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首页> 外文期刊>ACS Omega >Engineered (Lys)6-Tagged Recombinant Sulfide-Reactive Hemoglobin I for Covalent Immobilization at Multiwalled Carbon Nanotubes
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Engineered (Lys)6-Tagged Recombinant Sulfide-Reactive Hemoglobin I for Covalent Immobilization at Multiwalled Carbon Nanotubes

机译：工程化的（Lys） 6 标记的重组硫化物反应性血红蛋白I，用于共价固定在多壁碳纳米管上

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摘要

The recombinant HbI was fused with a poly-Lys tag ((Lys)_(6)-tagged rHbI) for specific-site covalent immobilization on two carbon nanotube transducer surfaces, i.e., powder and vertically aligned carbon nanotubes. The immobilization was achieved by following two steps: (1) generation of amine-reactive ester from the carboxylic acid groups of the surfaces and (2) coupling these groups with the amine groups of the Lys-tag. We analyzed the immobilization process using different conditions and techniques to differentiate protein covalent attachment from physical adsorption. Fourier transform infrared microspectroscopy data showed a 14 cm~(–1) displacement of the protein’s amide I and amide II peaks to lower the frequency after immobilization. This result indicates a covalent attachment of the protein to the surface. Differences in the morphology of the carbon substrate with and without (Lys)_(6)-tagged rHbI confirmed protein immobilization, as observed by transmission electron microscopy. The electrochemical studies, which were performed to evaluate the redox center of the immobilized protein, show a confinement suitable for an efficient electron transfer system. More importantly, the electrochemical studies allowed determination of a redox potential for the new (Lys)_(6)-tagged rHbI. The data show that the protein is electrochemically active and retains its biological activity toward H_(2)S.
机译：重组HbI与poly-Lys标签（（Lys）_（6）标记的rHbI）融合在一起，用于将特定位点共价固定在两个碳纳米管换能器表面（即粉末和垂直排列的碳纳米管）上。通过以下两个步骤实现固定化：（1）从表面的羧酸基团生成胺反应性酯，以及（2）将这些基团与Lys-tag的胺基偶联。我们分析了使用不同条件和技术区分蛋白质共价附着与物理吸附的固定过程。傅立叶变换红外光谱数据表明，蛋白质的酰胺I和酰胺II峰的位移为14 cm〜（–1），以降低固定后的频率。该结果表明蛋白质共价附于表面。如通过透射电子显微镜观察到的那样，带有和不带有（Lys）_（6）标签的rHbI的碳底物形态上的差异证实了蛋白质的固定化。进行电化学研究以评估固定化蛋白质的氧化还原中心，结果表明该限制适用于有效的电子转移系统。更重要的是，电化学研究可以确定新的（Lys）_（6）标记的rHbI的氧化还原电位。数据表明该蛋白质具有电化学活性，并保留了其对H_（2）S的生物学活性。

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