• 主题
高级搜索  >
历史搜索 清空历史
首页> 外文期刊>Bioprocess and Biosystems Engineering >Regulating unfolded protein response activator HAC1p for production of thermostable raw-starch hydrolyzing alpha-amylase in Pichia pastoris
【24h】

Regulating unfolded protein response activator HAC1p for production of thermostable raw-starch hydrolyzing alpha-amylase in Pichia pastoris

机译：调节未折叠的蛋白反应激活剂HAC1p在巴斯德毕赤酵母中生产热稳定的生淀粉水解α-淀粉酶

获取原文
获取原文并翻译 | 示例
           
页面导航
  • 摘要
  • 著录项
  • 相似文献
  • 相关主题

摘要

Unfolded protein response (UPR) usually happens when expressing heterologous proteins in high level, which may help cells to facilitate protein processing. Here, we evaluated the effects of the UPR activator HAC1p on a raw-starch hydrolyzing alpha-amylase (Gs4j-amyA), so as to improve heterologous production of the enzyme in Pichia pastoris. The gene (amyA) encoding Gs4j-amyA was first codon-optimized and expressed in P. pastoris under the control of the AOX1 promoter. A high gene dosage (12 copies) of amyA facilitated amylase expression which produced an enzyme activity of 305 U/ml. A spliced HAC1 encoding an UPR activator HAC1p was then co-expressed and the dosage effects of HAC1 on amylase expression was investigated. Six copies of HAC1 driven by AOX1 promoter produced a high amylase activity of 2200 U/ml, further increasing by 621%. However, excessive gene dosages driven by the same promoter led to a titration effect of its transcription factors and decreased the amount of amyA transcripts. Thus, constitutive expression of HAC1 by GAP promotor was further involved and Gs4j-amyA activity reached 3700 U/ml finally, which was further increased by 68.2%. Moreover, Gs4j-amyA was glycosylated in P. pastoris which generated higher enzyme activity than that in E. coli. Generally, regulating HAC1p expression by different strategies enhanced amylase production by 11.1 folds, indicating a reference for expression of other proteins in P. pastoris.
机译：高水平表达异源蛋白质时通常会发生未折叠的蛋白质反应（UPR），这可能有助于细胞促进蛋白质加工。在这里，我们评估了UPR活化剂HAC1p对生淀粉水解α-淀粉酶（Gs4j-amyA）的影响，从而提高了毕赤酵母中该酶的异源生产。编码Gs4j-amyA的基因（amyA）首先经过密码子优化，并在AOX1启动子的控制下在巴斯德毕赤酵母中表达。 amyA的高基因剂量（12个拷贝）促进了淀粉酶的表达，产生了305 U / ml的酶活性。然后共表达编码UPR激活剂HAC1p的剪接的HAC1，并研究HAC1对淀粉酶表达的剂量影响。由AOX1启动子驱动的六份HAC1产生了2200 U / ml的高淀粉酶活性，进一步增加了621％。然而，由相同启动子驱动的过量基因剂量导致其转录因子的滴定作用并减少了amyA转录物的量。因此，GAP启动子进一步参与了HAC1的组成型表达，Gs4j-amyA活性最终达到3700 U / ml，进一步增加了68.2％。而且，Gs4j-amyA在巴斯德毕赤酵母中被糖基化，其产生的酶活性高于大肠杆菌。通常，通过不同策略调节HAC1p表达可将淀粉酶产量提高11.1倍，这为在巴斯德毕赤酵母中表达其他蛋白质提供了参考。

著录项

  • 来源
    《Bioprocess and Biosystems Engineering》 |2017年第3期|341-350|共10页
  • 作者

    Huang Mengmeng; Gao Yanyun; Zhou Xiangshan; Zhang Yuanxing; Cai Menghao;

  • 作者单位

    East China Univ Sci & Technol, State Key Lab Bioreactor Engn, 130 Meilong Rd, Shanghai 200237, Peoples R China;

    East China Univ Sci & Technol, State Key Lab Bioreactor Engn, 130 Meilong Rd, Shanghai 200237, Peoples R China;

    East China Univ Sci & Technol, State Key Lab Bioreactor Engn, 130 Meilong Rd, Shanghai 200237, Peoples R China;

    East China Univ Sci & Technol, State Key Lab Bioreactor Engn, 130 Meilong Rd, Shanghai 200237, Peoples R China|Shanghai Collaborat Innovat Ctr Biomfg, 130 Meilong Rd, Shanghai 200237, Peoples R China;

    East China Univ Sci & Technol, State Key Lab Bioreactor Engn, 130 Meilong Rd, Shanghai 200237, Peoples R China;

  • 收录信息
  • 原文格式 PDF
  • 正文语种 eng
  • 中图分类
  • 关键词

    Unfolded protein response; HAC1p; Heterologous expression; P. pastoris; Raw-starch hydrolyzing alpha-amylase;

    机译：未折叠蛋白反应;HAC1p;异源表达;P。淀粉酶水解α-淀粉酶;

相似文献

  • 外文文献
  • 中文文献
  • 专利
获取原文

客服邮箱：kefu@zhangqiaokeyan.com

京公网安备：11010802029741号 ICP备案号：京ICP备15016152号-6 六维联合信息科技 (北京) 有限公司©版权所有

  • 客服微信

  • 服务号