Heterologous expression of Neurospora crassa cbh1 gene in Pichia pastoris resulted in production of a neutral cellobiohydrolase I

Yang Jie; Deng Lu; Zhao Chen; Fang Hao

摘要

Abstract > The high production cost of cellulase is one of the limitations that hinder the commercialization of lignocellulose‐based biorefineries. As one of the important cellulases, Neurospora crassa cellulase is not so intensively investigated as T. reesei cellulase. In this study, the cbh1 gene (NCU07340) cloned from N. crassa was expressed in Pichia pastoris under the control of alcohol oxidase 1 (AOX1) promotor. Six transformants with the highest resistance to G418 were selected by two rounds of transformant screening, among which the most robust producer of the recombinant cellobiohydrolase I (CBHI) has an Avicelase activity of 0.22?U/mL. After fermentation optimization, it was improved to 0.30?U/mL. Interestingly, the optimal temperature and pH of the recombinant CBHI were 60°C and 7.2, respectively, and it was quite stable within the wide ranges of temperature and pH. This work is a good example for the future improvement and optimization of N. crassa cellulase. </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> > 纤维素酶的高产量是阻碍基于木质纤维素的生物料理商业化的限制之一。作为重要的纤维素酶之一， neuroospora crassa </ i> 纤维素酶不会被密集地研究 t。 Reesei </ I> 纤维素酶。在这项研究中， cbh1 </ i> 基因（NCU07340）克隆 n。 Crassa </ i> 表达了 Pichia Pastoris </ i> 在醇氧化酶1（AXOX1）促进剂的控制下。通过两轮转化体筛选选择具有最高抗G418抗性抗性的六种转化体，其中重组纤维二聚酶I（CBHI）的最稳健的生产者具有0.22μl/ ml的百分之二酶活性。发酵优化后，它得到改善至0.30？U / ml。有趣的是，重组CBHI的最佳温度和pH分别为60℃和7.2，并且在宽范围和pH范围内非常稳定。这项工作是未来改善和优化的一个很好的例子 n。 Crassa </ i> 纤维素酶。 </ 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-15005/'>《Biotechnology Progress》</a> <b style="margin: 0 2px;">|</b><span>2019年第3期</span><b style="margin: 0 2px;">|</b><span>共9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=Yang Jie&option=202" target="_blank" rel="nofollow">Yang Jie;</a> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Deng Lu&option=202" target="_blank" rel="nofollow">Deng Lu;</a> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Zhao Chen&option=202" target="_blank" rel="nofollow">Zhao Chen;</a> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Fang Hao&option=202" target="_blank" rel="nofollow">Fang Hao;</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>College of Life SciencesNorthwest A&</p> <p>F UniversityYangling Shaanxi China;</p> <p>College of Life SciencesNorthwest A&</p> <p>F UniversityYangling Shaanxi China;</p> <p>College of Life SciencesNorthwest A&</p> <p>F UniversityYangling Shaanxi China;</p> <p>College of Life SciencesNorthwest A&</p> <p>F UniversityYangling Shaanxi China;</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/15.html" title="生物科学">生物科学;</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=cellobiohydrolase I&option=203" rel="nofollow">cellobiohydrolase I;</a> <a style="color: #3E7FEB;" href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=cellulase&option=203" rel="nofollow">cellulase;</a> <a style="color: #3E7FEB;" href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=glycosylation&option=203" rel="nofollow">glycosylation;</a> <a style="color: #3E7FEB;" href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Neurospora crassa&option=203" rel="nofollow">Neurospora crassa;</a> <a style="color: #3E7FEB;" href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Pichia pastoris&option=203" rel="nofollow">Pichia pastoris;</a> </p> <div class="translation"> 机译：纤维素酶I;纤维素酶;糖基化;神经孢子喃狂;Pichia Pastoris; </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" href="/academic-journal-cn_acta-biochimica-biophysica-sinica_thesis/0201254943041.html">Improved heterologous gene expression in Trichoderma reesei by cellobiohydrolase I gene (cbh1) promoter optimization</a> <b>[J]</b> . <span> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Ti Liu&option=202" target="_blank" rel="nofollow" class="tuijian_auth tuijian_authcolor">Ti Liu,</a> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Tianhong Wang&option=202" target="_blank" rel="nofollow" class="tuijian_auth tuijian_authcolor">Tianhong Wang,</a> <a 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Comprehensive final terminal report of the overall activities for past 20 years</a> <b>[R] </b> . <span> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Dutta, S K&option=202" target="_blank" rel="nofollow" class="tuijian_auth tuijian_authcolor">Dutta, S K </a> <span>1988</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：粗糙脉孢菌细胞类型中的差异基因表达：核糖体RNa基因。过去20年总体活动的综合终端报告</span> </p> </li> </ul> <ul style="display: none;"> <li> <div> <b>1. </b><a class="enjiyixqcontent" href="/academic-journal-cn_chinese-higher-education-academic-abstracts-biology_thesis/0201255401968.html">Heterologous expression and characterization of man gene from Bacillus Subtilis in Pichia Pastoris</a> <b>[J]</b> <span> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=&option=202" target="_blank" rel="nofollow" class="tuijian_auth tuijian_authcolor"> . </a> <span> <a href="/journal-cn-5107/" target="_blank" rel="nofollow" class="tuijian_authcolor"> . 中国高等学校学术文摘·生物学 </a> </span> <span> . 2008</span><span>,第001期</span> </span> </div> </li> <li> <div> <b>2. </b><a class="enjiyixqcontent" href="/academic-conference-cn_meeting-9664_thesis/020222341931.html">Effective and stable porcine interferon-a production by Pichia pastoris fed-batch cultivation with multi-variables clustering and analysis</a> <b>[C]</b> <span> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Ruisong Yu&option=202" target="_blank" rel="nofollow" class="tuijian_auth tuijian_authcolor"> . Ruisong Yu</a> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Shijuan Dong&option=202" target="_blank" rel="nofollow" class="tuijian_auth tuijian_authcolor">,Shijuan Dong</a> <a href="/search.html?doctypes=4_5_6_1-0_4-0_1_2_3_7_9&sertext=Yumin Zhu&option=202" target="_blank" rel="nofollow" class="tuijian_auth tuijian_authcolor">,Yumin Zhu</a> <span> <a href="/conference-cn-9664/" target="_blank" rel="nofollow" class="tuijian_authcolor"> . 上海市畜牧兽医学会2011年学术年会 </a> <span> <span> . 2011</span> </span> </div> </li> </ul> <ul style="display: none;"> <li> <div> <b>1. </b><a class="enjiyixqcontent" href="/patent-detail/06130444445925.html">Method of heterologous gene expression in yeast Pichia pastoris The expression vectors and transformed microorganisms.</a> <b>[P]</b> . <span> 外国专利：  ES2180528T3 </span> <span> . 2003-02-16</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：酵母巴斯德毕赤酵母中异源基因表达的方法表达载体和转化的微生物。 </span> </p> </li> <li> <div> <b>2. </b><a class="enjiyixqcontent" href="/patent-detail/06130444654375.html">approach to the expression of heterologous genes in the yeast pichia pastoris, expression vectors and transformed microorganisms</a> <b>[P]</b> . <span> 外国专利：  DK0438200T3 </span> <span> . 2002-11-11</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：毕赤酵母中表达异源基因的方法，表达载体和转化微生物 </span> </p> </li> <li> <div> <b>3. </b><a class="enjiyixqcontent" href="/patent-detail/06130446361225.html">Method for the expression of heterologous genes in the yeast Pichia pastoris, expression vectors and transformed microorganisms</a> <b>[P]</b> . <span> 外国专利：  EP0438200B1 </span> <span> . 2002-07-17</span> </div> <p class="zwjiyix translation" style="max-width: initial;height: auto;word-break: 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