Development of a dehalogenase-based protein fusion tag capable of rapid, selective and covalent attachment to customizable ligands.

Current chemical genomics Pub Date : 2012-01-01 Epub Date: 2012-10-05 DOI:10.2174/1875397301206010055
Lance P Encell, Rachel Friedman Ohana, Kris Zimmerman, Paul Otto, Gediminas Vidugiris, Monika G Wood, Georgyi V Los, Mark G McDougall, Chad Zimprich, Natasha Karassina, Randall D Learish, Robin Hurst, James Hartnett, Sarah Wheeler, Pete Stecha, Jami English, Kate Zhao, Jacqui Mendez, Hélène A Benink, Nancy Murphy, Danette L Daniels, Michael R Slater, Marjeta Urh, Aldis Darzins, Dieter H Klaubert, Robert F Bulleit, Keith V Wood
{"title":"Development of a dehalogenase-based protein fusion tag capable of rapid, selective and covalent attachment to customizable ligands.","authors":"Lance P Encell,&nbsp;Rachel Friedman Ohana,&nbsp;Kris Zimmerman,&nbsp;Paul Otto,&nbsp;Gediminas Vidugiris,&nbsp;Monika G Wood,&nbsp;Georgyi V Los,&nbsp;Mark G McDougall,&nbsp;Chad Zimprich,&nbsp;Natasha Karassina,&nbsp;Randall D Learish,&nbsp;Robin Hurst,&nbsp;James Hartnett,&nbsp;Sarah Wheeler,&nbsp;Pete Stecha,&nbsp;Jami English,&nbsp;Kate Zhao,&nbsp;Jacqui Mendez,&nbsp;Hélène A Benink,&nbsp;Nancy Murphy,&nbsp;Danette L Daniels,&nbsp;Michael R Slater,&nbsp;Marjeta Urh,&nbsp;Aldis Darzins,&nbsp;Dieter H Klaubert,&nbsp;Robert F Bulleit,&nbsp;Keith V Wood","doi":"10.2174/1875397301206010055","DOIUrl":null,"url":null,"abstract":"<p><p>Our fundamental understanding of proteins and their biological significance has been enhanced by genetic fusion tags, as they provide a convenient method for introducing unique properties to proteins so that they can be examinedin isolation. Commonly used tags satisfy many of the requirements for applications relating to the detection and isolation of proteins from complex samples. However, their utility at low concentration becomes compromised if the binding affinity for a detection or capture reagent is not adequate to produce a stable interaction. Here, we describe HaloTag® (HT7), a genetic fusion tag based on a modified haloalkane dehalogenase designed and engineered to overcome the limitation of affinity tags by forming a high affinity, covalent attachment to a binding ligand. HT7 and its ligand have additional desirable features. The tag is relatively small, monomeric, and structurally compatible with fusion partners, while the ligand is specific, chemically simple, and amenable to modular synthetic design. Taken together, the design features and molecular evolution of HT7 have resulted in a superior alternative to common tags for the overexpression, detection, and isolation of target proteins.</p>","PeriodicalId":88232,"journal":{"name":"Current chemical genomics","volume":"6 ","pages":"55-71"},"PeriodicalIF":0.0000,"publicationDate":"2012-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.2174/1875397301206010055","citationCount":"95","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Current chemical genomics","FirstCategoryId":"1085","ListUrlMain":"https://doi.org/10.2174/1875397301206010055","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"2012/10/5 0:00:00","PubModel":"Epub","JCR":"","JCRName":"","Score":null,"Total":0}
引用次数: 95

Abstract

Our fundamental understanding of proteins and their biological significance has been enhanced by genetic fusion tags, as they provide a convenient method for introducing unique properties to proteins so that they can be examinedin isolation. Commonly used tags satisfy many of the requirements for applications relating to the detection and isolation of proteins from complex samples. However, their utility at low concentration becomes compromised if the binding affinity for a detection or capture reagent is not adequate to produce a stable interaction. Here, we describe HaloTag® (HT7), a genetic fusion tag based on a modified haloalkane dehalogenase designed and engineered to overcome the limitation of affinity tags by forming a high affinity, covalent attachment to a binding ligand. HT7 and its ligand have additional desirable features. The tag is relatively small, monomeric, and structurally compatible with fusion partners, while the ligand is specific, chemically simple, and amenable to modular synthetic design. Taken together, the design features and molecular evolution of HT7 have resulted in a superior alternative to common tags for the overexpression, detection, and isolation of target proteins.

Abstract Image

Abstract Image

Abstract Image

一种基于脱卤酶的蛋白融合标签的开发,能够快速、选择性和共价地附着到可定制的配体上。
遗传融合标签增强了我们对蛋白质及其生物学意义的基本理解,因为它们提供了一种方便的方法来引入蛋白质的独特特性,以便分离检测它们。常用的标签满足许多与复杂样品中蛋白质的检测和分离有关的应用要求。然而,如果检测或捕获试剂的结合亲和力不足以产生稳定的相互作用,则它们在低浓度下的效用就会受到损害。在这里,我们描述了HaloTag®(HT7),这是一种基于修饰的卤代烷脱卤酶的基因融合标签,通过与结合配体形成高亲和力的共价附着来克服亲和标签的局限性。HT7及其配体具有额外的理想特性。该标签相对较小,是单体的,并且与融合伙伴在结构上兼容,而配体是特异性的,化学上简单的,并且适合模块化合成设计。综上所述,HT7的设计特点和分子进化导致了HT7在过表达、检测和分离靶蛋白方面优于普通标签。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
求助全文
约1分钟内获得全文 求助全文
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