{"title":"带有烷氧基天冬氨酸的 SpyTag 肽,可根据 pH 值激活 SpyCatcher/Tag 系统。","authors":"Sonji Che, Hiroyuki Konno and Koki Makabe*, ","doi":"10.1021/acs.bioconjchem.4c00052","DOIUrl":null,"url":null,"abstract":"<p >The SpyCatcher/SpyTag system is a protein pair that forms a covalent isopeptide bond without an additional energy supply. The ability to connect fused proteins makes this system an attractive tool for several protein engineering applications. Conditional activation of the SpyCatcher/SpyTag complex formation further expands the use of this system. Here, we evaluated the pH activation of SpyTag using alkoxyaspartic acids in the isopeptide-forming residue. We found that a peptide with an ethoxy group can be activated by hydrolysis under high pH conditions. However, the hydrolysis induces isoaspartate (isoAsp) formation, which is confirmed by an isoAsp-inserted short peptide. We overcame this problem by changing the C-terminal side of the aspartic acid position to Pro, which does not form isoAsp under high pH conditions. The findings of this study provide fundamental knowledge of the synthetic construction of the modified SpyTag peptide.</p>","PeriodicalId":29,"journal":{"name":"Bioconjugate Chemistry Bioconjugate","volume":null,"pages":null},"PeriodicalIF":4.0000,"publicationDate":"2024-04-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"SpyTag Peptide with Alkoxyl Aspartic Acids for pH-Dependent Activation of the SpyCatcher/Tag System\",\"authors\":\"Sonji Che, Hiroyuki Konno and Koki Makabe*, \",\"doi\":\"10.1021/acs.bioconjchem.4c00052\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p >The SpyCatcher/SpyTag system is a protein pair that forms a covalent isopeptide bond without an additional energy supply. The ability to connect fused proteins makes this system an attractive tool for several protein engineering applications. Conditional activation of the SpyCatcher/SpyTag complex formation further expands the use of this system. Here, we evaluated the pH activation of SpyTag using alkoxyaspartic acids in the isopeptide-forming residue. We found that a peptide with an ethoxy group can be activated by hydrolysis under high pH conditions. However, the hydrolysis induces isoaspartate (isoAsp) formation, which is confirmed by an isoAsp-inserted short peptide. We overcame this problem by changing the C-terminal side of the aspartic acid position to Pro, which does not form isoAsp under high pH conditions. The findings of this study provide fundamental knowledge of the synthetic construction of the modified SpyTag peptide.</p>\",\"PeriodicalId\":29,\"journal\":{\"name\":\"Bioconjugate Chemistry Bioconjugate\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":4.0000,\"publicationDate\":\"2024-04-25\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Bioconjugate Chemistry Bioconjugate\",\"FirstCategoryId\":\"1\",\"ListUrlMain\":\"https://pubs.acs.org/doi/10.1021/acs.bioconjchem.4c00052\",\"RegionNum\":2,\"RegionCategory\":\"化学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q1\",\"JCRName\":\"BIOCHEMICAL RESEARCH METHODS\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Bioconjugate Chemistry Bioconjugate","FirstCategoryId":"1","ListUrlMain":"https://pubs.acs.org/doi/10.1021/acs.bioconjchem.4c00052","RegionNum":2,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"BIOCHEMICAL RESEARCH METHODS","Score":null,"Total":0}
引用次数: 0
摘要
SpyCatcher/SpyTag 系统是一种无需额外能量供应即可形成共价异肽键的蛋白质对。连接融合蛋白的能力使该系统成为多种蛋白质工程应用中极具吸引力的工具。有条件地激活 SpyCatcher/SpyTag 复合物的形成进一步扩大了该系统的用途。在这里,我们利用异肽形成残基中的烷氧基天冬氨酸对 SpyTag 的 pH 活化进行了评估。我们发现,在高 pH 条件下,带有乙氧基的肽可通过水解激活。然而,水解会诱发异天冬氨酸(isoAsp)的形成,这一点已通过插入异天冬氨酸的短肽得到证实。我们通过将 C 端侧的天冬氨酸位置改为 Pro 来克服这一问题,因为 Pro 在高 pH 条件下不会形成异天冬氨酸。这项研究的结果为合成构建修饰的 SpyTag 肽提供了基础知识。
SpyTag Peptide with Alkoxyl Aspartic Acids for pH-Dependent Activation of the SpyCatcher/Tag System
The SpyCatcher/SpyTag system is a protein pair that forms a covalent isopeptide bond without an additional energy supply. The ability to connect fused proteins makes this system an attractive tool for several protein engineering applications. Conditional activation of the SpyCatcher/SpyTag complex formation further expands the use of this system. Here, we evaluated the pH activation of SpyTag using alkoxyaspartic acids in the isopeptide-forming residue. We found that a peptide with an ethoxy group can be activated by hydrolysis under high pH conditions. However, the hydrolysis induces isoaspartate (isoAsp) formation, which is confirmed by an isoAsp-inserted short peptide. We overcame this problem by changing the C-terminal side of the aspartic acid position to Pro, which does not form isoAsp under high pH conditions. The findings of this study provide fundamental knowledge of the synthetic construction of the modified SpyTag peptide.
期刊介绍:
Bioconjugate Chemistry invites original contributions on all research at the interface between man-made and biological materials. The mission of the journal is to communicate to advances in fields including therapeutic delivery, imaging, bionanotechnology, and synthetic biology. Bioconjugate Chemistry is intended to provide a forum for presentation of research relevant to all aspects of bioconjugates, including the preparation, properties and applications of biomolecular conjugates.