抗白三烯C4单链抗体Y54(L)W突变增加了对白三烯E4的亲和力

Y. Kawakami, Mai Kinoshita, Y. Mori, Shuji Okochi, Shiori Hirano, Ichika Shimoda, Keita Kanzaki, T. Suzuki-Yamamoto, M. Kimoto, M. Sugahara, T. Hori, H. Saino, M. Miyano, Shozo Yamamoto, Yoshitaka Takahashi
{"title":"抗白三烯C4单链抗体Y54(L)W突变增加了对白三烯E4的亲和力","authors":"Y. Kawakami, Mai Kinoshita, Y. Mori, Shuji Okochi, Shiori Hirano, Ichika Shimoda, Keita Kanzaki, T. Suzuki-Yamamoto, M. Kimoto, M. Sugahara, T. Hori, H. Saino, M. Miyano, Shozo Yamamoto, Yoshitaka Takahashi","doi":"10.1093/jb/mvw055","DOIUrl":null,"url":null,"abstract":"The X-ray crystal structure of an anti-leukotriene (LT) C4 monoclonal antibody (mAbLTC) in complex with LTC4 was determined, however, crystallographic studies alone are not enough to fully understand the structures of the antigen-binding site. To elucidate the individual contribution of Tyr-54 and Asn-58 in the light chain of mAbLTC, both of which formed a hydrogen bond with glutamic acid of LTC4, we examined whether substitution of the residues affects the antigen binding affinity and specificity using an anti-LTC4 single chain variable fragment (scFvLTC). Among the Tyr-54(L) mutants, Y54(L)W showed a dramatic increase in the affinity to LTE4 which was comparable to that to LTD4. Essentially the same results were obtained using the Y54(L)W mutant expressed in Escherichia coli and Pichia pastoris. The structural modeling suggested the formation of a novel hydrogen bond between the substituted tryptophan in the antibody and the cysteine residue in LTE4. The affinity of Y54(L)R, Y54(L)E and Y54(L)L to LTC4 was markedly reduced, whereas other tested Tyr-54(L) mutants as well as Asn-58(L) mutants did not show significant change in LT binding. The results may provide an insight into the molecular basis of specific LT recognition by the antibody.","PeriodicalId":22605,"journal":{"name":"The Journal of Biochemistry","volume":"52 1","pages":"79–86"},"PeriodicalIF":0.0000,"publicationDate":"2017-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"1","resultStr":"{\"title\":\"The Y54(L)W mutation of anti-leukotriene C4 single-chain antibody increases affinity to leukotriene E4\",\"authors\":\"Y. Kawakami, Mai Kinoshita, Y. Mori, Shuji Okochi, Shiori Hirano, Ichika Shimoda, Keita Kanzaki, T. Suzuki-Yamamoto, M. Kimoto, M. Sugahara, T. Hori, H. Saino, M. Miyano, Shozo Yamamoto, Yoshitaka Takahashi\",\"doi\":\"10.1093/jb/mvw055\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"The X-ray crystal structure of an anti-leukotriene (LT) C4 monoclonal antibody (mAbLTC) in complex with LTC4 was determined, however, crystallographic studies alone are not enough to fully understand the structures of the antigen-binding site. To elucidate the individual contribution of Tyr-54 and Asn-58 in the light chain of mAbLTC, both of which formed a hydrogen bond with glutamic acid of LTC4, we examined whether substitution of the residues affects the antigen binding affinity and specificity using an anti-LTC4 single chain variable fragment (scFvLTC). Among the Tyr-54(L) mutants, Y54(L)W showed a dramatic increase in the affinity to LTE4 which was comparable to that to LTD4. Essentially the same results were obtained using the Y54(L)W mutant expressed in Escherichia coli and Pichia pastoris. The structural modeling suggested the formation of a novel hydrogen bond between the substituted tryptophan in the antibody and the cysteine residue in LTE4. The affinity of Y54(L)R, Y54(L)E and Y54(L)L to LTC4 was markedly reduced, whereas other tested Tyr-54(L) mutants as well as Asn-58(L) mutants did not show significant change in LT binding. The results may provide an insight into the molecular basis of specific LT recognition by the antibody.\",\"PeriodicalId\":22605,\"journal\":{\"name\":\"The Journal of Biochemistry\",\"volume\":\"52 1\",\"pages\":\"79–86\"},\"PeriodicalIF\":0.0000,\"publicationDate\":\"2017-01-01\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"1\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"The Journal of Biochemistry\",\"FirstCategoryId\":\"1085\",\"ListUrlMain\":\"https://doi.org/10.1093/jb/mvw055\",\"RegionNum\":0,\"RegionCategory\":null,\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"\",\"JCRName\":\"\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"The Journal of Biochemistry","FirstCategoryId":"1085","ListUrlMain":"https://doi.org/10.1093/jb/mvw055","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"","JCRName":"","Score":null,"Total":0}
引用次数: 1

摘要

测定了一种与LTC4复合物的抗白三烯(LT) C4单克隆抗体(mAbLTC)的x射线晶体结构,但单靠晶体学研究不足以完全了解抗原结合位点的结构。为了阐明tyrr -54和Asn-58在与LTC4的谷氨酸形成氢键的mAbLTC轻链中的单独贡献,我们使用抗LTC4单链可变片段(scFvLTC)检测了残基的取代是否影响抗原结合亲和力和特异性。在Tyr-54(L)突变体中,Y54(L)W对LTE4的亲和力显著增加,与LTD4的亲和力相当。在大肠杆菌和毕赤酵母中表达的Y54(L)W突变体获得了基本相同的结果。结构模型表明在抗体中的取代色氨酸和LTE4中的半胱氨酸残基之间形成了一个新的氢键。Y54(L)R、Y54(L)E和Y54(L)L对LTC4的亲和力明显降低,而其他被试的Tyr-54(L)突变体和Asn-58(L)突变体对LT的结合没有明显变化。结果可能提供了一个深入了解特异性LT识别抗体的分子基础。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
The Y54(L)W mutation of anti-leukotriene C4 single-chain antibody increases affinity to leukotriene E4
The X-ray crystal structure of an anti-leukotriene (LT) C4 monoclonal antibody (mAbLTC) in complex with LTC4 was determined, however, crystallographic studies alone are not enough to fully understand the structures of the antigen-binding site. To elucidate the individual contribution of Tyr-54 and Asn-58 in the light chain of mAbLTC, both of which formed a hydrogen bond with glutamic acid of LTC4, we examined whether substitution of the residues affects the antigen binding affinity and specificity using an anti-LTC4 single chain variable fragment (scFvLTC). Among the Tyr-54(L) mutants, Y54(L)W showed a dramatic increase in the affinity to LTE4 which was comparable to that to LTD4. Essentially the same results were obtained using the Y54(L)W mutant expressed in Escherichia coli and Pichia pastoris. The structural modeling suggested the formation of a novel hydrogen bond between the substituted tryptophan in the antibody and the cysteine residue in LTE4. The affinity of Y54(L)R, Y54(L)E and Y54(L)L to LTC4 was markedly reduced, whereas other tested Tyr-54(L) mutants as well as Asn-58(L) mutants did not show significant change in LT binding. The results may provide an insight into the molecular basis of specific LT recognition by the antibody.
求助全文
通过发布文献求助,成功后即可免费获取论文全文。 去求助
来源期刊
自引率
0.00%
发文量
0
×
引用
GB/T 7714-2015
复制
MLA
复制
APA
复制
导出至
BibTeX EndNote RefMan NoteFirst NoteExpress
×
提示
您的信息不完整,为了账户安全,请先补充。
现在去补充
×
提示
您因"违规操作"
具体请查看互助需知
我知道了
×
提示
确定
请完成安全验证×
copy
已复制链接
快去分享给好友吧!
我知道了
右上角分享
点击右上角分享
0
联系我们:info@booksci.cn Book学术提供免费学术资源搜索服务,方便国内外学者检索中英文文献。致力于提供最便捷和优质的服务体验。 Copyright © 2023 布克学术 All rights reserved.
京ICP备2023020795号-1
ghs 京公网安备 11010802042870号
Book学术文献互助
Book学术文献互助群
群 号:481959085
Book学术官方微信