{"title":"Mechanistic Insights into How the Single Point Mutation Change the Autoantibody Repertoire","authors":"Zhong Ni, Fangyuan Song, Huimin Zhou, Ying Xu, Zhiguo Wang, Dongfeng Chen","doi":"10.1007/s10930-024-10225-w","DOIUrl":null,"url":null,"abstract":"<div><p>A recent study showed that just one point mutation F33 to Y in the complementarity-determining region 1 of heavy chain (H-CDR1) could lead to the auto-antibody losing its DNA binding ability. However, the potential molecular mechanisms have not been well elucidated. In this study, we investigated how the antibody lost the DNA binding ability caused by mutation F33 to Y in the H-CDR1. We found that the electrostatic force was not the primary driving force for the interaction between anti-DNA antibodies and the antigen single strand DNA (ssDNA), and that the H-CDR2 largely contributed to the binding of antigen ssDNA, even larger than H-CDR1. The H-F33Y mutation could increase the hydrogen-bond interaction but impair the pi-pi stacking interaction between the antibody and ssDNA. We further found that F33<sub>H</sub>, W98<sub>H</sub> and Y95<sub>L</sub> in the wiletype antibody could form the stable pi-pi stacking interaction with the nucleotide bases of ssDNA. However, the Y33 in mutant could not form the parallel sandwich pi-pi stacking interaction with the ssDNA. To further confirm the importance of pi-pi stacking, the wildtype antibody and the mutants (F33Y<sub>H</sub>, F33A<sub>H</sub>, W98A<sub>H</sub> and Y95A<sub>L</sub>) were experimentally expressed in CHO cells and purified, and the results from ELISA clearly showed that all the mutants lost the ssDNA binding ability. Taken together, our findings may not only deepen the understanding of the underlying interaction mechanism between autoantibody and antigen, but also broad implications in the field of antibody engineer.</p></div>","PeriodicalId":793,"journal":{"name":"The Protein Journal","volume":null,"pages":null},"PeriodicalIF":1.9000,"publicationDate":"2024-07-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"The Protein Journal","FirstCategoryId":"2","ListUrlMain":"https://link.springer.com/article/10.1007/s10930-024-10225-w","RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q4","JCRName":"BIOCHEMISTRY & MOLECULAR BIOLOGY","Score":null,"Total":0}
引用次数: 0
Abstract
A recent study showed that just one point mutation F33 to Y in the complementarity-determining region 1 of heavy chain (H-CDR1) could lead to the auto-antibody losing its DNA binding ability. However, the potential molecular mechanisms have not been well elucidated. In this study, we investigated how the antibody lost the DNA binding ability caused by mutation F33 to Y in the H-CDR1. We found that the electrostatic force was not the primary driving force for the interaction between anti-DNA antibodies and the antigen single strand DNA (ssDNA), and that the H-CDR2 largely contributed to the binding of antigen ssDNA, even larger than H-CDR1. The H-F33Y mutation could increase the hydrogen-bond interaction but impair the pi-pi stacking interaction between the antibody and ssDNA. We further found that F33H, W98H and Y95L in the wiletype antibody could form the stable pi-pi stacking interaction with the nucleotide bases of ssDNA. However, the Y33 in mutant could not form the parallel sandwich pi-pi stacking interaction with the ssDNA. To further confirm the importance of pi-pi stacking, the wildtype antibody and the mutants (F33YH, F33AH, W98AH and Y95AL) were experimentally expressed in CHO cells and purified, and the results from ELISA clearly showed that all the mutants lost the ssDNA binding ability. Taken together, our findings may not only deepen the understanding of the underlying interaction mechanism between autoantibody and antigen, but also broad implications in the field of antibody engineer.
期刊介绍:
The Protein Journal (formerly the Journal of Protein Chemistry) publishes original research work on all aspects of proteins and peptides. These include studies concerned with covalent or three-dimensional structure determination (X-ray, NMR, cryoEM, EPR/ESR, optical methods, etc.), computational aspects of protein structure and function, protein folding and misfolding, assembly, genetics, evolution, proteomics, molecular biology, protein engineering, protein nanotechnology, protein purification and analysis and peptide synthesis, as well as the elucidation and interpretation of the molecular bases of biological activities of proteins and peptides. We accept original research papers, reviews, mini-reviews, hypotheses, opinion papers, and letters to the editor.