Gómara María José, García-Moreno Cristina, Bárcenas Oriol, Castellanos-Moreira Raúl, Sarmiento Juan Camilo, Crehuet Ramon, Pérez Yolanda, Sanmartí Raimon, Haro Isabel
Unit of Synthesis and Biomedical Applications of Peptides, IQAC-CSIC, Jordi Girona 18-26, 08034 Barcelona, Spain.
Computational and Theoretical Chemistry Group, IQAC-CSIC, Jordi Girona 18-26, 08034 Barcelona, Spain.
Int J Mol Sci. 2025 Sep 16;26(18):9026. doi: 10.3390/ijms26189026.
Given the limited knowledge of the effect of post-translational modifications (PTMs) on protein structure, in this study we investigated whether introducing one-to-three RA-related PTMs into the α-fibrin (617-631) peptide influences the conformation and structure of the peptide antigen that could be responsible for the autoantibody recognition. Ten peptides containing a different number of PTMs within their primary structure were synthesized and their recognition by sera from RA patients was analyzed. The conformation of the peptides was studied by circular dichroism (CD) and the structure of the most relevant antigenic peptides was determined by nuclear magnetic resonance (NMR) and enhanced-sampling molecular dynamics (MD). Although peptides containing citrulline (Cit) showed a higher degree of binding to AMPAs than peptides containing only homocitrulline and/or acetyl-lysine, the latter were able to bind to AMPAs in sera that showed a small response to peptides with Cit, with the response being different depending on the position of each PTM. CD and NMR analyses indicated a series of half-turn conformations in the Lys620-Arg630 region. MD simulations generated a set of conformations compatible with the NMR NOEs. The effect of the PTMs was observed in intra-molecular contacts, hydrogen bonds and van der Waals interactions, generating more collapsed conformations. Differences in autoantibody reactivity between peptides bearing different PTMs within their primary structures are noted. Peptides with PTMs adopt different conformations than unmodified peptides, probably due to the lower net charge of peptides with multiple PTMs, which may explain their recognition by autoantibodies.
鉴于对翻译后修饰（PTM）对蛋白质结构影响的了解有限，在本研究中，我们调查了将一至三种与类风湿关节炎（RA）相关的PTM引入α-纤维蛋白（617 - 631）肽中是否会影响可能负责自身抗体识别的肽抗原的构象和结构。合成了十种在其一级结构中含有不同数量PTM的肽，并分析了RA患者血清对它们的识别情况。通过圆二色性（CD）研究肽的构象，并通过核磁共振（NMR）和增强采样分子动力学（MD）确定最相关抗原肽的结构。尽管含有瓜氨酸（Cit）的肽比仅含有高瓜氨酸和/或乙酰赖氨酸的肽与抗瓜氨酸化蛋白抗体（AMPA）的结合程度更高，但后者能够与对含有Cit的肽反应较小的血清中的AMPA结合，反应因每个PTM的位置而异。CD和NMR分析表明在Lys620 - Arg630区域存在一系列半圈构象。MD模拟生成了一组与NMR核Overhauser效应（NOE）兼容的构象。在分子内接触、氢键和范德华相互作用中观察到PTM的作用，产生了更多折叠的构象。注意到在其一级结构中带有不同PTM的肽之间自身抗体反应性的差异。带有PTM的肽采用与未修饰肽不同的构象，这可能是由于具有多个PTM的肽的净电荷较低，这可能解释了它们被自身抗体识别的原因。
