基于双臂型DNA四面体骨架的电子介体-辅酶-脱氢酶仿生传感界面的构建及应用

Dehydrogenases participate in the metabolism of many substances that involves the transfer of energy. The combination of electrochemical sensing methods to develop a dehydrogenase molecular device to simulate some important biochemical reactions in the body is one of the research hotspots in analytical chemistry and life sciences currently. However, the effective immobilization of coenzyme-dehydrogenase molecules and coenzyme regeneration at low potentials have limited its widespread development and application. This project proposed to use a two-arm tetrahedral DNA nanostructure as a skeleton to develop principles and methods for a controllable assembly of electron mediators on tetrahedral DNA nanostructures (TDNs) in order to construct an electron mediator-coenzyme-dehydrogenase accurate, fixed bionic sensing interface. This was used to study the electron transfer process between dehydrogenases, coenzymes, electron mediators, and electrodes that demonstrated the laws of how size and distribution density of the TDN, length and extension of the robotic arm, and type and embedding density of electron mediators affect the performance of the sensor. This demonstrates the advantages and mechanisms of the electron mediator/TDN in coenzyme regeneration at low potentials and accurate fixation of dehydrogenase-coenzyme. This study established the intrinsic relationship between material microstructure and macroscopic responses to finally develop a dehydrogenase electrode with strong resistance to interference, high sensitivity, and stability. The results are expected to provide a basis for the development of a new generation of high-performance biosensors and biomolecular devices.

脱氢酶参与生命体内的多种物质代谢和能量代谢过程，结合电化学传感手段构筑脱氢酶分子器件模拟研究生物体内某些重要的生化反应，是现代分析化学和生命科学领域的研究热点之一。然而辅酶-脱氢酶分子的有效固定和辅酶低电位下的再生限制了其广泛应用和发展。本项目拟以双臂型DNA四面体结构为骨架，发展电子介体在DNA四面体骨架上可控组装的原理和方法，构筑电子介体-辅酶-脱氢酶精准固定仿生传感界面。研究脱氢酶-辅酶-电子介体-电极之间的电子传递过程，揭示四面体的尺寸、分布密度、机械臂的长度和伸展方向、电子介体的种类和嵌入密度对传感器响应性能的影响规律，阐明电子介体/DNA四面体结构纳米材料在辅酶低电位下的再生和脱氢酶-辅酶精准固定方面的优势和机制；建立材料微观结构与宏观响应性能之间的内在关联，最终发展出抗干扰能力强、灵敏度高、稳定性好的脱氢酶电极，预期成果将为发展新一代高性能生物传感器件和生物分子器件提供依据。

发展高灵敏、高选择性和高稳定性的生物传感器和生物分子器件的研究是现代分析化学和生命科学的重要前沿领域之一，如何实现识别分子的精准高效固定以及如何提高检测的灵敏度是目前生物传感检测系统面临的两个难点。本项目针对分析方法中存在的瓶颈问题，基于电化学基本原理和生物化学传感技术，利用DNA四面体结构稳定，抗非特异性吸附能力强，探针设计灵活度高等优势，以生物识别分子为元件构筑纳米传感界面，发展了基于DNA纳米材料的传感新方法。项目筛选、设计、合成了不同尺寸的DNA四面体，发展了识别分子在DNA四面体骨架上可控组装的原理和方法，研究了其与识别探针的相互作用，揭示了四面体合成条件、四面体的尺寸、分布密度等条件对传感器响应性能的影响规律，阐述了四面体结构在识别分子精准固定方面的优势和机制，建立了材料微观结构与宏观响应性能之间的内在关联，为今后识别分子的精准固定提供理论和实验依据；合成了不同种类的碳基纳米材料，考察了不同材料对NADH的电化学响应，探讨了NADH在电极表面的响应机制，制备了脱氢酶传感器，研究了酶分子在材料表面的固定效率，掌握了纳米材料形状、尺寸等因素对传感器性能的影响规律，丰富了酶分析化学的研究内涵，为脱氢酶体系的构建提供新的机理和检测方法，具有重要的科学意义和实际应用价值。.共发表SCI论文5篇，申请发明专利1相；会议论文1篇；协助培养硕士2名；项目组成员参加学术会议1次。