Mediated spectroelectrochemical titration of proteins for redox potential measurements by a separator-less one-compartment bulk electrolysis method.
Mediated spectroelectrochemical titration of proteins for redox potential measurements by a separator-less one-compartment bulk electrolysis method.
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DOI：
10.1016/j.ab.2004.11.017
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发表时间：
2005-02
影响因子：
2.9
通讯作者：
S. Tsujimura;Atsushi Kuriyama;N. Fujieda;K. Kano;T. Ikeda
中科院分区：
文献类型：
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作者：
S. Tsujimura;Atsushi Kuriyama;N. Fujieda;K. Kano;T. Ikeda
单室本体电解和同时光谱测量是在一个传统的光谱比色皿中实现的，没有分离器，通过使用具有极大表面积的网状工作电极和具有非常小表面积的线状对电极。分光光度监测显示完全电解的一阶动力学。将该技术应用于细胞色素c和胆红素氧化酶的介导滴定，测定它们的氧化还原电位。描述了蛋白质和介体之间溶液氧化还原反应的动力学。由于重现性高，减去介体吸附引起的光谱背景非常容易。实验可以在完全厌氧的条件下进行。低吸光度蛋白质样品（低浓度或小吸收系数）和疏水蛋白质（如膜结合蛋白质）可用于测量。
One-compartment bulk electrolysis and simultaneous spectroscopic measurements are realized in a conventional spectroscopic cuvette without separator by using a mesh-type working electrode with extremely large surface area and a wire-type counter electrode with very small surface area. Spectrophotometric monitoring revealed complete electrolysis in a first-order kinetics. This technique was applied to mediated titration of cytochrome c and bilirubin oxidase for determining their redox potentials. Kinetics for the solution redox reaction between protein and mediator is described. The subtraction of spectral background due to mediator adsorption is very easy because of high reproducibility. The experiments can be done under completely anaerobic conditions. Low-absorbance protein samples (of low concentrations or small absorption coefficients) and hydrophobic proteins (such as membrane-bound proteins) are acceptable for measurements.