Leiden Institute of Chemistry, Leiden University, 2300 RA Leiden, The Netherlands.
J Am Chem Soc. 2010 May 12;132(18):6550-7. doi: 10.1021/ja101515y.
A method for the electrical contacting of redox enzymes that obtain oxidizing or reducing equivalents from small electron-transfer proteins is demonstrated. The electrochemical contacting of redox enzymes through their immobilization onto electrode supports offers great potential for technological applications and for fundamental studies, but finding appropriate methods to immobilize the enzymes in an orientation allowing rapid electron transfer with the electrode has proven difficult. The copper enzyme nitrite reductase (NiR) and its natural electron-exchange partner pseudoazurin (Paz) are conjugated to a specific DNA tag and immobilized to a gold electrode into a stoichiometrically defined assembly. The DNA tethered to the electrode surface acts as flexible place-holder for the protein components, allowing both proteins to move within the construct. It is shown that Paz efficiently shuttles electrons between the electrode and the NiR enzyme, allowing the electrochemically driven NiR catalysis to be monitored. The activity of the NiR enzyme remains unperturbed by the immobilization. The rate-limiting step of the system is tentatively ascribed to the dissociation of the Paz/NiR complex. The electrochemical response of the system reports not only on the NiR catalysis and on interfacial electron transfer but also on the interaction between NiR and Paz.
一种从小型电子转移蛋白中获取氧化还原当量的氧化还原酶的电接触方法得到了证实。通过将氧化还原酶固定在电极支持物上进行电化学接触，为技术应用和基础研究提供了巨大的潜力，但找到合适的方法将酶以允许与电极快速电子转移的方向固定，事实证明是困难的。铜酶亚硝酸盐还原酶（NiR）及其天然电子交换伙伴假血蓝蛋白（Paz）与特定的 DNA 标签缀合，并固定在金电极上，形成化学计量定义的组装体。与电极表面相连的 DNA 充当蛋白质组件的柔性占位符，允许两种蛋白质在构建体内移动。结果表明，Paz 有效地在电极和 NiR 酶之间传递电子，从而可以监测电驱动的 NiR 催化作用。NiR 酶的活性不受固定化的影响。该系统的限速步骤暂时归因于 Paz/NiR 复合物的解离。该系统的电化学响应不仅报告了 NiR 催化作用和界面电子转移，还报告了 NiR 和 Paz 之间的相互作用。
