癌变·畸变·突变2026，Vol.38Issue(3)：197-204,8.DOI:10.3969/j.issn.1004-616x.2026.03.004
LCN2 K162琥珀酰化修饰对LCN2蛋白结构和功能变化的预测及特异识别该修饰抗体的制备
Prediction of structural and functional changes of lysine succinylation-induced LCN2 at position 162 and preparation of a specific antibody recognizing this modification
摘要
Abstract
OBJECTIVE:To investigate changes in structure and function of LCN2 protein which was induced by succinylation modification at lysine 162(K162),and to prepare a specific antibody targeting this modification,thereby laying an experimental foundation for subsequent detection of its expression and distribution at the cellular and tissue levels.METHODS:AlphaFold3 was used to predict and simulate the three-dimensional structures of wild-type LCN2 and LCN2 with succinylation at K162.Subsequently,visualization and comparative analysis were carried out to reveal impact of this modification on LCN2 structure and function.Three peptides containing K162 were designed and synthesized based on the LCN2 amino acid sequence:two peptides with succinylated K162,and one non-modified K162 peptide as a control.The two modified peptides were conjugated into keyhole limpet hemocyanin(KLH),and rabbits were immunized to prepare antibodies specifically recognizing LCN2 K162 succinylation.Antibody titer and specificity were evaluated by enzyme-linked immunosorbent assay(ELISA),Dot blot and Western blot.RESULTS:Bioinformatics analysis revealed significant differences in structure and function between succinylated LCN2 K162 and wild-type LCN2.Three-dimensional structure prediction identified four distinct non-overlapping regions between the two models.Succinylation at K162 drastically altered the internal hydrogen bonds and solvent-accessible surface area of LCN2,exerting a profound impact on its functional structure.Immunization of rabbits with the two modified peptides and one control peptide successfully yielded antibodies.ELISA results showed serum antibody titers of 1∶2.56×105,1∶2.56×105,and 1∶6.4×104,respectively.The titer of the modification-specific antibody was 243-fold higher than that of the control antibody,and the hybridization signal intensity was 34-fold stronger.Ab4,with the higher titer,was selected for subsequent experiments,as it specifically recognized the LCN2 K162 succinylation site.CONCLUSION:Bioinformatics analysis indicates that succinylation at K162 affected structure and function of LCN2 protein;the prepared antibody specifically recognized LCN2 K162 succinylation modification.关键词
脂质运载蛋白-2/赖氨酸琥珀酰化/肺腺癌/抗体特异性Key words
LCN2/lysine succinylation/lung adenocarcinoma/antibody specificity分类
医药卫生引用本文复制引用
陈湖婷,何柳燕,陈丽丽,余华军,张海涛,伍俊..LCN2 K162琥珀酰化修饰对LCN2蛋白结构和功能变化的预测及特异识别该修饰抗体的制备[J].癌变·畸变·突变,2026,38(3):197-204,8.基金项目
湛江市科技发展专项资金竞争性分配项目(2022A01197) （2022A01197）
