争小分子危害物，在竞争免疫分析中具有良好的应用前景。当前Nb-Ab2β制备存在筛选效率低、抗体序列多样性差等问题，不利于从分子水平研究Nb-Ab2β与抗体的相互作用，而明确抗体与抗原互作的亲和力调控机制对于提高检测灵敏度至关重要。本课题拟以赭曲霉毒素A（OTA）为研究对象，通过构建噬菌体展示鲨源新抗原受体可变域（VNAR）天然库，以具有更高抗体可变区占比的羊驼纳米抗体（VHH）为靶标并优化淘选及洗脱策略，来提高OTA竞争物VNAR-Ab2β的筛选效率。基于VHH和VNAR结构简单、易基因改造的特性，综合运用同源建模、分子对接和定点饱和突变等技术，分析VHH与VNAR-Ab2β的识别模式，并探究两者互作的亲和力双向调控机制。研究能为高效筛选小分子危害物的新型竞争物VNAR-Ab2β及抗原抗体互作的亲和力双向调控提供模式依据，拓宽高灵敏竞争免疫分析的研究思路。

The anti-idiotope nanobody Nb-Ab2β can compete with small molecule hazards and has a good application prospect in the competitive immunoassay. Currently, there are some problems in the preparation of Nb-Ab2β, such as low screening efficiency and poor diversity of antibody sequences, which influence the study of the interaction between Nb-Ab2β and antibody at the molecular level. Meanwhile, clarifying the affinity regulation mechanism of antibody-antigen interaction is vital to improving the detection sensitivity. This project intends to select a cereal mycotoxin ochratoxin A as the model small molecule compound, construct the phage-displayed shark nanobody (VNAR) naive library, use the alpaca nanobody (VHH) to replace the conventional antibody as the target, and optimize the strategy of biopanning and elution for improving the screening efficiency of OTA competitor VNAR-Ab2β. Based on the characteristics of simple structure and ease of genetic modification of VHH and VNAR, homologous modeling, molecular docking, and site-saturated mutation are used in combination to analyze the recognition mode of VHH and VNAR-Ab2β, and explore the affinity bidirectional regulation mechanism of the interaction between them. This research can provide the model basis for the efficient screening of the novel competitor VNAR-Ab2β of small molecule compounds, and broaden the research ideas of the highly sensitive competitive immunoassay.