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首页> 外文期刊>Biotechnology and Bioengineering >ΔFlucs: Brighter Photinus pyralisPhotinus pyralis firefly luciferases identified by surveying consecutive single amino acid deletion mutations in a thermostable variant
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ΔFlucs: Brighter Photinus pyralisPhotinus pyralis firefly luciferases identified by surveying consecutive single amino acid deletion mutations in a thermostable variant

机译：Δflucs：更亮的 photinus pyralis 通过测量连续的单一氨基酸缺失突变在恒温变体中鉴定的photinus pyralis萤火虫酶

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Abstract <section xml:id="bit26451-sec-0001" numbered="no"> > The bright bioluminescence catalyzed by Photinus pyralis firefly luciferase (Fluc) enables a vast array of life science research such as bio imaging in live animals and sensitive in vitro diagnostics. The effectiveness of such applications is improved using engineered enzymes that to date have been constructed using amino acid substitutions. We describe ΔFlucs: consecutive single amino acid deletion mutants within six loop structures of the bright and thermostable ×11 Fluc. Deletion mutations are a promising avenue to explore new sequence and functional space and isolate novel mutant phenotypes. However, this method is often overlooked and to date there have been no surveys of the effects of consecutive single amino acid deletions in Fluc. We constructed a large semi‐rational ΔFluc library and isolated significantly brighter enzymes after finding ×11 Fluc activity was largely tolerant to deletions. Targeting an “omega‐loop” motif (T352‐G360) significantly enhanced activity, altered kinetics, reduced Km for D‐luciferin <sub>,</sub> altered emission colors, and altered substrate specificity for redshifted analog DL‐infraluciferin. Experimental and in silico analyses suggested remodeling of the Ω‐loop impacts on active site hydrophobicity to increase light yields. This work demonstrates the further potential of deletion mutations, which can generate useful Fluc mutants and broaden the palette of the biomedical and biotechnological bioluminescence enzyme toolbox. </section> </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 XMLNS =“http://www.wiley.com/namespaces/wiley”type =“main”xml：lang =“en”> <title type =“main”>抽象</ title> <section XML：ID =“bit26451-sec-0001”编号=“否”> >通过 photinus pyralis催化的亮生物发光</ i>萤火虫荧光素酶（Fluc）使得大量的生命科学研究，如生物成像动物和体外诊断敏感。使用迄今为止使用氨基酸取代构建的工程酶改善了这种应用的有效性。我们描述了Δflucs：连续的单个氨基酸缺失突变体在明亮和热稳定的×11 fluc的六个环结构内。删除突变是一个有希望的途径，用于探索新的序列和功能空间并分离新的突变表型。然而，这种方法通常被忽略，迄今为止，悬浮液中连续单氨基酸缺失的影响没有任何调查。我们构建了一个大型半标率ΔFluc库，并在发现×11烙丝活动后孤立明显更亮的酶在很大程度上耐缺失。靶向“Omega-Loop”主题（T352-G360）显着增强的活性，改变的动力学，D-Luciferin <sub>减少km，</ sub>改变的发光颜色，以及用于红移的模拟D1-Inflyucerin的改变的底物特异性。实验性和硅藻分析表明ω-环路对活性位点疏水性的影响，以增加光收率。该工作证明了缺失突变的进一步潜力，其可以产生有用的熔琥突变体并拓宽生物医学和生物技术生物发光酶工具箱的调色板。 </ p> </ section> </ abstract> </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-15017/'>《Biotechnology and Bioengineering》</a> <b style="margin: 0 2px;">|</b><span>2018年第1期</span><b style="margin: 0 2px;">|</b><span>共10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=Halliwell Lisa M.&option=202" target="_blank" rel="nofollow">Halliwell Lisa M.;</a> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Jathoul Amit P.&option=202" target="_blank" rel="nofollow">Jathoul Amit P.;</a> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Bate Jack P.&option=202" target="_blank" rel="nofollow">Bate Jack P.;</a> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Worthy Harley L.&option=202" target="_blank" rel="nofollow">Worthy Harley L.;</a> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Anderson James C.&option=202" target="_blank" rel="nofollow">Anderson James C.;</a> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Jones D. Dafydd&option=202" target="_blank" rel="nofollow">Jones D. Dafydd;</a> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Murray James A.H.&option=202" target="_blank" rel="nofollow">Murray James A.H.;</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>School of BiosciencesUniversity of CardiffCardiff UK;</p> <p>School of BiosciencesUniversity of CardiffCardiff UK;</p> <p>School of BiosciencesUniversity of CardiffCardiff UK;</p> <p>School of BiosciencesUniversity of CardiffCardiff UK;</p> <p>Department of ChemistryUniversity College LondonLondon UK;</p> <p>School of BiosciencesUniversity of CardiffCardiff UK;</p> <p>School of BiosciencesUniversity of CardiffCardiff UK;</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/180.html" title="生物工程学（生物技术）">生物工程学（生物技术）;</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=bioimaging&option=203" rel="nofollow">bioimaging;</a> <a style="color: #3E7FEB;" href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=bioluminescence&option=203" rel="nofollow">bioluminescence;</a> <a style="color: #3E7FEB;" href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=bioreporter&option=203" rel="nofollow">bioreporter;</a> <a style="color: #3E7FEB;" href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=deletion mutagenesis&option=203" rel="nofollow">deletion mutagenesis;</a> <a style="color: #3E7FEB;" href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=firefly luci ferase&option=203" rel="nofollow">firefly luci ferase;</a> <a style="color: #3E7FEB;" href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=infraluciferi&option=203" rel="nofollow">infraluciferi;</a> </p> <div class="translation"> 机译：生物分析;生物发光;Biooreporter;缺失诱变;萤火虫Luci Ferase;Inflyuciferi; </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> </ul> </div> <div class="similarity_details"> <ul > <li> <div> <b>1. </b><a class="enjiyixqcontent" href="/journal-foreign-detail/0704021882139.html">ΔFlucs: Brighter <i xmlns="http://www.wiley.com/namespaces/wiley">Photinus pyralisPhotinus pyralis firefly luciferases identified by surveying consecutive single amino acid deletion mutations in a thermostable variant</a> <b>[J]</b> . <span> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Halliwell Lisa M.&option=202" target="_blank" rel="nofollow" class="tuijian_auth tuijian_authcolor">Halliwell Lisa M.,</a> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Jathoul Amit P.&option=202" target="_blank" rel="nofollow" class="tuijian_auth tuijian_authcolor">Jathoul Amit P.,</a> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Bate Jack P.&option=202" target="_blank" rel="nofollow" class="tuijian_auth tuijian_authcolor">Bate Jack P.,</a> <a href="/journal-foreign-15017/" target="_blank" rel="nofollow" class="tuijian_authcolor">Biotechnology and Bioengineering .</a> <span>2018</span><span>,第1期</span> </span> </div> <p class="zwjiyix translation" style="max-width: initial;height: auto;word-break: break-all;white-space: initial;text-overflow: initial;overflow: initial;"> <span>机译：Δflucs：更亮的<i xmlns =“http://www.wiley.com/namespaces/wiley”> photinus pyralis </ i>通过测量连续的单一氨基酸缺失突变在恒温变体中鉴定的photinus pyralis萤火虫酶</span> </p> </li> <li> <div> <b>2. </b><a class="enjiyixqcontent" href="/journal-foreign-detail/0704024020378.html">Gas‐phase chemical ionization of 4‐alkyl branched‐chain carboxylic acids and 3‐methylindole using H <sub xmlns="http://www.wiley.com/namespaces/wiley">3</sub>3 O <sup xmlns="http://www.wiley.com/namespaces/wiley">++ , NO <sup xmlns="http://www.wiley.com/namespaces/wiley">++ , and O <sub xmlns="http://www.wiley.com/namespaces/wiley">2</sub>2 <sup xmlns="http://www.wiley.com/namespaces/wiley">++ ions</a> <b>[J]</b> . <span> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Castada Hardy Z.&option=202" target="_blank" rel="nofollow" class="tuijian_auth tuijian_authcolor">Castada Hardy Z.,</a> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Barringer Sheryl A.&option=202" target="_blank" rel="nofollow" class="tuijian_auth tuijian_authcolor">Barringer Sheryl A.,</a> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Wick Macdonald&option=202" target="_blank" rel="nofollow" class="tuijian_auth tuijian_authcolor">Wick Macdonald </a> <a href="/journal-foreign-26627/" target="_blank" rel="nofollow" class="tuijian_authcolor">Rapid Communications in Mass Spectrometry: RCM .</a> <span>2017</span><span>,第19期</span> </span> </div> <p class="zwjiyix translation" style="max-width: initial;height: auto;word-break: break-all;white-space: initial;text-overflow: initial;overflow: initial;"> <span>机译：使用H <sub xmlns =“http://www.wiley.com/namespaces/wiley”> 3 </ sub> 3 o <sup xmlns的4-烷基支链羧酸和3-甲基吲哚的气相化学电离 =“http://www.wiley.com/namespaces/wiley”> + </ sup> +，no <sup xmlns =“http://www.wiley.com/namespaces/wiley”> + </ sup> +，和O <sub xmlns =“http://www.wiley.com/namespaces/wiley”> 2 </ sub> 2 <sup xmlns =“http://www.wiley.com/namespaces/wiley”> + </ sup> +离子</span> </p> </li> <li> <div> <b>3. </b><a class="enjiyixqcontent" href="/journal-foreign-detail/0704025121631.html"><fi xmlns="http://www.wiley.com/namespaces/wiley"> <fc>HLA</fc> ‐B*40:01:45 </fi> <fc xmlns="http://www.wiley.com/namespaces/wiley">HLA</fc>HLA ‐B*40:01:45 , a novel variant of <fi xmlns="http://www.wiley.com/namespaces/wiley"> <fc>HLA</fc> ‐B*40:01 </fi> <fc xmlns="http://www.wiley.com/namespaces/wiley">HLA</fc>HLA ‐B*40:01 , discovered in a Taiwanese hematopoitic stem cell donor</a> <b>[J]</b> . <span> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Yang K. 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