An Excimer Clamp for Measuring Damaged‐Base Excision by the DNA Repair Enzyme NTH1

Jun Yong Woong; Wilson David L.; Kietrys Anna M.; Lotsof Elizabeth R.; Conlon Savannah G.; David Sheila S.; Kool Eric T.

摘要

Abstract >Direct measurement of DNA repair enzyme activities is important both for the basic study of cellular repair pathways as well as for potential new translational applications in their associated diseases. NTH1, a major glycosylase targeting oxidized pyrimidines, prevents mutations arising from this damage, and the regulation of NTH1 activity is important in resisting oxidative stress and in suppressing tumor formation. Herein, we describe a novel molecular strategy for the direct detection of damaged DNA base excision activity by a ratiometric fluorescence change. This strategy utilizes glycosylase‐induced excimer formation of pyrenes, and modified DNA probes, incorporating two pyrene deoxynucleotides and a damaged base, enable the direct, real‐time detection of NTH1 activity in?vitro and in cellular lysates. The probe design was also applied in screening for potential NTH1 inhibitors, leading to the identification of a new small‐molecule inhibitor with sub‐micromolar potency. </abstract> </span> <span class="z_kbtn z_kbtnclass hoverxs" style="display: none;">展开▼</span> </div> <div class="translation abstracttxt"> <span class="zhankaihshouqi fivelineshidden" id="abstract"> <span>机译：</span><Abstract Type =“Main”XML：Lang =“en”> <标题类型=“main”>抽象</ title> > DNA修复酶活性的直接测量对于细胞修复途径的基本研究非常重要，以及在其相关疾病中潜在的新翻译应用。 NTH1是靶向氧化嘧啶的主要糖基酶，防止来自这种损伤引起的突变，并且NTH1活性的调节对于抗氧化应激和抑制肿瘤形成是重要的。在此，我们描述了通过比例荧光变化直接检测受损DNA基础切除活性的新型分子策略。该策略利用糖基酶诱导的比蛋白的准分子形成，并改性DNA探针，掺入两种芘脱氧核苷酸和受损基础，使得体外和细胞裂解物中的NTH1活性直接实时检测NTH1活性。探针设计也用于筛选潜在的NTH1抑制剂，导致鉴定具有亚微摩尔效力的新的小分子抑制剂。</ p> </摘要> </span> <span class="z_kbtn z_kbtnclass hoverxs" style="display: none;">展开▼</span> </div> </div> <div class="record"> <h2 class="all_title" id="enpatent33" >著录项</h2> <ul> <li> <span class="lefttit">来源</span> <div style="width: 86%;vertical-align: text-top;display: inline-block;"> <a href='/journal-foreign-19011/'>《Angewandte Chemie》</a> <b style="margin: 0 2px;">|</b><span>2020年第19期</span><b style="margin: 0 2px;">|</b><span>共6页</span> </div> </li> <li> <div class="author"> <span class="lefttit">作者</span> <p id="fAuthorthree" class="threelineshidden zhankaihshouqi"> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Jun Yong Woong&option=202" target="_blank" rel="nofollow">Jun Yong Woong;</a> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Wilson David L.&option=202" target="_blank" rel="nofollow">Wilson David L.;</a> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Kietrys Anna M.&option=202" target="_blank" rel="nofollow">Kietrys Anna M.;</a> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Lotsof Elizabeth R.&option=202" target="_blank" rel="nofollow">Lotsof Elizabeth R.;</a> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Conlon Savannah G.&option=202" target="_blank" rel="nofollow">Conlon Savannah G.;</a> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=David Sheila S.&option=202" target="_blank" rel="nofollow">David Sheila S.;</a> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Kool Eric T.&option=202" target="_blank" rel="nofollow">Kool Eric T.;</a> </p> <span class="z_kbtnclass z_kbtnclassall hoverxs" id="zkzz" style="display: none;">展开▼</span> </div> </li> <li> <div style="display: flex;"> <span class="lefttit">作者单位</span> <div style="position: relative;margin-left: 3px;max-width: 639px;"> <div class="threelineshidden zhankaihshouqi" id="fOrgthree"> <p>Department of ChemistryStanford UniversityStanford CA 94305 USA;</p> <p>Department of ChemistryStanford UniversityStanford CA 94305 USA;</p> <p>Department of ChemistryStanford UniversityStanford CA 94305 USA;</p> <p>Department of ChemistryUniversity of California DavisDavis CA 95616 USA;</p> <p>Department of ChemistryUniversity of California DavisDavis CA 95616 USA;</p> <p>Department of ChemistryUniversity of California DavisDavis CA 95616 USA;</p> <p>Department of ChemistryStanford UniversityStanford CA 94305 USA;</p> </div> <span class="z_kbtnclass z_kbtnclassall hoverxs" id="zhdw" style="display: none;">展开▼</span> </div> </div> </li> <li > <span class="lefttit">收录信息</span> <span style="width: 86%;vertical-align: text-top;display: inline-block;"></span> </li> <li> <span class="lefttit">原文格式</span> <span>PDF</span> </li> <li> <span class="lefttit">正文语种</span> <span>eng</span> </li> <li> <span class="lefttit">中图分类</span> <span><a href="https://www.zhangqiaokeyan.com/clc/1188.html" title="应用化学">应用化学;</a></span> </li> <li class="antistop"> <span class="lefttit">关键词</span> <p style="width: 86%;vertical-align: text-top;"> <a style="color: #3E7FEB;" href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=base excision repair&option=203" rel="nofollow">base excision repair;</a> <a style="color: #3E7FEB;" href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=DNA damage&option=203" rel="nofollow">DNA damage;</a> <a style="color: #3E7FEB;" href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=fluorescent probes&option=203" rel="nofollow">fluorescent probes;</a> <a style="color: #3E7FEB;" href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=glycosylases&option=203" rel="nofollow">glycosylases;</a> <a style="color: #3E7FEB;" href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=inhibitors&option=203" rel="nofollow">inhibitors;</a> </p> <div class="translation"> 机译：基本切除修复;DNA损伤;荧光探针;糖基酶;抑制剂; </div> </li> </ul> </div> </div> <div class="literature cardcommon"> <div class="similarity "> <h3 class="all_title" id="enpatent66">相似文献</h3> <div class="similaritytab clearfix"> <ul> <li class="active" >外文文献</li> <li >中文文献</li> <li >专利</li> </ul> </div> <div class="similarity_details"> <ul > <li> <div> <b>1. </b><a class="enjiyixqcontent" 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