基于表位屏蔽理论的Co-60辐照结合糖基化降低OVA致敏性的分子机制

前期研究表明，辐照可以促进卵清蛋白（OVA）内部氨基酸的糖基化进而降低其致敏性，这与其过敏表位的屏蔽密切相关。为了探清其内在机制，以OVA和还原糖为研究对象，经Co-60辐照结合糖基化处理，采用食品分析、ELISA、免疫印迹和细胞学等方法分析其糖基化和致敏性的变化规律，制备低敏性OVA；运用质谱、光谱和同位素标记等方法研究其一级和高级结构的变化；通过氧-18标记、Orbitrap MS等技术对糖基化位点进行定性定量分析；借助抗体捕获OVA中的主要过敏组分，采用质谱、nano光谱、HDX等方法表征其结构和抗原结合位点。并与天然OVA对比，解析辐照条件-糖基化位点/程度-致敏性-过敏表位之间的相关性，从已知过敏表位、糖基化屏蔽位点/屏蔽程度和抗原结合位点三个角度揭示Co-60辐照结合糖基化降低OVA致敏性的分子机制，从而完善糖基化蛋白质的表位屏蔽理论，为新型脱敏技术的研发提供理论指导。

Our previous research showed that irradiation could promote the glycation of ovalbumin (OVA) and reduced its allergenicity, which was closely related to the shielding of glyation sites on allergic epitopes. In order explore its internal mechamism, OVA and reducing sugar were used as research objects to treat by Co-60 irradiation. The change of glycation and allergenicity were analyzed by the technology such as food analysis, ELISA, western blot and cytology, and obtained the hypoallergenic OVA sample. The changes of their primary and higher structures were studied by mass spectrometry, spectrum and isotope labeling methods. The glycation sites were qualitatively and quantitatively analyzed by oxygen-18 labeling and Orbitrap MS and other methods. The main allergic components in OVA were captured by antibodies, and their structures and antigen binding sites were characterized by mass spectrometry, nano-spectroscopy and HDX. The correlation among the changes of irradiation conditions-glycation sites/degree – allergenicity - allergic epitopes was analyzed. By comparing with natural OVA, the molecular mechanism of reducing OVA allergenicity by Co-60 irradiation combined with glycation was revealed from three aspects including in known allergic epitopes, glycation shielding sites/extent of shielding and antigen binding sites, so as to improve the protein glycation table of shielding theory, provide theoretical guidance for the development of new desensitization technology.