The Development of Interface Molecules for Enzyme Electrochemical Reaction

酶电化学反应界面分子的研究进展

基本信息

批准号：
14350431
负责人：
SODE Koji
金额：
$ 9.09万
依托单位：
National University Corporation Tokyo University of Agriculture and Technology
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2002
资助国家：
日本
起止时间：
2002 至 2004
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-14350431/
关键词：
Enzyme reaction Catalyst chemical processes Cytochrome Nanobio Electron transfer Bioelectronics Biosensors

项目摘要

In most of oxidoreductases, the prosthetic group is buried deeply and within the protein shell. Thus a direct electrochemical recycling of the enzyme's prosthetic group at the electrode surface leading to a corresponding current signal is rarely encountered. In order to improve enzyme sensor signal property, we focused on the application of electron transfer proteins, cytochromes as the molecular interface between oxidorcductase redox center and electrode surface. In nature, various dehydrogenases harboring either PQQ or FAD are coupled with the respiratory chain via cytochromes. Based on this electron transfer property, we considered the utilization of cytochrome as the interface molecule to facilitate the electron transfer between enzyme and the artificial electron mediator or the electrode in the enzyme based biosensor. In this project, we focused on the application and engineering of cytochrome b_<562> as the interface molecule for novel bio-electronic devices and processes employi … More ng various oxidoreductases. The impact of the presence of cytochrome b_<562> on the various oxidoreductases was investigated, and its properties as the interface molecule were confirmed, by observing improved electrochemical response in the sensors employing these combinations. The engineering of cytochrome b_<562> was then attempted. The improvement of affinity toward PQQ glucose dehydrogenase was achieved by either modifying its surface charge or by introducing novel bio-affinity ligands. The employment of engineered interface molecules resulted in the bioelectrochemical reaction with PQQGDH, consequently improved the sensor properties. Finally, the co-expression of PQQ glucose dehydrogenase and cytochromeb_<562> was attempted to construct novel process employing interface molecules. The co-expression of PQQ glucose dehydrogenase and cytochromeb_<562> resulted in the construction of engineered E.coli cell which was able to transfer electron directly with electrode in the absence of artificial synthetic electron mediators. The results suggested the construction of novel bio-electrochemical process employing in vivo expressing interface molecules. Less

在大多数氧化还原酶中，辅基深埋在蛋白质外壳内，因此很少遇到酶辅基在电极表面直接电化学回收而产生相应的电流信号的情况。我们重点研究电子转移蛋白、细胞色素作为氧化还原酶氧化还原中心和电极表面之间的分子界面的应用。在自然界中，各种含有PQQ或FAD的脱氢酶与呼吸系统偶联。基于这种电子传递特性，我们考虑利用细胞色素作为界面分子来促进酶和基于酶的生物传感器中的人工电子介体或电极之间的电子传递。细胞色素 b_<562> 作为采用各种氧化还原酶的新型生物电子器件和工艺的界面分子的应用和工程细胞色素 b_<562> 的存在对研究了各种氧化还原酶，并通过观察使用这些组合的传感器中电化学响应的改善来确认它们作为界面分子的性质，然后尝试对细胞色素b_<562>进行改造，从而实现了对PQQ葡萄糖脱氢酶的亲和力的改善。通过改变其表面电荷或引入新型生物亲和配体，可以与 PQQGDH 发生生物电化学反应，从而改善传感器的性能。尝试使用界面分子共表达PQQ葡萄糖脱氢酶和细胞色素b_<562>构建新的过程PQQ葡萄糖脱氢酶和细胞色素b_<562>的共表达导致构建能够转移的工程化大肠杆菌细胞。在没有人工合成电子介体的情况下，电子直接与电极接触，结果表明采用体内表达界面分子构建新型生物电化学过程。