Direct Electrochemistry of Phanerochaete chrysosporium Cellobiose Dehydrogenase Covalently Attached onto Gold Nanoparticle Modified Solid Gold Electrodes
Direct Electrochemistry of Phanerochaete chrysosporium Cellobiose Dehydrogenase Covalently Attached onto Gold Nanoparticle Modified Solid Gold Electrodes
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DOI：
10.1021/la3018858
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发表时间：
2012-07-24
期刊：
影响因子：
3.9
通讯作者：
Gorton, Lo
中科院分区：
文献类型：
--
作者：
Matsumura, Hirotoshi;Ortiz, Roberto;Gorton, Lo
在氧化还原酶和各种电极材料之间实现有效的电化学通信是生物电化学的主要挑战之一，对于开发电子应用具有重要意义。纤维二糖脱氢酶 (CDH) 是一种细胞外黄素细胞色素，由含有催化 FAD 的脱氢酶结构域 (DHCDH)、含有血红素 b 的细胞色素结构域 (CYTCDH) 和连接这两个结构域的柔性接头区域组成。提出了来自担子菌黄孢原毛平革菌 (PcCDH) 的 CDH 的高效直接电子转移 (DET)，该 CDH 共价连接到混合自组装单层 (SAM) 修饰的金纳米颗粒 (AuNP) 电极。使用的硫醇如下：4-氨基苯硫酚(4-ATP)、4-巯基苯甲酸(4-MBA)、4-巯基苯酚(4-MP)、11-巯基-1-十一烷胺(MUNH2)、11-巯基-1-十一烷酸(MUCOOH)和11-巯基-1-十一烷醇(MUOH)。使用戊二醛作为交联剂，在混合 SAM 中的 PcCDH 和 4-ATP 或 MUNH2 之间形成共价键。 PcCDH 的共价固定和表面覆盖通过表面等离子共振 (SPR) 得到证实。为了提高电流密度，将金纳米粒子浇铸在多晶金电极的顶部。对于所有固定化的 PcCDH 修饰的 AuNPs 电极，循环伏安法在没有底物（乳糖）的情况下表现出清晰的 CYTcmi 电化学响应，具有快速的电子转移（ET）速率，并且在 pH 4.50 时，形式电位评估为 +162 mV vs NHE。首次估计了 CDH 的标准 ET 速率常数 (k)，发现 4ATP/4-MBA、4-ATP/4-MP、MUNH2/MUCOOH 和 MUNH2/MUOH 修饰电极的标准 ET 速率常数 (k) 分别为 52.1、59.8、112 和 154 s(-1)。在所有混合 SAM 修饰的 AuNP 电极中，PcCDH 仅通过 CYTcDH 显示 DET。未发现 DHcDH 结构域和电极之间存在 DET 通讯。通过引入 AuNP，乳糖氧化的电流密度显着增加。 4-ATP/4-MBA修饰的AuNPs的电流密度高达30 pA cm(-2)，是4-ATP/4-MBA修饰的多晶金电极的数倍。这些结果提供了对共价固定在金电极上的CDH的基本电化学特性的深入了解，并促进了CDH在生物传感器、生物膜细胞和生物电催化方面的进一步应用。
Achieving efficient electrochemical communication between redox enzymes and various electrode materials is one of the main challenges in bioelectrochemistry and is of great importance for developing electronic applications. Cellobiose dehydrogenase (CDH) is an extracellular flavocytochrome composed of a catalytic FAD containing dehydrogenase domain (DHCDH), a heme b containing cytochrome domain (CYTCDH), and a flexible linker region connecting the two domains. Efficient direct electron transfer (DET) of CDH from the basidiomycete Phanerochaete chrysosporium (PcCDH) covalently attached to mixed self-assembled monolayer (SAM) modified gold nanoparticle (AuNP) electrode is presented. The thiols used were as follows: 4-aminothiophenol (4-ATP), 4-mercaptobenzoic acid (4-MBA), 4-mercaptophenol (4-MP), 11-mercapto-l-undecanamine (MUNH2), 11-mercapto-1-undecanoic acid (MUCOOH), and 11-mercapto-l-undecanol (MUOH). A covalent linkage between PcCDH and 4-ATP or MUNH2 in the mixed SAMs was formed using glutaraldehyde as cross-linker. The covalent immobilization and the surface coverage of PcCDH were confirmed with surface plasmon resonance (SPR). To improve current density, AuNPs were cast on the top of polycrystalline gold electrodes. For all the immobilized PcCDH modified AuNPs electrodes, cyclic voltammetry exhibited clear electrochemical responses of the CYTcmi with fast electron transfer (ET) rates in the absence of substrate (lactose), and the formal potential was evaluated to be +162 mV vs NHE at pH 4.50. The standard ET rate constant (k) was estimated for the first time for CDH and was found to be 52.1, 59.8, 112, and 154 s(-1) for 4ATP/4-MBA, 4-ATP/4-MP, MUNH2/MUCOOH, and MUNH2/MUOH modified electrodes, respectively. At all the mixed SAM modified AuNP electrodes, PcCDH showed DET only via the CYTcDH. No DET communication between the DHcDH domain and the electrode was found. The current density for lactose oxidation was remarkably increased by introduction of the AuNPs. The 4-ATP/4-MBA modified AuNPs exhibited a current density up to 30 pA cm(-2), which is times higher than that obtained for a 4-ATP/4-MBA modified polycrystalline gold electrode. The results provide insight into fundamental electrochemical properties of CDH covalently immobilized on gold electrodes and promote further applications of CDHs for biosensors, biofiiel cells, and bioelectrocatalysis.